Efeito das estações do ano nas alterações celulares de oócitos e embriões da espécie ovina

Abstract

This study aimed to determine the effect of seasons on nuclear maturation in vitro and inductions of cell death by apoptosis in sheep oocytes as well as the effect on the ability of develop from oocytes in vitro sheep embryos. The ewes ovaries during the dry season (October to March) and rainy season (April-September) were collected in a slaughterhouse and transported to the Laboratory of Biotechnical Reproduction of UFRPE. The cumulus oophorus complexes (COCs) were collected by the technique of "slicing" of the follicles from 2 to 6 mm in diameter and selected based on morphology. In this stage were performed 12 repetitions, where the COCs were subjected to maturation, and in vitro fertilization. The percentage of cleaved oocytes was determined on day 3 (D-3) and the oocytes that developed to the stages of 8-16 cells (D-4), morula (D-5) and blastocyst (D-8) after fertilization. The blastocysts quality was assessed with the pigment DAPI and the determination of blastomeres positive for apoptosis and DNA fragmentation by TUNEL test. It was found significant difference (P <0.05) during D-3 and D-4 fertilization stages. No significant difference (P>0.05) was observed on morula and blastocyst in vitro maturation phases and DNA fragmentation (TUNEL). In the stage 25 oocytes were placed in maturity basic medium (MBM). After maturation, we determined the stage of maturity with the nuclear dye DAPI, the enzyme activity of group II caspases with reagent PhiPhiLux-G1D2 and DNA fragmentation (TUNEL). In the stages of nuclear maturation stages of germinal vesicle, metaphase I, telophase I and metaphase II showed no significant difference (P> 0.05). Significant difference (P <0.05) in enzyme activity of caspases in vitro matured oocytes. The DNA fragmentation was not significantly different (P> 0.05). Under the conditions observed in this study, the results indicate that there was no effect of season on in vitro nuclear maturation and apoptosis of oocytes as well as on the development of oocytes and in vitro production of sheep embryos.