mRNA expression profile of the TNF-alpha system in LH-induced bovine preovulatory follicles and effects of TNF-alpha on gene expression, ultrastructure and expansion of cumulus-oocyte complexes cultured in vitro

摘要

This study evaluated (1) the effects of in vivo GnRH treatment on mRNA expression of TNF-alpha system (TNF-alpha, TNFR1 and TNFR2) in granulosa cells of bovine preovulatory follicles, (2) the in vitro influence of gonadotropins on mRNA expression of TNF-alpha system in cultured cumulus cells, (3) the protein expression of the TNF-a system in late antral follicles and, (4) the influence of TNF-alpha on cumulus cells expansion, ultrastructure and on expression of HAS2, CASP3 and CASP6 in follicular cells cultured for 24 h. An increased expression of TNF-alpha and TNFR1 was observed after 3, 6 and 12 h of GnRH treatment when compared to 0 and 24h. Higher TNFR2 mRNA levels were observed 3, 6 and 12 h after GnRH, when compared to 0 and 24 h. Proteins of TNF-a system were also expressed in late antral follicles. In vitro, TNF-alpha did not affect cumulus cells expansion, but reduced the HAS2, CASP3 and CASP6 mRNA levels in cumulus cells after 12 h. After 24 h of culture, TNF-alpha increased the mRNA levels for CASP6 in mural granulosa cells, while the TNF-alpha, TNFR1 and TNFR2 mRNA levels were increased in cumulus-oocyte complexes (COCs) cultured for 12 h with gonadotropins, but not after 24 h. Ultrastructural analysis confirmed the integrity of COCs cultured in presence of TNF-alpha. In conclusion, TNF-alpha system members are present in bovine antral follicles and expression of TNF-alpha is influenced by gonadotropins in vivo and in vitro. In vitro, TNF-alpha maintained cumulus cells ultrastructure during COC culture. (C) 2016 Elsevier Inc. All rights reserved.