Effects of follicle-stimulating hormone, insulin-like growth factor 1, fibroblast growth factor 2, and fibroblast growth factor 9 on sirtuins expression and histone deacetylase activity in bovine granulosa cells.

Granulosa cells (GC) are critical regulators of fertility. During the process of ovarian folliculogenesis, these cells undergo profound changes while producing steroid hormones that are important to control follicular growth, oocyte maturation, and ovulation. Sirtuins are enzymes that regulate several biological processes and have been associated with control of GC function. However, how sirtuins are regulated in GC during ovarian folliculogenesis remains to be unveiled. The present study was designed to investigate effects of hormones that control GC proliferation, differentiation, and steroidogenesis on expression of the seven members of the mammalian sirtuins family (SIRT1-7) and on histone deacetylase activity of nuclear sirtuins (SIRT1, 6, and 7) in GC. Bovine granulosa cells were isolated from small antral follicles (1-5 mm) and were treated with or without follicle-stimulating hormone (FSH), insulin-like growth factor 1 (IGF-1), and fibroblast growth factors 2 (FGF2) and 9 (FGF9). Following treatments, cell proliferation was determined via a cell analyzer, estradiol synthesis and histone deacetylase activity were determined via ELISA, and sirtuins mRNA expression was determined via qPCR. Treatments with FSH and IGF-1 stimulated cell proliferation while addition of FGF2 or FGF9 suppressed estradiol production stimulated by FSH plus IGF-1. In terms of treatments that regulated sirtuins expression in GC, fibroblast growth factors were the most impactful: FGF2 alone increased SIRT1 mRNA expression in comparison to several treatments and increased mRNA abundance of SIRT2 and SIRT7 when added to the combination of FSH and IGF-1; the addition of FGF9 to the combination of FSH and IGF-1 increased mRNA expression of SIRT2, SIRT3, SIRT4, SIRT6, and SIRT7 and increased mRNA expression of SIRT5 in comparison to the negative control group that received no treatment. Also, FGF2 alone increased histone deacetylase activity of sirtuins in comparison to all treatments that contained FSH and/or IGF-1. Furthermore, several correlations were observed between treatments and sirtuins expression and activity, between estradiol or GC numbers and sirtuins expression, and between expression of sirtuins. As FGF2 and FGF9 are considered anti-differentiation factors of GC that stimulate GC proliferation while suppressing estradiol production in combination with FSH and IGF-1, data of this study suggest that sirtuins are associated with control of differentiation of bovine GC.

Effects of asprosin on estradiol and progesterone secretion and proliferation of bovine granulosa cells.

Asprosin is an adipokine synthesized by the white adipose tissue that regulates glucose homeostasis and that has been reported to affect bovine theca cell function and follicular growth, but its role on granulosa cell functions remains to be unveiled. Hence, the objective of this study was to investigate asprosin impacts on granulosa cell steroidogenesis. Bovine granulosa cells from small ovarian follicles were cultured in vitro to investigate the effects of asprosin on cell proliferation, production of steroids, mRNA abundance of genes that encode steroidogenic enzymes and cell cycle regulators, and protein relative abundance of steroidogenic signaling pathways. Asprosin was shown to affect granulosa cell functions in a dose-dependent manner. In the presence of FSH, asprosin enhanced estradiol production and stimulated an increase in mRNA expression of FSHR and CYP19A1 in a dose-dependent manner. In the presence of IGF1, asprosin decreased estradiol production, increased progesterone production, altered PKA relative protein expression, and tended to alter the ratio of p-ERK1/2/total ERK1/2 protein expression in a dose-dependent manner. Furthermore, asprosin increased p-53 gene expression in basal culture conditions and with or without FSH and IGF1. Taken together, findings of this study show that asprosin is a regulator of granulosa cell functions and the effects of asprosin depend on dose and cell culture conditions.