Navegando por Palavras-chave "Germ Cel"

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    Papel da caspase-3 na quiescência dos gonócitos de ratos
    (Universidade Federal de São Paulo (UNIFESP), 2019-01-31) Santos, Marina Nunes Dos [UNIFESP]; Teixeira, Taiza Stumpp [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)
    The germ cell lineage arises from a progenitor cell population in the epiblast called primordial germ cells (PGC). These cells erase their somatic program and enter a germ cell program after what they migrate to the gonads through the gut endoderm and their dorsal mesentery. During their migration and just after they enter the gonadal ridges, the male PGC proliferate for a short period and then enter in quiescence. These cells are then called gonocytes. Because gonocytes are quiescent cells, very few studies about them are available, so that the mechanisms of the quiescence establishment and regulation are not well known. Studies by our group showed that rat gonocytes are mitotic until 18 days post coitum (dpc) and enter in quiescence at 19dpc. During this period, they stay arrested in G0/G1 and no death or proliferation is observed. Our group also showed very abundant and intense caspase-3 (Casp3) labeling in the cytoplasm of all gonocytes, suggesting that this enzyme may play a non-apoptotic role during rat gonocyte quiescence. In addition, there is evidence that Casp3 cleaves the pluripotent marker NANOG in embryonic stem cells, which is also expressed by the male germ cells. Aim: The aim of this study was to investigate if Casp3 and NANOG interact and whether the inhibition of Casp3 activity leads to alterations of the expression of cell cycle genes in the quiescent gonocyte. Methods: For this, 19dpc testis were collected, dissociated and the isolated gonocytes were incubated in vitro with Casp3 inhibitor for 24h. The expression of cell cycle, apoptosis and pluripotency markers was analyzed by qPCR. Gonocyte viability was investigated by propidium iodide and flow cytometry. Protein extraction was performed from whole 19dpc testes for the investigation of Casp3 and NANOG protein expression. Results: The analysis of gonocyte viability showed that the cultures treated with Casp3 inhibitor presented more dead gonocytes than the control cultures. The inhibition of Casp3 also lead to an increase of Pcna expression as well as a decrease of p21cip, p27kip, Bcl2 and Kras expression. When compared with gonocytes that were not maintained in culture for 24h, the cultured gonocytes showed an increase of p21cip expression in control and treated cultures and of p27kip in the control cultures. The expression of Sox2 and Nanog was not detected in the cultured gonocytes, although Sox2 have been detected in non-cultured gonocytes. Casp3 protein was detected in 19dpc testis extract, although NANOG protein was not detected. Conclusion: The results obtained here lead us to conclude that Casp3 inhibition affects the expression of classical cell cycle markers, suggesting that this enzyme plays a non-apoptotic role in male germ cell development by controlling the cell cycle.