RNA干扰HAS2基因表达对多囊卵巢综合征大鼠卵巢颗粒细胞生物学特性及免疫因子的影响

目的:探讨透明质酸合成酶2(HAS2)基因在多囊卵巢综合征(PCOS)大鼠表达及RNA干扰其表达对卵巢颗粒细胞活力、凋亡及免疫抑制因子TGF-β1和VEGF表达的影响。方法:建立DHEA致雌性SD大鼠PCOS模型,Western blot检测PCOS大鼠卵巢组织中HAS2的蛋白表达;培养原代PCOS大鼠卵巢颗粒细胞,将合成的靶向抑制HAS2表达的siRNA转染到细胞中,Western blot检测转染效果及转染后48 h各组细胞中免疫抑制因子TGF-β1、VEGF和NF-κB信号通路相关蛋白的表达。CCK8及流式细胞仪分别检测细胞活力和凋亡率。结果:HAS2在PCOS大鼠卵巢组织中高表达,转染si-HAS2的大鼠卵巢颗粒细胞中HAS2的表达显著低于NC组和对照组(P <0. 05);与NC组比较,si-HAS2组细胞活力显著降低,凋亡率增加,TGF-β1、VEGF及NF-κB信号通路p-NF-κB、p-IκBα和下游cyclin D1、survivin的蛋白表达均显著降低(P <0. 05)。结论:HAS2在PCOS大鼠卵巢组织中高表达,抑制大鼠卵巢颗粒细胞中HAS2表达可降低细胞活力,诱导细胞凋亡,下调免疫抑制因子TGF-β1、VEGF及NF-κB信号通路表达。

Effect of HAS2 gene by RNA interference on biological characteristics and inflammatory immune factors of ovarian granulosa cells in rats with polycystic ovary syndrome

Objective: To investigate the expression of HAS2 gene in polycystic ovary syndrome(PCOS) rats and the effect of HAS2 gene by RNA interference on the viability,apoptosis and expression of TGF-β1 and VEGF of ovarian granulosa cells.Methods:The rat PCOS model was established,and the expression of HAS2 protein was detected in PCOS rats by Western blot.Primary PCOS rat ovarian granulosa cells were cultured and siRNA that inhibit the expression of HAS2 was transfected in cells.Western blot were used to detect TGF-β1,VEGF and NF-κB signaling pathway related proteins expression in transfected cells after transfected for 48 h.Cell viability and apoptosis rate were detected by CCK8 and flow cytometry.Results: HAS2 was highly expressed in the ovarian tissue of PCOS rats.The expression of HAS2 in rat ovarian granulosa cells transfected with si-HAS2 was significantly lower than that in NC group and control group(P < 0.05).Compared with NC group,cell viability significantly decreased in si-HAS2 group,apoptosis rate significantly increased.The expression of TGF-β1,VEGF,p-NF-κB,p-IκBα,cyclin D1 and survivin protein were significantly decreased(P <0.05).Conclusion: HAS2 was highly expressed in the ovarian tissue of PCOS rats.Inhibition of HAS2 expression in rat ovarian granulosa cells can reduce cell viability and induce apoptosis,and downregulate the expression of TGF-β1 and VEGF and NF-κB signaling pathway.