Neurofunctional Evaluation of Young Male Offspring of Rat Dams with Diabetes Induced by Streptozotocin

Diabetes mellitus (DM) is a complex disease, being one of the most prevalent diseases worldwide. As a consequence, pregnancy-associated diabetes is increasingly common. Given the numerous studies about the influence of diabetes on offspring of diabetic rat dams, the neurological outcome is of outmost importance. This paper aimed at evaluating the neurofunctional performance of young male offspring of rat dams with diabetes induced by streptozotocin. Diabetes was induced in Wistar female rats by streptozotocin administration, while control groups received vehicle injection. At two-month survival period, male offspring from each group were randomized to the water maze Morris test, in order to assess their neurofunctional status. There was no significant difference between the groups as assessed by the Morris water maze test for spatial reference task. Our results point to the need of further investigation on the offspring neurofunctional performance

Hyperthermia induced after recirculation triggers chronic neurodegeneration in the penumbra zone of focal ischemia in the rat brain

Chronic neurodegenerative processes have been identified in the rat forebrain after prolonged survival following hyperthermia (HT) initiated a few hours after transient global ischemia. Since transient global ischemia and ischemic penumbra share pathophysiological similarities, this study addressed the effects of HT induced after recirculation of focal brain ischemia on infarct size during long survival times. Adult male Wistar rats underwent intra-luminal occlusion of the left middle cerebral artery for 60 min followed by HT (39.0-39.5°C) or normothermia. Control procedures included none and sham surgery with and without HT, and middle cerebral artery occlusion alone. Part I: 6-h HT induced at recirculation. Part II: 2-h HT induced at 2-, 6-, or 24-h recirculation. Part III: 2-h HT initiated at recirculation or 6-h HT initiated at 2-, 6- or 24-h recirculation. Survival periods were 7 days, 2 or 6 months. The effects of post-ischemic HT on cortex and striatum were evaluated histopathologically by measuring the area of remaining tissue in the infarcted hemisphere at -0.30 mm from bregma. Six-hour HT initiated from 6-h recirculation caused a significant decrease in the remaining cortical tissue between 7-day (N = 8) and 2-month (N = 8) survivals (98.46 ± 1.14 to 73.62 ± 8.99%, respectively). When induced from 24-h recirculation, 6-h HT caused a significant reduction of the remaining cortical tissue between 2- (N = 8) and 6-month (N = 9) survivals (94.97 ± 5.02 vs 63.26 ± 11.97%, respectively). These data indicate that post-ischemic HT triggers chronic neurodegenerative processes in ischemic penumbra, suggesting that similar fever-triggered effects may annul the benefit of early recirculation in stroke patients over the long-term.Universidade Federal de São Paulo (UNIFESP) Laboratório de Fisiopatologia Clínica e ExperimentalUniversidade Federal de São Paulo (UNIFESP) Departamento de Neurologia e NeurocirurgiaUniversidade Federal de São Paulo (UNIFESP) Departamento de MedicinaUniversidade Federal de São Paulo (UNIFESP) Centro de Microscopia EletrônicaUniversidade Federal de São Paulo (UNIFESP) Escola Paulista de Medicina COLSAN-UNIFESPUNIFESP, Laboratório de Fisiopatologia Clínica e ExperimentalUNIFESP, Depto. de Neurologia e NeurocirurgiaUNIFESP, Depto. de MedicinaUNIFESP, Centro de Microscopia EletrônicaUNIFESP, EPM, COLSAN-UNIFESPSciEL

Effect of chronic sleep restriction and aging on calcium signaling and apoptosis in the hippocampus of young and aged animals

Aging leads to progressive deterioration of physiological function and diminished responses to environmental stress. Organic and functional alterations are frequently observed in elderly subjects. Although chronic sleep loss is observed during senescence, little is known about the impact of insufficient sleep on cellular function in aging neurons. Disruption of neuronal calcium (Ca2+) signaling is related to impaired neuronal function and cell death. It has been hypothesized that sleep deprivation may compromise neuronal stability and induce cell death in young neurons; however, it is necessary to evaluate the impact of aging on this process. Therefore, the aim of this study was to evaluate the effects of chronic sleep restriction (CSR) on Ca2+ signaling and cell death in the hippocampus of young and aged animals. We found that glutamate and carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) induced a greater elevation in cytosolic Ca2+ ([Ca2+](c)) in hippocampal slices from aged rats subjected to CSR compared to age-matched controls. Interestingly, aged-matched controls showed a reduced Ca2+ response to glutamate and FCCP, relative to both CSR and control young animals. Apoptotic nuclei were observed in aged rats from both treatment groups; however, the profile of apoptotic nuclei in aged CSR rats was highly variable. Bax and Bc1-2 protein expression did not change with aging in the CSR groups. Our study indicates that aging promotes changes in Ca2+ signaling, which may also be affected by CSR. These age-dependent changes in Ca2+ signaling may increase cellular vulnerability during CSR and contribute to Ca2+ signaling dysregulation, which may ultimately induce cell death. (c) 2012 Elsevier Inc. All rights reserved.Associacao Fundo de Incentivo a Pesquisa (AFIP)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)CEPIDConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Universidade Federal de São Paulo UNIFESP, Dept Psicobiol, BR-04024002 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Farmacol, BR-04044020 São Paulo, BrazilUN1FESP, Ctr Microscopia Eletron, São Paulo, BrazilUniversidade Federal de São Paulo UNIFESP, Dept Psicobiol, BR-04024002 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Farmacol, BR-04044020 São Paulo, BrazilFAPESP: 08/50424-3CEPID: 98/14303-3Web of Scienc

Epilepsy and neuroprotection: employment of the RPia during status epilepticus induced by pilocarpine

OBJECTIVE: The aim of this study was to characterize the neuroprotection of the RPia in rats subjected to status epilepticus (SE) induced by pilocarpine (Pilo). METHODS: We evaluated the mismatch between local cerebral glucose utilisation (LCGU) and local cerebral blood flow (LCBF) 4 hours after SE induction. Neuronal loss was evaluated by Fluoro Jade-B (FJB) 24 hours and 90 days after SE. Four groups were studied: Saline, Pilo, RPia+Saline and RPia+Pilo. RESULTS AND CONCLUSIONS: Significant increases in the LCGU were observed in the almost all brain regions of Pilo and RPia+Pilo groups compared to control. However, significant reduction in the LCGU occurred in the substantia nigra pars reticulata and hippocampal formation of RPia+Pilo group versus Pilo. There was significant increase of the LCBF in all the studied areas, comparing the Pilo and RPia+Pilo groups with the control. The increases of LCBF was more intense in rats from RPia+Pilo compared to Pilo, and located mainly in CA2, CA3, dentate gyrus, entorhinal cortex, thalamic nuclei, mammillary body, red nucleus, zone incerta, pontine nucleus and visual cortex. A great number FJB stained cells was observed in the Pilo group and RPia pretreatment reduced the staining in the hippocampal formation, piriform cortex, basolateral amygdala and substantia nigra pars compacta.OBJETIVO: O objetivo desse estudo foi caracterizar a neuroproteção do RPia em ratos submetidos ao status epilepticus (SE) induzido pela pilocarpina (Pilo). MÉTODOS: Avaliou-se o balanço entre utilização local da glicose cerebral (ULGC) e fluxo sanguíneo cerebral local (FSCL) após 4 horas de SE, e a marcação por Fluoro Jade-B (FJB), 24 horas e 90 dias após SE. Quatro grupos foram avaliados: Salina, Pilo, RPia+Salina e RPia+Pilo. RESULTADOS E CONCLUSÃO: Aumentos significantes na ULGC foram observados na maioria das regiões avaliadas nos grupos Pilo e RPia+Pilo quando comparados ao controle. Entretanto, redução significante na ULGC ocorreu na substância negra pars reticulata e giro denteado do grupo RPia+Pilo versus Pilo. Houve aumento significante do FSCL em todas as áreas estudadas, comparando-se os grupos Pilo e RPia+Pilo com o controle. Foi observado um aumento significante do FSCL durante SE em CA2, CA3, giro denteado, córtex entorrinal, corpo mamilar, núcleos talâmicos, núcleo rubro, zona incerta, núcleo oral da ponte e córtex visual, no grupo pré-tratado com RPia comparado ao tratado somente com Pilo. Grande número de células marcadas com FJB foi observado no grupo Pilo e o pré-tratamento com RPia reduziu essa marcação na formação hipocampal, córtex piriforme, amígdala basolateral e substância negra pars compacta.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)UNIFESP Departamento de Neurologia e NeurocirurgiaUniversité Louis Pasteur INSERM Unité 465UNIFESP Centro de Microscopia EletrônicaUNIFESP, Depto. de Neurologia e NeurocirurgiaUNIFESP, Centro de Microscopia EletrônicaSciEL

A total transcriptome profiling method for plasma-derived extracellular vesicles: applications for liquid biopsies

Extracellular vesicles (EVs) are key mediators of intercellular communication. Part of their biological effects can be attributed to the transfer of cargos of diverse types of RNAs, which are promising diagnostic and prognostic biomarkers. EVs found in human biofluids are a valuable source for the development of minimally invasive assays. However, the total transcriptional landscape of EVs is still largely unknown. Here we develop a new method for total transcriptome profiling of plasma-derived EVs by next generation sequencing (NGS) from limited quantities of patient-derived clinical samples, which enables the unbiased characterization of the complete RNA cargo, including both small- and long-RNAs, in a single library preparation step. This approach was applied to RNA extracted from EVs isolated by ultracentrifugation from the plasma of five healthy volunteers. Among the most abundant RNAs identified we found small RNAs such as tRNAs, miRNAs and miscellaneous RNAs, which have largely unknown functions. We also identified protein-coding and long noncoding transcripts, as well as circular RNA species that were also experimentally validated. This method enables, for the first time, the full spectrum of transcriptome data to be obtained from minute patient-derived samples, and will therefore potentially allow the identification of cell-to-cell communication mechanisms and biomarkers.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Gillson-Longenbaugh FoundationNational Institutes of Health (NIH/NCATS) through the NIH Common Fund, Office of Strategic Coordination (OSC)AC Camargo Canc Ctr, Lab Med Genom, Sao Paulo, SP, BrazilAC Camargo Canc Ctr, Lab Computat Biol, Sao Paulo, SP, BrazilUniv Sao Paulo, Inst Biomed Sci, Dept Cell & Dev Biol, Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Electron Microscopy Ctr, Sao Paulo, SP, BrazilUniv Texas MD Anderson Canc Ctr, Dept Expt Therapeut, Houston, TX 77030 USAUniv Texas MD Anderson Canc Ctr, Ctr RNA Interference & Non Coding RNAs, Houston, TX 77030 USAUniv New Mexico, Comprehens Canc Ctr, Albuquerque, NM 87131 USAUniv New Mexico, Sch Med, Div Hematol Oncol, Dept Internal Med, Albuquerque, NM 87131 USAUniv New Mexico, Sch Med, Div Mol Med, Dept Internal Med, Albuquerque, NM 87131 USARockefeller Univ, Lab Mol Immunol, 1230 York Ave, New York, NY 10021 USAFMUSP, Lab Neurociencias Alzira Denise Hertzog Silva LIM, Inst Psiquiatria, Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Electron Microscopy Ctr, Sao Paulo, SP, BrazilFAPESP: 2011/09172-3FAPESP: 2014/26897-0Web of Scienc

Analysis of the Virulence of an Atypical Enteropathogenic Escherichia coli Strain In Vitro and In Vivo and the Influence of Type Three Secretion System

Atypical enteropathogenic Escherichia coli (aEPEC) inject various effectors into intestinal cells through a type three secretion system (T3SS), causing attaching and effacing (A/E) lesions. We investigated the role of T3SS in the ability of the aEPEC 1711-4 strain to interact with enterocytes in vitro (Caco-2 cells) and in vivo (rabbit ileal loops) and to translocate the rat intestinal mucosa in vivo. A T3SS isogenic mutant strain was constructed, which showed marked reduction in the ability to associate and invade but not to persist inside Caco-2 cells. After rabbit infection, only aEPEC 1711-4 was detected inside enterocytes at 8 and 24 hours pointing to a T3SS-dependent invasive potential in vivo. In contrast to aEPEC 1711-4, the T3SS-deficient strain no longer produced A/E lesions or induced macrophage infiltration. We also demonstrated that the ability of aEPEC 1711-4 to translocate through mesenteric lymph nodes to spleen and liver in a rat model depends on a functional T3SS, since a decreased number of T3SS mutant bacteria were recovered from extraintestinal sites. These findings indicate that the full virulence potential of aEPEC 1711-4 depends on a functional T3SS, which contributes to efficient adhesion/invasion in vitro and in vivo and to bacterial translocation to extraintestinal sites

Os livros brancos da defesa da República Popular da China 1998-2010

Este estudo é uma análise da evolução das perceções de (in)segurança da República Popular da China (RPC), através da aferição quantitativa e qualitativa de expressões idiomáticas caracterizadoras da evolução do sistema internacional, as quais foram selecionadas e associadas a tais perceções, e que constam das sete edições do Livro Branco da Defesa publicadas pelo Conselho de Estado entre 1998 e 2010. Procura-se através de um enquadramento conceptual e metodológico derivado da análise crítica do discurso baseado nas teorias de Michel Foucault e de Norman Fairclough, bem como do da perceção de ameaças por parte dos Estados no sistema internacional formulado por Robert Jervis, identificar e justificar variações nas perceções de (in)segurança da RPC entre 1998 e 2010, concluindo-se que estas refletem uma visão de natureza essencialmente realista estrutural e Lockeana quanto à evolução do sistema internacional

Characteristics of chronic physiopathologic process of experimental model of global transient ischemic and the epilepsy of tempral lobe in rats

O estudo que constitui a parte inicial desta tese caracterizou, sob o ponto de vista histopatologico e imunohistoquimico, um processo neurodegenerativo cronico desencadeado pela inducao de 2 periodos consecutivos de hipertermia por aquecimento externo (entre a quarta e a nona hora no primeiro dia de recuperacao e entre a vigesima-primeira e a trigesima hora de recuperacao) apos a isquemia global de 10 minutos de duracao no prosencefalo de ratos wistar, sacrificados com 7 dias, 2 ou 6 meses. Verificou-se que o processo e acompanhado por varios sinais encontrados no cerebro humano afetado pela Doenca de Alzheimer, incluindo-se as placas neuriticas e os novelos neurofibrilares, alem da persistencia da ativacao microglial e do sistema citolitico do complemento. O aquecimento ambientar da gaiola nao foi efetivo para a inducao de hipertermia em animais controle, submetidos apenas ao procedimento cirurgico, o que impede o controle do efeito da hipertermia durante testes comportamentais. Assim, deu-se inicio um segundo estudo, o qual incluiu outros 18 grupos de animais, desta vez obtendo-se a inducao de um unico periodo de hipertermia (39,5§C entre a segunda e a nona hora de recirculacao) atraves de uma almofada de aquecimento e sob anestesia com halotano, para caracterizar-se o efeito funcional da hipertermia pos-isquemica atraves do teste do labirinto aquatico de Morris. Os grupos de animais foram destinados a 3 sobrevidas: 7 dias (6 grupos), 2 meses (6 grupos) e 6 meses (6 grupos). Os 6 grupos destinados para cada sobrevida foram assim constituidos: (a) animais normais, (b) animais submetidos apenas a cirurgia sem isquemia seguida de hipertermia sob halotano, (c) animais submetidos apenas a cirurgia sem isquemia seguida de normotermia sob halotano, (d) animais submetidos a isquemia sem tratamento posterior, (e) animais isquemicos submetidos a anestesia com halotano sob temperatura normal, (f) animais submetidos a isquemia seguida de hipertermia sob anestesia com halotano. Os grupos destinados as sobrevidas de 2 e 6 meses foram avaliados sob o ponto de vista comportamental, respectivamente, apos l e 5 meses de recuperacao, atraves de testes de memoria de referencia, Os resultados de latencia e trajeto, avaliados segundo a analise de variancia para medidas repetidas, demonstraram que, em ambas as sobrevidas de 1 e 5 meses, os animais submetidos a hipertermia pos-isquemica diferenciam-se larga e significativamente...(au)BV UNIFESP: Teses e dissertaçõe