基于DNA甲基化调控GATA转录因子活性研究內异方改善子宫内膜异位症炎症微环境的作用机制

Endometriosis (EM) is a major research topic in the field of gynecology, of which the incidence and recurrence rate are very high. However, its pathogenesis is not clear so far. Inflammatory response provides microenvironment for lesions, which has been proved to be the major factor to promote the development of EM. Previous studies have found that the abnormality of DNA methylation, which regulated the expression of GATA-2/6, is the important reason for inflammation response in EM. On the other side, we observed that Chinese medicine NeiYiFang could effectively improve the inflammation microenvironment and suppress the development of EM. However, the upstream regulatory mechanism of DNA methylationon GATA-2/6 promoters and the working mechanism of NeiYiFang are not clear. According to the results of bioinformatics, NeiYiFang shows the activity of epigenetic regulation. Thus we propose a hypothesis: NeiYiFang could correct the abnormality of DNA methylation on GATA-2/6 promoters to improve the inflammation microenvironment and suppress the development of EM. In this study, we plan to establish the EM rat models and endometrial stromal cells to find out the regulatory mechanism of DNA methylationon GATA-2/6 promoters and the working mechanism of NeiYiFang by using siRNA, MSP, flow cytometry and other molecular biology techniques. The research will provide new thoughts for further research on the pathogenesis of EM and the theoretical basis for clinical application of Chinese medicine NeiYiFang.

子宫内膜异位症（EM）发病率及复发率高且发病机制不明，是妇科领域重大研究课题。炎症反应为EM病灶组织提供微环境，是促进EM发生发展的重要因素。前期研究发现DNA甲基化异常调控GATA-2/6转录因子表达是EM产生炎症反应的重要原因，而中药内异方可有效改善病灶组织炎症微环境，抑制EM发生发展。但GATA启动子DNA甲基化异常修饰的上游调控机制以及内异方改善炎症反应的作用机制尚不明确。生物信息学结果显示，内异方具有表观遗传学调控活性。由此我们提出假说：中药內异方可通过纠正GATA-2/6启动子DNA甲基化紊乱，从而改善EM炎症微环境达到治疗目的。本项目拟建立大鼠EM模型以及原代培养内膜基质细胞，采用siRNA、MSP、流式细胞等技术，探索GATA-2/6启动子DNA甲基化紊乱的调控模式，明确内异方改善炎症微环境的作用机制，为深入研究EM的发病机理提供新思维，同时为內异方临床应用提供理论依据。

子宫内膜异位症（endometriosis, EM）是一种子宫内膜腺体和基质种植生长于子宫腔以外的疾病，具有症状多样性、病变广泛性、侵袭性、易复发性等特点，被认为是一种难治性疾病。因此临床迫切需要副作用小、可长期服用、有效治疗EM的新药。本课题组长期致力于中医药治疗 EM 的探索，内异方为治疗EM 的临床经验方，临床取得的疗效较好，本研究基于DNA甲基化探讨内异方治疗子宫内膜异位症机制研究。结果表明:1)内异方治疗EM在缓解EM相关疼痛、缩小子宫内膜异位囊肿、降低CA125水平、改善生活质量等方面具有良好的疗效，整体有效率高且安全性好，不良反应较少。2)EM异位灶组织存在独特的表观印迹，DNA甲基化调控可能是EM的发生发展机制之一。影响的基因及信号通路（激素相关通路、免疫炎症相关通路、凋亡相关通路等），可以作为我们后续研究内异方作用机制的切入点。3)内异方可以有效抑制异位灶组织中GATA-6 表达异常导致的炎症反应，抑制炎症因子生成及相关信号通路的启动，多靶点、整体改善盆腔炎症微环境，有效抑制异位灶的生长，从而治疗EM。4)内异方药物血清对异位子宫内膜基质细胞有促凋亡、抗增殖的药理学效应，其机制可能是通过抑制PI3K/AKT/GSK-3β凋亡信号通路实现。5)GATA6在EM异位灶中过表达，又可能是作为PI3K/AKT/GSK-3β通路的下游靶蛋白，参与转录调控，在EM发生发展中可能起抑制凋亡、促进细胞增殖的作用。