Tese

Efeito do cortisol na produção in vitro de embriões bovinos

The aim of this thesis was to evaluate the effect of cortisol in oocyte maturation and bovine embryo culture. In Chapter 1 we analyzed the distribution and location of glucocorticoid receptor (GR) by immunocytochemistry in oocytes, cumulus cells, embryonic cells 2-4, 8-16 cell, morula and blastocyst...

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Autor principal: ALMEIDA, Nathalia Nogueira da Costa de
Grau: Tese
Idioma: por
Publicado em: Universidade Federal do Pará 2017
Assuntos:
Acesso em linha: http://repositorio.ufpa.br/jspui/handle/2011/8330
Resumo:
The aim of this thesis was to evaluate the effect of cortisol in oocyte maturation and bovine embryo culture. In Chapter 1 we analyzed the distribution and location of glucocorticoid receptor (GR) by immunocytochemistry in oocytes, cumulus cells, embryonic cells 2-4, 8-16 cell, morula and blastocyst, and was also verified the presence of mRNA (qualitative RT-PCR) for GR in the stages. The results showed that the GR is present in all cells analyzed. In order to verify the functionality GR in preimplantation embryo development, translation of mRNA for GR zygotes was silenced in the RNAi technique, and subsequent embryo development was analyzed. The embryonic development decreased (P <0.05) after silencing of GR mRNA. The presence of GR oocyte and the cumulus cells indicates that these cells are sensitive to the use of CG. Given this, in Chapter 2, the effects of different cortisol concentrations during in vitro maturation (IVM) of bovine oocytes and embryonic development, apoptosis rate and gene expression (NRF1, COX, TFAM, GLUT1, FASN and HSP70) were analiyzed. The concentrations of cortisol used were 0.01, 0.1 and 1 mg / mL . There was no statistical difference in the number of cells and cleavage rate, but the concentration of 0.1 mg/ml of cortisol increased the blastocyst rate when compared to the control group (without cortisol in IVM) (41 ± 10 versus 21 ± 1.2; p <0.05, respectively). The rate of apoptosis and gene expression in oocytes, cumulus cells and blastocysts was only assessed at a concentration of 0.1 mg / ml of cortisol. There was no statistical difference in the apoptotic index, and not with respect to gene expression in oocytes and cumulus cells for COX genes, NRF1, HSP70 and FASN (p> 0, 05). Regarding embryonic gene expression, only the measurements relative mRNA FASN, GLUT1 and HSP70 were increased in blastocysts treated with 0.1 g/ml during IVM when compared to embryos of the Control group (p <0.05), the other genes showed no change (p> 0.05). In Chapter 3, we investigated the use of cortisol during in vitro culture (IVC) of bovine embryos. As in Chapter 1 was identified GR in all embryonic stages and when the GR expression was silenced and embryonic development was impaired. Thus, in the experiment three different concentrations (0.01, 0.1 and 1 mg / mL) of cortisol in IVC and embryonic development was evaluated same parameters of Chapter 2. There was no statistical difference in the embryos treated with cortisol in different concentrations when compared to the control for the parameters analyzed (p> 0.05). The concentration of 0.1 mg/ml was chosen to evaluate other parameters of embryo quality. Thus, IVC embryos with or without 0.1 mg/mL of cortisol, were analyzed for apoptosis rate and gene expression, not being statistically significant difference in any of the analyzes (p> 0.05). After these studies conclude that oocytes and embryos are responsive to GC, and the addition of cortisol in IVM improves oocyte competence, but the supplementation of IVC with cortisol may not have influence on embryo development.