Du Jing, Liang Xiaoyan, Zeng Haitao, Shu Yimin, Yao Shuzhong, Zhu Bo, Zhuang Guanglun
Center for Reproductive Medicine, First Affiliated Hospital of Sun Yat-sen University, Zhongshan Er Road, Guangzhou, Guangdong, China.
Cell Biol Int. 2008 Apr;32(4):469-72. doi: 10.1016/j.cellbi.2007.12.011. Epub 2008 Jan 10.
Polycystic ovary syndrome (PCOS) is associated with a variety of endocrinologic and metabolic abnormalities, with clinical features of hyperandrogenism and hyperandrogenemia. Cytochrome P450 17alpha-hydroxylase/17,20-lyase (CYP17) is critical in androgen biosynthesis, and CYP17 mRNA expression was proven augmented in PCOS theca cells. To demonstrate whether RNA interference (RNAi) could lower the androgen concentration in theca cells, small interfering RNA (siRNA) targeting the CYP17 gene was co-cultured with exogenous CYP17 in HeLa cells and endogenous CYP17 of theca cells. CYP17 gene expression was measured by fluorescent microscopy, flow cytometry and real-time reverse transcription-polymerase chain reaction analysis. Androstenedione and progesterone concentrations were measured by ELISA. RNAi effectively reduced the expression of exogenous CYP17 in HeLa cells by up to 50%. The CYP17 mRNA and androstenedione production of theca cells were slightly, but not significantly, reduced when compared with non-specific siRNA.
多囊卵巢综合征(PCOS)与多种内分泌和代谢异常相关,具有高雄激素血症和高雄激素的临床特征。细胞色素P450 17α-羟化酶/17,20-裂解酶(CYP17)在雄激素生物合成中起关键作用,并且已证实在PCOS卵泡膜细胞中CYP17 mRNA表达增加。为了证明RNA干扰(RNAi)是否能够降低卵泡膜细胞中的雄激素浓度,将靶向CYP17基因的小干扰RNA(siRNA)与HeLa细胞中的外源性CYP17以及卵泡膜细胞的内源性CYP17共同培养。通过荧光显微镜、流式细胞术和实时逆转录-聚合酶链反应分析来测定CYP17基因表达。通过酶联免疫吸附测定法测量雄烯二酮和孕酮浓度。RNAi有效地将HeLa细胞中外源性CYP17的表达降低了多达50%。与非特异性siRNA相比,卵泡膜细胞的CYP17 mRNA和雄烯二酮生成略有减少,但不显著。