基于PI3K/Akt/GSK3β及JAK2/STAT3通路探讨芍药甘草汤治疗PCOS的分子机制

Polycystic ovary syndrome (PCOS) is a complex, common reproductive endocrine diseases in women, unfortunately, there is no ideal treatment drug. Our previous results and systematic reviews and meta-analyses showed that Shaoyao Gancao decoction can decrease androgen level and increase insulin sensitivity, as well as improve reproductive dysfunction. However, the molecular mechanism for Shaoyao Gancao Decoction is not yet clear. Due to close association with insulin resistance and hyperandrogenism, PI3K/Akt/GSK3β and JAK2-STAT3 pathways become the reasonable selection of research of Shaoyao Gancao Decoction which can decrease androgen level and increase insulin sensitivity. It is first time for this study to explore treatment effect of Shaoyao Gancao Decoction on PI3K/Akt/GSK3β and JAK2-STAT3 pathways to investigate its pharmacological mechimism in PCOS, protein and gene expression (mRNA) of PI3K, AKT, GSK3β, STAT3, Cyclin D1, SOCS3 and androgen synthesis enzymes，which are close to PI3K/Akt/GSK3β and JAK2-STAT3 pathways, were detected by Western Blot and RT-PCR. Through these methods, the interaction regulation of Shaoyao Gancao Decoction improving the PI3K/Akt/GSK3β and JAK2-STAT3 pathways were clarify objectively and systematacially, and which can not only evaluate the pharmacological mechimism of Shaoyao Gancao Decoction, but also provide evidence for extended clinical practice of classic formula.

PCOS是影响女性生殖内分泌健康的现代常见疑难疾病，且尚无理想的治疗药物。我们前期实验研究及系统评价荟萃分析结果均显示芍药甘草汤具有较好的降雄增敏，改善生殖障碍的作用，但对于其疗效的分子机制尚不清楚。PI3K/Akt/GSK3β与JAK2-STAT3信号通路与PCOS胰岛素抵抗和高雄激素血症的发生密切相关，以此为着眼点，研究芍药甘草汤降雄增敏的疗效靶点成为理想的选择。本研究首次以GSK3β和STAT3为疗效关键点，探讨芍药甘草汤治疗PCOS的作用机制，应用Western Blot和qRT-PCR方法检测PI3K、AKT、GSK3β、STAT3、Cyclin D1、SOCS3等关键因子以及雄激素合成关键酶的蛋白表达和mRNA表达，从分子水平客观评价芍药甘草汤对GSK3β和STAT3活性的影响及治疗PCOS的作用机制，为拓展经方的临床适应症提供依据。

多囊卵巢综合征（Polycystic ovary syndrome，PCOS）是影响女性生殖内分泌健康的现代常见疑难疾病。PCOS以胰岛素抵抗和高雄激素血症为主要特征，影响着世界6-21%女性患者。由于本病病因和机制尚未完全阐明，亦未研制出理想的治疗药物，给社会和家庭带来了巨大的负担。前期研究发现，芍药甘草汤纠正PCOS效果明确，但对于其疗效的分子机制尚不清楚，严重阻碍了芍药甘草汤临床应用推广和国际认可。因此，亟待开展芍药甘草汤治疗PCOS疗效机制挖掘，为经方的临床应用提供科学依据。本课题围绕芍药甘草汤治疗PCOS的疗效机制，开展了体内和体外两部分研究。体内研究结果显示芍药甘草汤对PCOS大鼠生育功能异常、甾体激素及糖代谢异常具有良好调节作用；明确了芍药甘草汤对 PCOS大鼠卵巢组织的病理改变具有较好的改善作用；发现芍药甘草汤能够上17β-HSD、Cyclin D1、p-STAT3蛋白表达，下调SOCS3、 p-IRS、p-GSK3β、CYP11、CYP17的蛋白表达；上调 PI3K、CYP17、CYP19mRNA 水平，下调 GSK3βmRNA表达水平。体外研究部分探究了芍药甘草汤对 PCOS 大鼠卵巢卵泡膜颗粒细胞的疗效机制，发现芍药甘草汤能够增强p-STAT3、Cyclin D1、PI3K 的蛋白表达，降低SOCS3的蛋白表达；下调17β-HSDmRNA和 PI3KmRNA水平。综合体内和体外研究我们发现芍药甘草汤能够改善PCOS胰岛素抵抗和高雄激素，其作用机制主要与下调雄激素素合成关键酶CYP11、CYP17蛋白表达，增强p-STAT3、PI3K、Cyclin D1的蛋白表达，上调STAT3mRNA水平，抑制SOCS3的蛋白表达有关。该机制的阐明，拓展了芍药甘草汤防治AD的作用机理，提示该方在治疗多囊卵巢综合征方面具有多靶点的作用。