Leptin enhances adult neurogenesis and reduces pathological features in a transgenic mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is the most common dementia worldwide and is characterized by the presence of senile plaques by amyloid-beta (Aß) and neurofibrillary tangles of hyperphosphorylated Tau protein. These changes lead to progressive neuronal degeneration and dysfunction, resulting in severe brain atrophy and cognitive deficits. With the discovery that neurogenesis persists in the adult mammalian brain, including brain regions affected by AD, studies of the use of neural stem cells (NSCs) for the treatment of neurodegenerative diseases to repair or prevent neuronal cell loss have increased. Here we demonstrate that leptin administration increases the neurogenic process in the dentate gyrus of the hippocampus as well as in the subventricular zone of lateral ventricles of adult and aged mice. Chronic treatment with leptin increased NSCs proliferation with significant effects on proliferation and differentiation of newborn cells. The expression of the long form of the leptin receptor, LepRb, was detected in the neurogenic niches by reverse qPCR and immunohistochemistry. Moreover, leptin modulated astrogliosis, microglial cell number and the formation of senile plaques. Additionally, leptin led to attenuation of Aß-induced neurodegeneration and superoxide anion production as revealed by Fluoro-Jade B and dihydroethidium staining. Our study contributes to the understanding of the effects of leptin in the brain that may lead to the development of new therapies to treat Alzheimer's disease.

Evaluation of the isoflavones and estrogen effects on the rat adrenal.

The aim of this study was to evaluate the morphometry and the gene expression of Ki-67, VEGF and caspase 3 and the stress oxidative in the adrenal gland of ovariectomized rats treated with estrogen or isoflavones. We used 15 Wistar rats ovariectomized treated with isoflavones or estrogen during 30 days. At the end of the treatment, the left adrenal gland was removed for subsequent histological studies and the right was used to evaluate gene expression of angiogenesis (VEGF-A), cell proliferation (Ki-67), apoptose (caspase 3 clivated) and oxidative stress. Treatment with estrogen showed a largest increase in the layers of the adrenal cortex than with isoflavones. These hypertrofic effects agree with higher expression elevation of Ki67 and VEGF, which did not occur with the caspase 3, indicating that isoflavones have great proliferative effect on the adrenal gland. Similar results were also observed on superoxide quantification show that isoflavone has a protective effect against oxidative stress. Our results indicate positively the trophic therapeutic potential of isoflavones has a protective effect and can contribute to the development of effective therapies to decrease the symptoms of menopause.

Essential Elements as Biomarkers of Acute Kidney Injury and Spontaneous Reversion.

Acute kidney injury (AKI) is an important health problem and can be caused by number of factors. The use of aminoglycosides, such as gentamicin, is one of these factors. Recently, an effort has been made to find biomarkers to guide treatment protocols. Inductively coupled plasma optical emission spectroscopy (ICP-OES) was used to estimate the contents of Ca, Cu, Fe, K, Mg, Mn, Na, P, and Zn in serum and urine of the healthy, AKI, and spontaneous recovery (SR) groups of animals. The animal model of AKI and SR was validated by measuring serum and urinary urea and creatinine. The quantitative determination of the elements showed a decrease in serum levels of Ca, and Fe in the AKI group (P<0.01 vs. healthy), with a return to normal levels in the SR group, without a significant difference between the healthy and SR groups. In the urine samples, there was a decrease in P and Na levels in the AKI group (P<0.001 and P<0.01 vs. healthy), but Ca levels were increased in this group compared with the healthy and SR groups (P<0.01). These findings indicate that mineral elements might be useful as biomarkers for AKI.

Central administration of a nitric oxide precursor abolishes both the hypothalamic serotonin release and the hypophagia induced by interleukin-1beta in obese Zucker rats.

Serotonin-induced anorexia has long been recognized as an important part of the CNS mechanisms controlling energy balance. More recently, interleukin-1beta and nitric oxide have been suggested to influence this control, possibly through modulation of hypothalamic serotonin. The present work aimed at investigating the interaction of these systems. We addressed whether 5-HT is affected during IL-1beta-induced anorexia in obese Zucker rats and the influence of the central NO system on this IL-1beta/5-HT interaction. Using microdialysis, we observed that an intracerebroventricular injection of 10 ng IL-1beta significantly stimulated 5-HT extracellular levels in the VMH, with a peak variation of 102+/-41% above baseline. IL-1beta also significantly reduced the 4-h feeding by 33% and the 24-h feeding by 42%. Contrarily, these effects were absent when IL-1beta was injected 2 h after the i.c.v. administration of 20 microg of the NO precursor L-arginine. The results suggest that, in obese Zucker rats, activation of the serotonergic system in the medial hypothalamus participates in IL-1beta-induced anorexia. Since L-arginine, probably through NO stimulation, abolished both the anorexia and the serotonergic activation, it can be proposed that the NO system, either directly or indirectly, counteracts IL-1beta anorexia. The hypothalamic serotonergic system is likely to mediate this NO effect.

Transplantation of bone marrow mesenchymal stem cells decreases oxidative stress, apoptosis, and hippocampal damage in brain of a spontaneous stroke model.

Stroke is the most common cause of motor disabilities and is a major cause of mortality worldwide. Adult stem cells have been shown to be effective against neuronal degeneration through mechanisms that include both the recovery of neurotransmitter activity and a decrease in apoptosis and oxidative stress. We chose the lineage stroke-prone spontaneously hypertensive rat (SHRSP) as a model for stem cell therapy. SHRSP rats can develop such severe hypertension that they generally suffer a stroke at approximately 1 year of age. The aim of this study was to evaluate whether mesenchymal stem cells (MSCs) decrease apoptotic death and oxidative stress in existing SHRSP brain tissue. The results of qRT-PCR assays showed higher levels of the antiapoptotic Bcl-2 gene in the MSC-treated animals, compared with untreated. Our study also showed that superoxide, apoptotic cells, and by-products of lipid peroxidation decreased in MSC-treated SHRSP to levels similar those found in the animal controls, Wistar Kyoto rats. In addition, we saw a repair of morphological damage at the hippocampal region after MSC transplantation. These data suggest that MSCs have neuroprotective and antioxidant potential in stroke-prone spontaneously hypertensive rats.

Differential Shh, Bmp and Wnt gene expressions during craniofacial development in mice.

In this study, Bmp-4, Wnt-5a and Shh gene expressions were compared during early craniofacial development in mice by comparative non-isotopic in situ hybridization. Wild-type C57BL/6J mice were studied at various stages of embryonic development (from 8.5- to 13.5-day-old embryos--E8.5-13.5). During early odontogenesis, transcripts for Bmp-4, Shh and Wnt-5a were co-localised at the tooth initiation stage. At E8.5, Shh mRNA expression was restricted to diencephalon and pharyngeal endoderm. Before maxillae and mandible ossification, Bmp-4 and Wnt-5a signals were detected in the mesenchymal cells and around Meckel's cartilage. During palatogenesis, Shh was expressed only in the epithelium and Wnt-5a only in the mesenchyme of the elevating palatal shelves. During tongue development, Shh expression was found in mesenchyme, probably contributing to tongue miogenesis, while Wnt-5a signal was in the epithelium, possibly during placode development and papillae formation. Taken together, these findings suggest that Bmp-4, Shh and Wnt-5a gene expressions may act together on the epithelial-mesenchymal interactions occurring in several aspects of the early mouse craniofacial development, such as odontogenesis, neuronal development, maxillae and mandible ossification, palatogenesis and tongue formation.

Localization of Bmp-4, Shh and Wnt-5a transcripts during early mice tooth development by in situ hybridization.

A comparative nonisotopic in situ hybridization (ISH) analysis was carried out for the detection of Bmp-4, Shh and Wnt-5a transcripts during mice odontogenesis from initiation to cap stage. Bmp-4 was expressed early in the epithelium and then in the underlying mesenchyme. Shh expression was seen in the odontogenic epithelial lining thickening, being stronger in the enamel knot area, during the cap stage. Wnt-5a transcripts were expressed only in the mesenchyme during the initiation, bud and cap stages, with strong expression in the dental mesenchyme during the bud stage. The present results showed that Bmp-4, Shh and Wnt-5a are expressed since the very early stages of tooth development, and they suggest that the Wnt-5a gene is expressed in different cell populations than Bmp-4 and Shh.

Methods of acute biological assays in guinea-pigs for the study of toxicity and innocuity of drugs and chemicals

In this study, 602 samples were tested by the following assays performed at the animal facilities (Cedeme) of the Federal University of São Paulo (UNIFESP): 385 for dermal irritability, 90 for ocular irritability (discontinued in 1995), 31 for systemic toxicity by injection, 26 for oral acute toxicity, 15 for toxicity by intracutaneous injection, 15 for skin sensitization, 15 for toxicity of serum and vaccines for human use, 14 for toxicity by intramuscular implantation, 7 for pyrogens, 2 for acute dermal toxicity, and 2 for irritation of mucous membrane. The following agents were tested: cosmetics and related substances (42.0 percent), chemicals used in industry (32.9 percent), plastics, rubber, and other polymers (15.9 percent), agrotoxics (4.0 percent), medicines (2.7 percent), and vaccines (2.5 percent). In the present description, emphasis was given to tests of dermal irritability and sensitization. This work was conducted entirely in animal facilities, according to our general belief that animal facilities at universities, while considering ethic principles and sanitary, genetic, nutritional, and pathophysiological controls, also require laboratories specialized in areas such as transgenics, cryopreservation, ambiental physiology, functional genomics, alternative models, and mainly activities and research on methods in toxicology, as focused in this study.

Descrevemos os testes usados em ensaios biológicos de curta duração para estudo de toxicidade e inocuidade de cosméticos, fármacos e outras substâncias químicas, feitos no Biotério Central/Cedeme da Unifesp, de 1986 a 2000. Testamos 602 amostras nos seguintes ensaios: 385 de irritação cutânea, 90 de irritação ocular (até 1995), 31 de toxicidade sistêmica por injeção, 26 de toxicidade oral aguda, 15 de toxicidade por aplicação intracutânea, 15 de sensibilização da pele, 15 de toxicidade de soros e vacinas de uso humano, 14 de toxicidade por implantação intramuscular, 7 de pirogênio, 2 de toxicidade dérmica aguda e 2 de irritação da mucosa. Os agentes testados foram: cosméticos e suas matérias-primas (42,0 por cento), substâncias químicas industriais (32,9 por cento), plásticos, borrachas e outros polímeros (15,9 por cento), defensivos agrícolas (4,0 por cento), medicamentos (2,7 por cento) e vacinas (2,5 por cento). Aqui daremos ênfase aos ensaios de irritação e sensibilização cutânea. Este trabalho foi feito inteiramente em biotério, em consonância com a idéia de que os biotérios em universidades, sem deixar de considerar os princípios éticos pertinentes e sem desconsiderar a presença de laboratórios para controles sanitário, genético, nutricional e fisiopatológico, devem ter também laboratórios para pesquisa em transgênicos, criopreservação, fisiologia ambiental, genômica funcional, modelos alternativos e fundamentalmente toxicologia, entre outros.

Localization of Bmp-4, Shh and Wnt-5a transcripts during early mice tooth development by in situ hybridization

A comparative nonisotopic in situ hybridization (ISH) analysis was carried out for the detection of Bmp-4, Shh and Wnt-5a transcripts during mice odontogenesis from initiation to cap stage. Bmp-4 was expressed early in the epithelium and then in the underlying mesenchyme. Shh expression was seen in the odontogenic epithelial lining thickening, being stronger in the enamel knot area, during the cap stage. Wnt-5a transcripts were expressed only in the mesenchyme during the initiation, bud and cap stages, with strong expression in the dental mesenchyme during the bud stage. The present results showed that Bmp-4, Shh and Wnt-5a are expressed since the very early stages of tooth development, and they suggest that the Wnt-5a gene is expressed in different cell populations than Bmp-4 and Shh.

No presente trabalho, realizou-se uma análise comparativa não isotópica por hibridização in situ a fim de se detectar a presença de transcritos de Bmp-4, Shh e Wnt-5a durante as fases iniciais da odontogênese em camundongos, desde a iniciação até o estágio de capuz. No estágio de iniciação, observou-se expressão precoce de Bmp-4 no epitélio e no mesênquima subjacente, enquanto que a expressão de Shh ocorreu durante o estágio de capuz, na região de espessamento do revestimento epitelial odontogênico, tornando-se mais intensa na área de nó do esmalte. Os transcritos de Wnt-5a foram expressos somente no mesênquima durante os estágios de iniciação, botão e capuz, com intenso sinal na região no mesênquima na fase de botão. Estes resultados mostraram que Bmp-4, Shh e Wnt-5a são expressos desde os estágios mais precoces do desenvolvimento dentário, sugerindo que o gene Wnt-5a seja expresso em populações celulares distintas daquelas que expressam Bmp-4 e Shh.