Identification of neuronal alterations induced by Shank3 mutations using iPS cells from Phelan-McDermid Patients' Fibroblasts

A família de proteínas Shank é o principal conjunto de proteinas de suporte e está localizada na densidade pós-sináptica das sinapses excitatórias. Existem 3 genes na família Shank, Shank1, Shank2 e Shank3 e são caracterizados por múltiplos domínios repetidos de anquirina próximo ao N-terminal seguido pelos domínios Src homologo 3 e PDZ, uma região longa rica em prolina e um domínio de motivo α estéril próximo ao C-terminal. Shank proteínas conectam duas subunidades de receptors glutamatérgicos, recetores NMDA e recetores metabotrópicos de glutamato do tipo-I (mGluRs). O domínio PDZ da Shank conecta-se ao C-terminal do GKAP e este, liga-se, ao complexo recetor PSD-95-NMDA. Por outro lado, a proteína Homer interage com o domínio rico em prolina para confirmar a associação entre a proteína Shank com o mGluR tipo-I. A proteína específica em estudo, Shank3, é haploinsuficiente em pacientes com sindrome Phelan-McDermid devido à deleções no braço comprido do cromossoma 22 levando à danos intelectuais, ausência ou atraso no discurso, comportamentos semelhantes ao autismo, hipotonia e características dismórficas. Neste trabalho, investigamos o papel da Shank3 na função sináptica para compreender a relação entre alterações nesta proteína e as características neurológicas presente em Pacientes com síndrome Phelan-McDermid. Foram utilizados dois modelos diferentes, ratinhos knockout Shank3 e hiPSC de pacientes com PMS. Ratinhos geneticamente modificados são ferramentas uteis no estudo de genes e na compreensão dos mecanismos que experiências in vitro não são capazes de reproduzir, mas de maneira a compreender melhor as patologias humanas, decidimos trabalhar também com células humanas. Os fibroblastos dos pacientes com síndrome Phelan-McDermid fora reprogramados em hiPS cells, diferenciados em neurónios e comparados com os neurónios obtidos a partir de doadores saudavéis e da mesma idade. A reprogramação em iPSC foi realizada por infecção de lentivirus com quatro genes de reprogramação OCT4, c-MYC, SOX2 e KFL4 para posteriormente serem diferenciados em neurónios, com cada passo sendo positivamente confirmado através de marcadores neuronais. Através dos neurónios diferenciados, analisamos a expressão de proteínas sinápticas. Pacientes com haploinsuficiencia na proteína Shank3 apresentam níveis elevados de proteína mGluR5 e decrescidos de proteína Homer sugerindo que a haploinsuficiencia leva a desregulação do complexo mGluR5-Homer-Shank3 conduzindo também, a defeitos na maturação sináptica. Assim, a expressão da proteína mGluR5 está alterada nos pacientes com PMS podendo estar relacionada com defeitos encontrados na diferenciação neuronal e maturação sináptica observados nos neurónios de pacientes. Conclusivamente, iPS cells representam um modelo fundamental no estudo da proteína Shank3 e a sua influência no sindrome de Phelan-McDermid.

Nephrotoxicity effects of the wood preservative chromium copper arsenate on mice: histopathological and quantitative approaches

Chromium copper arsenate(CCA)was used for the protection of wood building material suntil the restriction by EPA in2002. During a short period of time 14–24h,a comparative nephrotoxicity study was performed regarding the effects of CCA and its compounds per se. Histopathological and histochemical features were correlated with the concentration of the total arsenic and chromium in mice kidney. Animals were subcutaneously injected with CCA(7.2mg/kg arsenic and 10.2mg/kg chromium per body weight), CrO3 (10.2 mg/kg),As2O5 (7.2 mg/kg)andNaCl(0.9%) per se. The histopathological examination of the renal sections evidenced acute tubular necrosis in the groups of animals exposed to CCA(in both periods of time). Although the same contents of pentavalent arsenic and hexavalent chromium were injected in treated animals with CCA and with the prepared solutions of As2O5 and CrO3, the arsenic concentration on kidneys of CCA-exposed animals was much higher than those in animals exposed to As2O5 (32- and28-fold higher at 14 and 24h,respectively). However,the elimination of chromium seems to occur similarly in the kidneys of animals treated with CCA and CrO3 per se. Interactions among the components of CCA result in a marked decrease of the ability of kidney to eliminate simultaneously both analytes.The nephrotoxicity of CCA was higher than its components per se, evidencing a possible synergetic effect.

Subacute effects of the thiodicarb pesticide on target organs of male wistar rats: biochemical, histological, and flow cytometry studies

Thiodicarb, a carbamate pesticide widely used on crops, may pose several environmental and health concerns. This study aimed to explore its toxicological profile on male rats using hematological, biochemical, histopathological, and flow cytometry markers. Exposed animals were dosed daily at 10, 20, or 40 mg/kg/body weight (group A, B, and C, respectively) during 30 d. No significant changes were observed in hematological parameters among all groups. After 10 d, a decrease of total cholesterol levels was noted in rats exposed to 40 mg/kg. Aspartate aminotransferase (AST) activity increased (group A at 20 d; groups A and B at 30 d) and alkaline phosphatase (ALP) (group B at 30 d) activity significantly reduced. At 30 d a decrease of some of the other evaluated parameters was observed with total cholesterol and urea levels in group A as well as total protein and creatinine levels in groups A and B. Histological results demonstrated multi-organ dose-related damage in thiodicarb-exposed animals, evidenced as hemorrhagic and diffuse vacuolation in hepatic tissue; renal histology showed disorganized glomeruli and tubular cell degeneration; spleen was ruptured with white pulp and clusters of iron deposits within red pulp; significant cellular loss was noted at the cortex of thymus; and degenerative changes were observed within testis. The histopathologic alterations were most prominent in the high-dose group. Concerning flow cytometry studies, an increase of lymphocyte number, especially T lymphocytes, was seen in blood samples from animals exposed to the highest dose. Taken together, these results indicate marked systemic organ toxicity in rats after subacute exposure to thiodicarb.

Metabolic alterations in pregnant women: gestational diabetes

Gestational diabetes mellitus (GDM) and controversy are old friends. The impact of GDM on maternal and fetal health has been increasingly recognized. Nevertheless, universal consensus on the diagnostic methods and thresholds has long been lacking. Published guidelines from major societies differ significantly from one another, with recommendations ranging from aggressive screening to no routine screening at all. As a result, real-world practice is equally varied. This article recaps the latest evidence-based recommendations for the diagnosis and classification of GDM. It reviews the current evidence base for intensive multidisciplinary treatment of GDM and provides recommendations for postpartum management to delay and/or prevent progression to type 2 diabetes.

Role of oxidative stress-induced systemic and cavernosal molecular alterations in the progression of diabetic erectile dysfunction

Background Erectile dysfunction (ED) is a prevalent complication of diabetes, and oxidative stress is an important feature of diabetic ED. Oxidative stress-induced damage plays a pivotal role in the development of tissue alterations. However, the deleterious effects of oxidative stress in the corpus cavernosum with the progression of diabetes remain unclear. The aim of this study was to evaluate systemic and penile oxidative stress status in the early and late stages of diabetes. Methods Male Wistar streptozotocin-diabetic rats (and age-matched controls) were examined 2 (early) and 8 weeks (late) after the induction of diabetes. Systemic oxidative stress was evaluated by urinary H2O2 and the ratio of circulating reduced/oxidized glutathione (GSH/GSSG). Penile oxidative status was assessed by H2O2 production and 3-nitrotyrosine (3-NT) formation. Cavernosal endothelial nitric oxide synthase (eNOS) was analyzed by quantitative immunohistochemistry. Dual immunofluorescence was also performed for 3-NT and α-smooth muscle actin (α-SMA) and eNOS–α-SMA. Results There was a significant increase in urinary H2O2 levels in both diabetic groups. The plasma GSH/GSSG ratio was significantly augmented in late diabetes. In cavernosal tissue, H2O2 production was significantly increased in late diabetes. Reactivity for 3-NT was located predominantly in cavernosal smooth muscle (SM) and was significantly reduced in late diabetes. Quantitative immunohistochemistry revealed a significant decrease in eNOS levels in cavernosal SM and endothelium in late diabetes. Conclusions The findings indicate that the noxious effects of oxidative stress are more prominent in late diabetes. Increased penile protein oxidative modifications and decreased eNOS expression may be responsible for structural and/or functional deregulation, contributing to the progression of diabetes-associated ED.

Aerobic Exercise Does Not Predict Brain Derived Neurotrophic Factor And Cortisol Alterations in Depressed Patients

The pathophysiology of depression is related to neurobiological changes that occur in the monoamine system, hypothalamic-pituitary-adrenal axis, neurogenesis system and the neuroimmune system. In recent years, there has been a growing interest in the research of the effects of exercise on brain function, with a special focus on its effects on brainderived neurotrophic factor (BDNF), cortisol and other biomarkers. Thus, the aim of this study is to present a review investigating the acute and chronic effects of aerobic exercise on BDNF and cortisol levels in individuals with depression. It was not possible to establish an interaction between aerobic exercise and concentration of BDNF and cortisol, which may actually be the result of the divergence of methods, such as type of exercises, duration of the sessions, and prescribed intensity and frequency of sessions.