L-半胱氨酸对绵羊卵巢颗粒细胞增殖、凋亡和类固醇激素分泌的影响OA北大核心CSTPCD

This study was conducted to investigate the effects of L-cysteine(L-Cys)on proliferation,apoptosis and the secretion of steroid hormones(progesterone and estrogen,P4 and E2)in ovine ovarian granulosa cells(GCs).Thirty sexually mature ewes aged between 1-1.5 years with the similar body weight and the same feeding condition were selected,and the fresh ovaries were collected.Then,GCs were collected from the dominant follicles(2-8 mm),and the GCs were randomly divided into 5 groups with 6 replicates per group.GCs were treated with 0,100,300,500 and 700 μmol·L-1 L-Cys for 48 h.The cell proliferation,apoptosis assay and the secretion of steroid hormones were measured using the CCK-8 kit,Annexin V-FITC/PI flow cytometry and ELISA,respectively.Real-time quantitative PCR and Western blot were used to analyze the mRNA and protein expression of the cell proliferation related genes(PCNA,CCND2,CCNB1),apoptosis-related genes(BAX,Caspase-3,Bcl-2)and steroid-secretion-related genes(STAR,3β-HSD,CYP 11 A1,CYP19A1).The results showed that 100,300 and 500 μmol·L-1 L-Cys significantly promoted the cell viability(P＜0.05)and significantly inhibited the apoptotic rate(P＜0.05)of GCs.The addition of 300 µmol·L-1 L-Cys significantly inhibited the secretion of P4(P＜0.05).The mRNA expression of proliferation-related genes(PCNA,CCND2,CCNB1)were upregulated in the 300 μmol·L-1 and 500 μmol·L-1L-Cys groups,while the protein expression of proliferation-related proteins(PCNA,CCND2,CCNB1)were increased in the 300 μmol·L-1 L-Cys group(P＜0.05).The mRNA and protein expressions of apoptosis-related genes(BAX,Caspase-3)were significantly downregulated in 100 µmol'L-1 and 300 μmol'L-1 L-Cys groups(P＜0.05).The mRNA and protein expressions of the steroid-related genes(STAR,3β-HSD)were significantly decreased in 300 μmol·L-1 L-Cys group(P＜0.05),but the mRNA and protein expressions of the steroid-related genes(CYP11A1,CYP19A1)were significantly increased in 100 μmol·L-1 and 300 μmol·L-1 L-Cys groups(P＜0.05).In summary,L-Cys promoted cell proliferation and inhibited the apoptosis of ovine ovarian GCs by upregulating the mRNA and protein expressions of PCNA,CCNB1,CCND2,Bcl-2 and downregulating the mRNA and protein expressions of BAX,Caspase-3;L-Cys inhibited secretion of P4 by downregulating the mRNA and protein expressions of STAR,3β-HSD and upregulating the mRNA and protein expressions of CYP11A1.