miR-195-5p targeting CLDN6 regulates the proliferation and apoptosis of ovarian granulosa cells in rats with polycystic ovary syndrome
Objective To investigate the regulatory mechanism of microRNA-195-5p(miR-195-5p)/CLDN6 on the proliferation and apoptosis of ovarian granulocyte cells in rats with polycystic ovary syndrome(PCOS).Methods Ten SD rats were divided into normal group and PCOS group with 5 rats in each group.Construct the PCOS model.Isolation and culture of PCOS rat ovarian granulosa cells and divided into Con group,miR-NC group,miR-195-5p group,si-NC group,si-CLDN6 group,miR-195-5p+pcDNA group and miR-195-5p+pcDNA-CLDN6 group.Dual luciferase reporting experiment verified the targeting relationship between miR-195-5p and CLDN6.The proliferative activity of ovarian granulosa cells in PCOS rats was detected by MTT assay.The cloning ability of ovarian granulosa cells in PCOS rats was detected by plate cloning assay.The apoptosis rate of ovarian granulosa cells in PCOS rats was determined by flow cytometry.The expression levels of CLDN6,activated cysteine protease 9(cleaved-caspase 9),activated cysteine protease 3(cleaved-caspase 3),and B-lymphoblastoma 2(Bcl-2)associated X protein(Bax)were detected by Western blot.Results The expression level of miR-195-5p in ovarian granulosa cells of PCOS rats was lower than that of normal rats,and the expression level of CLDN6 was higher then that of normal rat ovarian granulosa cells(P＜0.05).MiR-195-5p can target the expression of CLDN6 protein.Overexpression of miR-195-5p or interference with CLDN6 expression inhibited the proliferation of ovarian granulosa cells in PCOS rats and promoted cell apoptosis(P＜0.05).The apoptosis rate and the expression levels of cleaved-caspase 3,cleaved-caspase 9 and Bax protein were significantly increased(P＜0.05).Up-regulation of CLDN6 partially reversed the effects of overexpression of miR-195-5p on proliferation of ovarian granulosa cells in PCOS rats(P＜0.05).Conclusion MiR-195-5p may regulate the pro-liferation and apoptosis of ovarian granulosa cells in PCOS rats by targeting CLDN6.
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