养精种玉汤有效部位对PCOS卵巢局部ERK/c-fos/CYP17的调控

Exposed excess androgen by subcutaneously injecting pregnant rats with testosterone propionate in late gestation to construct a offspring rat model for PCOS. And construct the rat cell models for PCOS synthetizing excess androgen by inducing rat theca cells insulin resistance with low dose of dexamethasone, and by silencing c-fos of granulose cells with RNA interference technology. Thus, this project investigate the cellular and molecular mechanisms of YangJing ZhongYu decoction and its effective-fractions reducing androgen that synthetized in ovary through the key enzymes of androgen synthesis, CYP17, and it`s regulatory factor, c-fos, and MAPK signal pathway. This study will proclaim the effective target ,pathway and process of YangJingZhouYu decoction reducing excess androgen from dynamic balance of signal pathway-gene expression-biological effect .Then, clarifying the material base of YangJingZhouYu decoction reducing excess androgen and the characteristics of its effective-fractions, providing the basis for rational clinical application, and the ideas and methods for modern research of prescriptions. This study will provide the evidence and expand its application in traditional Chinese medicine of active material reducing excess androgen.

采用孕期大鼠雄激素处理，构建子代PCOS产生高雄激素的动物模型，采用产生胰岛素抵抗的大鼠卵泡膜细胞和RNA干扰技术沉默c-fos的大鼠颗粒细胞构建PCOS产生高雄激素的细胞模型，从雄激素生成关键酶CYP17的基因表达及其调控因子c-fos与主要信号转导途径（MAPK），研究养精种玉汤及其主要有效部位降低卵巢源性雄激素生成的细胞与分子机制。从信号通路-基因表达-生物效应动态平衡角度揭示其作用靶点、途径和方式。阐明该方抗雄激素的物质基础，明确各类有效部位作用的特点，为方剂的现代化研究提供思路与方法。为扩大该方应用、挖掘新的抗雄激素生成的中药活性物质群提供实验依据。

本项目按计划完成了养精种玉汤有效部分对卵巢局部雄激素生成机制的研究。本课题成功构建了pGMLV-SC1 RNAi慢病毒载体，并筛选出RNAi有效靶点，成功抑制了卵巢颗粒细胞c-fos的表达，实验表明卵巢颗粒细胞和膜细胞ERK、c-Fos蛋白和mRNA表达下调时，CYP17蛋白和mRNA表达上调，T生成明显升高。提示可能存在ERK/c-Fos/CYP17途径调节卵巢雄激素的生成。进一步研究证实养精种玉汤正丁醇和乙酸乙酯有效部位可通过ERK/c-Fos/CYP17途径降低雄激素的生成。本研究为扩大该方应用、挖掘新的抗雄激素生成的中药活性物质群提供实验依据。