Effects of thimerosal on NGF signal transduction and cell death in neuroblastoma cells.

Signaling through neurotrophic receptors is necessary for differentiation and survival of the developing nervous system. The present study examined the effects of the organic mercury compound thimerosal on nerve growth factor signal transduction and cell death in a human neuroblastoma cell line (SH-SY5Y cells). Following exposure to 100 ng/ml NGF and increasing concentrations of thimerosal (1 nM-10 microM), we measured the activation of TrkA, MAPK, and PKC-delta. In controls, the activation of TrkA MAPK and PKC-delta peaked after 5 min of exposure to NGF and then decreased but was still detectable at 60 min. Concurrent exposure to increasing concentrations of thimerosal and NGF for 5 min resulted in a concentration-dependent decrease in TrkA and MAPK phosphorylation, which was evident at 50 nM for TrkA and 100 nM for MAPK. Cell viability was assessed by the LDH assay. Following 24-h exposure to increasing concentrations of thimerosal, the EC50 for cell death in the presence or absence of NGF was 596 nM and 38.7 nM, respectively. Following 48-h exposure to increasing concentrations of thimerosal, the EC50 for cell death in the presence and absence of NGF was 105 nM and 4.35 nM, respectively. This suggests that NGF provides protection against thimerosal cytotoxicity. To determine if apoptotic versus necrotic cell death was occurring, oligonucleosomal fragmented DNA was quantified by ELISA. Control levels of fragmented DNA were similar in both the presence and absence of NGF. With and without NGF, thimerosal caused elevated levels of fragmented DNA appearing at 0.01 microM (apoptosis) to decrease at concentrations >1 microM (necrosis). These data demonstrate that thimerosal could alter NGF-induced signaling in neurotrophin-treated cells at concentrations lower than those responsible for cell death.     

Evaluation of combined fibroblast growth factor-2 and moderate hypothermia therapy in traumatically brain injured rats

Both the exogenous administration of fibroblast growth factor-2 (FGF-2) or the induction of moderate hypothermia have been shown to attenuate histopathology and improve functional outcome after traumatic brain injury (TBI). Since combined therapeutic strategies may be more beneficial than single therapies, we examined the potential synergistic effect of FGF-2 combined with moderate hypothermia treatment induced 10 min after TBI on functional and histological outcome following controlled cortical impact (CCI) injury. Fifty male Sprague-Dawley rats were randomized to one sham and four CCI treatment groups: Sham+vehicle (VEH); FGF-2 (45 microg/kg/h for 3 h i.v.)+Normothermia (37+/-0.5 degrees C); FGF-2+Hypothermia (32+/-0.5 degrees C for 3 h); VEH+Norm; VEH+Hypo. Vestibulomotor performance on the beam balance and beam-walk (BW) tasks on post-operative days 1-5 and spatial memory acquisition in the Morris water maze (MWM) on days 14-18 were assessed. After 4 weeks survival, histological evaluations (CA(1) and CA(3) cell counts and lesion volume) were performed. MWM performance improved in all treatment groups, but combined treatment was not more efficacious than either alone. The FGF-2+Hypo group performed significantly better than the other injured treatment groups in the BW task. Lastly, no significant group differences in beam balance or histological outcome were observed. These data suggest a suboptimal and incomplete synergy of combined FGF-2 and hypothermia treatment. These data may indicate that either our dose of FGF-2 or combination therapy was not optimized in our model.     

Increased susceptibility to vancomycin-resistant Enterococcus intestinal colonization persists after completion of anti-anaerobic antibiotic treatment in mice.

BACKGROUND: Antibiotic-associated disruption of the indigenous intestinal microflora may persist beyond the treatment period. Although piperacillin/tazobactam inhibits the establishment of vancomycin-resistant Enterococcus (VRE) stool colonization in mice during treatment, we hypothesized that this agent and other anti-anaerobic antibiotics would increase susceptibility to colonization during the period of recovery of the intestinal microflora. DESIGN: Mice received 10(4) colony-forming units of vancomycin-resistant E. faecium by orogastric inoculation 2, 5, or 10 days after completing 5 days of subcutaneous antibiotic treatment, or both during and 2 days after the completion of treatment. Denaturing gradient gel electrophoresis (DGGE) was performed to assess changes in the intestinal microflora. RESULTS: Anti-anaerobic antibiotics (ie, piperacillin/ tazobactam, cefoxitin, and clindamycin) caused significant disruption of the indigenous microflora (mean DGGE similarity indices < or = 27% in comparison with saline controls) and promoted the establishment of high-density colonization when VRE was inoculated 2 or 5, but not 10, days following treatment (P < .001). Piperacillin/tazobactam exhibited a biphasic effect on the establishment of colonization (ie, inhibition when exposed to VRE during treatment and promotion when exposed to VRE after discontinuation of treatment), resulting in greater overall promotion of colonization than did agents with minimal anti-anaerobic activity (ie, levofloxacin, cefepime, and aztreonam) when VRE was inoculated both during and 2 days after treatment (P < .001). CONCLUSION: Patients receiving anti-anaerobic antibiotics, including piperacillin/tazobactam, may be susceptible to the establishment of high-density VRE colonization during the period of recovery of the anaerobic microflora.     

A placebo controlled, double-blind study of mecamylamine treatment for cocaine dependence in patients enrolled in an opiate replacement program.

A placebo controlled, double-blind trial of mecamylamine treatment of cocaine dependence was performed in methadone or LAAM maintained subjects who met DSM-IV criteria for cocaine dependence. After an eight-week placebo run-in screening period, 35 subjects were randomly assigned to receive either mecamylamine (6 mg/day) or placebo transdermal patches for a 16-week treatment period. Outcome measures included quantitative urine benzoylecognine (BE) levels, self-report of cocaine use, cocaine craving, global impression scores, mood, retention, and safety. Mecamylamine was well tolerated, and study retention did not differ by treatment group. Evidence for cocaine use, based on urine BE levels and cocaine abstinence rates, did not differ by treatment group. Self reported cocaine use, cocaine craving, and global impression scores showed moderate improvement in both groups, with a significantly greater reduction in cocaine craving (p < 0.05) and self-rated severity of cocaine dependence (p < 0.05) in the placebo group. This pilot study does not support the effectiveness of mecamylamine for the treatment of cocaine dependence in methadone or LAAM maintained patients.     

Epoxide hydrolases in the rat epididymis: possible roles in xenobiotic and endogenous fatty acid metabolism.

Epoxide hydrolases play an important role in detoxifying epoxides that arise from the metabolism of xenobiotic and endogenous compounds. Both the soluble and microsomal forms of epoxide hydrolase (sEH and mEH, respectively) have been detected in the rat testis. Because of the important role the epididymis plays in sperm maturation and protection, the present study evaluated the presence and activity of these two epoxide hydrolases in the rat epididymis. Using Western blotting, protein bands consistent in size with both mEH and sEH were detected in the caput, corpus, and cauda of the epididymis. The mEH immunoreactive bands in the epididymis ( approximately 50 kDa) were consistent with mEH detected in the liver and kidney. The sEH immunoreactive bands in the epididymis ( approximately 65 kDa) were consistent with a recombinant sEH standard and sEH detected in the liver, kidney, and testis. The presence of mEH and sEH in the epididymis was supported by observations from substrate-based enzyme assays. Results indicated that epididymal mEH can hydrolyze [(3)H]-cis-stilbene oxide to the corresponding diol at levels approximately 9% of the kidney. Epididymal sEH hydrolyzed the substrate [(3)H]-trans-diphenylpropene oxide to the corresponding diol and this activity was inhibited by cyclohexyl-dodecyl urea. Arachidonic acid epoxygenase activity was detected in epididymal S9 fractions, suggesting that fatty acid metabolism by epididymal cytochrome P450s can form epoxides that subsequently become substrates for epididymal sEH. Results from the present study indicate that the epididymis contains at least two active forms of epoxide hydrolase. The role of these enzymes in the detoxification of xenobiotic epoxides is well known, although it is unclear what cellular role they may play in the formation of biologically active metabolites in the epididymis.     

Le syndrome de pseudo-Bartter néonatal d? à des désordres alimentaires maternels: peut-être plus fréquent qu’il n’y para?t

Pseudo-Bartter Syndrome is defined by a hypochloremic-hypokaliemic metabolic alkalosis but without primary tubular renal defects. This metabolic imbalance can result from a state of often hidden chronic vomiting in some psychiatric disorders. It can affect both the pregnant mother and her foetus. Owing to the paucity of symptoms due to the slow installation of these biochemical disturbances, a diagnosis of pseudo-Bartter syndrome could be missed at birth in the mother and her infant. This potential misdiagnosis could expose the mother and her baby to serious complications during the first hours of life. We report on two cases of neonatal pseudo-Bartter in which the diagnosis was made a posteriori in the mothers.