miR-495-3p对山羊卵巢颗粒细胞功能的影响

旨在探究miR-495-3p对山羊卵巢颗粒细胞功能的影响及作用机制.本研究选取健康的3～4月龄大足黑山羊母羊,收集卵巢颗粒细胞,利用miR-495-3p模拟物(mimics)和抑制物(inhibitor)构建过表达和抑制模型,通过流式细胞术检测细胞凋亡和周期,ELISA分析颗粒细胞的雌二醇(E2)和孕酮(P4)分泌,采...

Effects of miR-495-3p on Ovarian Granulosa Cell Functions in Goat

The objective of this study was to explore the effect of miR-495-3p on the function of goat ovarian granulosa cells and its regulation mechanism.In this study,healthy 3-4 months female Dazu black goats were selected to collect ovarian granulosa cells,miR-495-3p mimics and inhibitor were used to construct overexpression and inhibition cell models.The phenotype of apoptosis and cell cycle were detected by flow cytometry.The secretion of estradiol(E2)and progesterone(P4)in granulosa cells were tested by ELISA kits.The qRT-PCR and Western blot were used to detect mRNA and protein expression levels of genes involved in cell apoptosis,proliferation,cycle and steroid hormone synthesis,respectively.The results showed that miR-495-3p significantly promoted ovarian granulosa cell apoptosis(P<0.01),enhanced BAX expression(P<0.05)and inhibited BCL2 expression(P<0.01)in ovarian granulosa cell.Moreover,overexpression of miR-495-3p inhibited cell proliferation in a time-dependent manner and significantly reduced the expression of PCNA(P<0.05).Meanwhile,overexpression of miR-495-3p significantly increased the percentage of cells at G2/M phase(P<0.05),and significantly reduced the mRNA level of cell cycle-related genes CDK4(P<0.001)and CCND1(P<0.05).Others,inhibition of miR-495-3p significantly reduced the mRNA level of CCNE1 gene(P<0.01).Meanwhile,miR-495-3p could promote the secretion of E2 and P4 in granulosa cells.Overexpression of miR-495-3p significantly increased the mRNA and protein expression levels of 3β-HSD(P<0.001),CYP11A1(P<0.01),CYP19A1(P<0.05).Interference with miR-495-3p significantly inhibited 3β-HSD(P<0.01),promoted CYP11A1 expression(P<0.01).Overall,the above results indicated that miR-495-3p can promote granulosa cell apoptosis,inhibit proliferation,arrest in the G2/M phase of cell cycle,stimulate the secretion of steroid hormones,thereby affect the follicular development of goat.