Bioanalytical LC-QTOF/MS Method for a N-phenylpiperazine Derivate (LQFM05): An Anxiolytic- and Antidepressant-like Prototype Drug Applied to Pharmacokinetic and Biodistribution Studies.

The LQFM05 is a prototype drug designed for treatment of psychiatric disorders, such as schizophrenia, exhibiting anxiolytic- and antidepressant-like (12 or 24 µmol/kg) effects in classical behavioral tests. In order to evaluate its pharmacokinetic properties, a liquid chromatography method coupled to a quadrupole time of flight mass spectrometry system (LC-QTOF/MS) was developed and fully validated for LQFM05 analysis in rat plasma and tissue samples (brain, heart, liver, and kidneys). Liquid-liquid extraction, solid phase extraction and protein precipitation were assessed as clean-up procedures for biological samples and analyte enrichment. Plasma and tissue samples underwent protein precipitation as a preliminary step, using acetonitrile. Linearity was fully demonstrated for the dynamic range (10.0 to 900.0 ng/mL), with r2 values higher than 0.99 (RSDslope ≤ 2%, Fcal < Ftab, Ccal < Ctab). Biodistribution studies in rats revealed high brain tissue concentrations (12.4 µg/g), suggesting elevated drug affinity to the main therapeutic target tissue, showing a blood partition coefficient of 1.9. Kidneys also showed great exposure and tissue affinity, suggesting a potential extrahepatic clearance. Likewise, all examined tissues exhibited satisfactory LQFMF05 distribution. The mass fragmentation spectrum indicated the presence of its main metabolite, LQFM235, yielded by high hepatic hydroxylation route, an equally bioactive derivative. Lastly, the developed LC-QTOF/MS method was shown to be sensitive (LOQ = 10 ng/mL), precise and accurate for LQFM05 determination in tissue homogenates and plasma samples.

Predictors of renal function recovery among patients undergoing renal replacement therapy following orthotopic liver transplantation.

Renal dysfunction frequently occurs during the periods preceding and following orthotopic liver transplantation (OLT), and in many cases, renal replacement therapy (RRT) is required. Information regarding the duration of RRT and the rate of kidney function recovery after OLT is crucial for transplant program management. We evaluated a sample of 155 stable patients undergoing post-intensive care hemodialysis (HD) from a patient population of 908 adults who underwent OLT. We investigated the average time to renal function recovery (duration of RRT required) and determined the risk factors for remaining on dialysis > 90 days after OLT. Log-rank tests were used for univariate analysis, and Cox proportional hazards models were used to identify factors associated with the risk of remaining on HD. The results of our analysis showed that of the 155 patients, 28% had pre-OLT diabetes mellitus, 21% had pre-OLT hypertension, and 40% had viral hepatitis. Among the patients, the median MELD (Model for End-Stage Liver Disease) score was 27 (interquartile range [IQR] 22-35). When they were listed for liver transplantation, 32% of the patients had serum creatinine (Scr) levels > 1.5 mg/dL or were on HD, and 50% had serum creatinine (Scr) levels > 1.5 mg/dL or were on HD at the time of OLT. Of the transplanted patients, 25% underwent pre-OLT intermittent HD, and 14% and 41% underwent continuous renal replacement therapy (CRRT) pre-OLT and post-OLT, respectively. At 90 days post-OLT, 118 (76%) patients had been taken off dialysis, and 16 (10%) patients had died while undergoing HD. The median recovery time of these post-OLT patients was 33 (IQR 27-39) days. In the multivariate analysis, fulminant hepatic failure as the cause of liver disease (p<0.001), the absence of pre-OLT hypertension (p = 0.016), a lower intraoperative fresh-frozen plasma (FFP) transfusion volume (p = 0.019) and not undergoing pre-OLT intermittent HD (p = 0.032) were associated with performing RRT for less than 90 days. Therefore, a high proportion of OLT patients showed improved renal function after OLT, and those who were diagnosed with fulminant hepatic failure, had no pre-OLT hypertension, received a lower transfused volume of intraoperative FFP and did not undergo pre-OLT intermittent HD had a higher probability of recovery.

Central H1 and H2 receptor participation in the control of water and salt intake in rats.

The aim of the present study was to evaluate the participation of brain H1 and H2 histaminergic receptors on water and salt intake induced by water deprivation (24 h), furosemide-induced sodium depletion and central angiotensinergic pharmacological stimulation in rats. Third ventricle injections of the H1 and H2 receptor antagonists, mepyramine (50, 100, 200 and 400 nmol) and cimetidine (100, 200 and 400 nmol), were unable to modify water intake induced by water deprivation and sodium depletion. Salt intake elicited by water deprivation and sodium depletion was reduced by the central administration of mepyramine, while intracerebroventricular administration of cimetidine had no effect. Water and salt intake evoked by central angiotensinergic stimulation (10 ng) was diminished by third ventricle injections of both mepyramine and cimetidine. Inhibition of the ingestive behaviors observed here is not a result of any illness-like effect produced by the intracerebroventricular injections of the histaminergic antagonists used, as demonstrated by an avoidance test. It was also shown that third ventricle injections of these compounds were unable to modify the hedonic behavior that leads rats to drink a tasty saccharin solution. We conclude that central histaminergic receptors participate in the control of salt intake induced by distinct physiological and pharmacological stimuli.

Superoxide overproduction and kidney fibrosis: a new animal model.

OBJECTIVE: To establish whether the mutation in the Immp2L gene induces renal fibrosis and whether aging exacerbates renal morphology in mice. METHODS: Female mutant mice with mutation in the inner mitochondrial membrane peptidase 2-like protein at 3 and 18 months of age were used. Renal fibrosis was analyzed using classic fibrosis score, Masson's trichrome staining, and analysis of profibrotic markers using real time polymerase chain reaction (superoxide dismutase 1, metalloproteinase-9, erythropoietin, transforming growth factor beta), and immunostaining (fibroblasts and Type IV collagen). Oxidative stress markers were determined by immunohistochemistry. The number of renal apoptotic cells was determined. Renal function was estimated by serum creatinine. RESULTS: Young mutant mice had significantly more glomerulosclerosis than age-matched mice (p=0.034). Mutant mice had more tubular casts (p=0.025), collagen deposition (p=0.019), and collagen type IV expression (p<0.001). Superoxide dismutase 1 expression was significantly higher in young mutants (p=0.038). Old mutants exhibited significantly higher expression of the fibroblast marker and macrophage marker (p=0.007 and p=0.012, respectively). The real time polymerase chain reaction of metalloproteinase-9 and erythropoietin were enhanced 2.5- and 6-fold, respectively, in old mutants. Serum creatinine was significantly higher in old mutants (p<0.001). CONCLUSION: This mutation altered renal architecture by increasing the deposition of extracellular matrix, oxidative stress, and inflammation, suggesting a protective role of Immp2L against renal fibrosis.

Central 5-HT2B/2C and 5-HT3 receptor stimulation decreases salt intake in sodium-depleted rats.

In the present study, we investigated the participation of central 5-HT(2B/2C) and 5-HT(3) receptors in the salt intake induced by sodium depletion in Wistar male rats. Sodium depletion was produced by the administration of furosemide associated with a low salt diet. Third ventricle injections of mCPP, a 5-HT(2B/2C) agonist, at doses of 80, 160 and 240 nmol, promoted a dose-dependent reduction in salt intake in sodium-depleted rats. The inhibitory effect produced by central administration of mCPP was abolished by the central pretreatment with SDZ SER 082, a 5-HT(2B/2C) antagonist. Similar results were obtained with third ventricle injections of m-CPBG (80, 160 and 240 nmol), a selective 5-HT(3) agonist that also induced a dose-related decrease in salt intake in sodium-depleted rats. The central pretreatment with LY-278,584, a selective 5-HT(3) receptor antagonist, was able to impair the salt intake inhibition elicited by third ventricle injections of m-CPBG. Central administration of each one of the antagonists alone or a combination of both antagonists together did not significantly change salt intake after sodium depletion. On the other hand, the central administration of both mCPP and m-CPBG, in the highest dose used to test their effect on salt intake (240 nmol), was unable to modify blood pressure in sodium-depleted rats. It is concluded that: (1) pharmacological activation of central 5-HT(2B/2C) and 5-HT(3) receptors diminishes salt intake during sodium depletion, (2) an inhibitory endogenous drive exerted by central 5-HT(2B/2C) and 5-HT(3) receptors does not seem to exist and (3) the reduction in salt intake generated by the pharmacological activation of these central receptors is not produced by an acute hypertensive response.

Histaminergic H1 and H2 receptors located within the ventromedial hypothalamus regulate food and water intake in rats.

The aim of the present study was to investigate the effect of the pharmacological blockade of histamine H1 and H2 receptors located within the ventromedial hypothalamus (VMH) on overnight food and water intake and on water intake elicited by two physiological stimuli: hyperosmolarity induced by an acute intragastric salt load and water deprivation. During the overnight period, the pharmacological blockade of both H1 and H2 VMH receptors significantly increased food intake and decreased water intake. In hyperosmotic rats, the blockade of H1 VMH receptors reduced water intake, while the blockade of H2 receptors in this same region yielded no significant effect. Additionally, in water-deprived rats, the blockade of both H1 and H2 receptors located within the VMH induced a significant decrease in water intake. The inhibitory effects on drinking behavior observed in this study do not seem to be a consequence of any "illness-inducing" effect provoked by the central administration of the antihistaminergic agents employed here, because an aversion test indicated that the injection of those compounds into the VMH does not induce any "illness-like" effect. In addition, the central administration of either mepyramine or cimetidine to dehydrated and hyperosmotic rats did not produce any reduction in locomotor activity measured in an open-field arena. Injections of the antihistaminergic agents used here into the regions that circumscribe the VMH produced no significant effects on water or food intake, indicating that the actions observed here may be specifically attributed to the set of histaminergic receptors situated within the VMH.

Superoxide overproduction and kidney fibrosis: a new animal model / Produção aumentada de superóxido e fibrose renal: novo modelo animal

Objective To establish whether the mutation in the Immp2L gene induces renal fibrosis and whether aging exacerbates renal morphology in mice. Methods Female mutant mice with mutation in the inner mitochondrial membrane peptidase 2-like protein at 3 and 18 months of age were used. Renal fibrosis was analyzed using classic fibrosis score, Masson’s trichrome staining, and analysis of profibrotic markers using real time polymerase chain reaction (superoxide dismutase 1, metalloproteinase-9, erythropoietin, transforming growth factor beta), and immunostaining (fibroblasts and Type IV collagen). Oxidative stress markers were determined by immunohistochemistry. The number of renal apoptotic cells was determined. Renal function was estimated by serum creatinine. Results Young mutant mice had significantly more glomerulosclerosis than age-matched mice (p=0.034). Mutant mice had more tubular casts (p=0.025), collagen deposition (p=0.019), and collagen type IV expression (p<0.001). Superoxide dismutase 1 expression was significantly higher in young mutants (p=0.038). Old mutants exhibited significantly higher expression of the fibroblast marker and macrophage marker (p=0.007 and p=0.012, respectively). The real time polymerase chain reaction of metalloproteinase-9 and erythropoietin were enhanced 2.5- and 6-fold, respectively, in old mutants. Serum creatinine was significantly higher in old mutants (p<0.001). Conclusion This mutation altered renal architecture by increasing the deposition of extracellular matrix, oxidative stress, and inflammation, suggesting a protective role of Immp2L against renal fibrosis. .

Objetivo Estabelecer se a mutação no gene Immp2L induz à fibrose renal e se o envelhecimento exacerba a morfologia renal em camundongos. Métodos Foram usadas fêmeas de camundongos mutantes para proteína semelhante à peptidase 2 da camada interna da mitocôndria, com 3 e 18 meses de idade. Para analisar a fibrose renal, foram usados o escore clássico de fibrose, a coloração com tricrômio de Masson, e a análise de marcadores profibróticos, por meio da reação em cadeia de polimerase em tempo real (superóxido dismutase 1, metalonoproteinase-9, eritropoietina e fator transformador de crescimento beta), e a imunocoloração (fibroblastos e colágeno IV). Marcadores de estresse oxidativo foram determinados por imuno-histoquímica. O número de células apoptóticas renais foi analisado. A função renal foi estimada por creatinina sérica. Resultados Camundongos mutantes jovens apresentaram glomeruloesclerose em quantidade significativamente maior que animais da mesma idade (p=0,034). Os mutantes mostraram maior formação de cilindros tubulares (p=0,025), deposição de colágeno (p=0,019) e maior expressão de colágeno do tipo IV (p<0,001). A expressão de superóxido dismutase 1 foi maior em mutantes jovens (p=0,038). Mutantes idosas exibiram maior expressão dos marcadores de fibroblastos e macrófagos (p=0,007 e p=0,012, respectivamente). As reações da cadeia de polimerase em tempo real da metalanoproteinase-9 e da eritropoietina estavam aumentadas em 2,5 e 6 vezes, respectivamente, em mutantes idosas. A creatinina sérica foi significantemente maior em animais idosos mutantes (p<0,001). Conclusão Essa mutação alterou a arquitetura renal pelo aumento da deposição de matriz extracelular, estresse oxidativo e inflamação, sugerindo papel de proteção de Immp2L contra a fibrose renal. .

Academética: um novo método de estudo continuado sobre Ética Médica e Bioética / Academética: a new method of continuous study in Medical Ethics and Bioethics

A consciência de que a prática deve estar fundamentada na postura ética dos profissionais leva à crescente importância de um ensino continuado e aprofundado, que ressalte os valores morais e humanos e o comprometimento social dos estudantes. Buscando adequar-se a esse novo paradigma, estudantes de Medicina da Universidade Federal da Bahia aceitaram a sugestão de criação, em novembro de 2000, de um grupo denominado Academética - Associação de Acadêmicos para o estudo da Ética Médica e Bioética. Os objetivos traçados pela equipe são: continuar o contato com o conteúdo da disciplina Ética Médica, aprofundando e atualizando as discussões acerca de Ética Médica e Bioética, e desenvolver trabalhos ligados a este assunto. As discussões de temas são realizadas através do estudo de artigos, vídeos e estímulo à participação dos estudantes em congressos, palestras e afins. Desta forma, a Academética se constitui em mais uma ferramenta para que os alunos durante a graduação na faculdade de Medicina possam desenvolver o senso crítico e conhecimento acerca de assuntos que englobem a Ética Médica e Bioética.