基于iTRAQ技术的“平冲降逆方”抑制子宫内膜异位症异位内膜间质细胞迁移和侵袭的分子机制研究

Endometriosis is a complex chronic gynecological disease. In the previous study, we proposed a new pathogenesis of EMs: “adversely ascending qi of thoroughfare vessel and static blood blocking collaterals”, we determined the therapy strategy and its detailed formula “Ping-Chong-Jiang-Ni formula” based on this theory; this therapy strategy has been approved by the State Administration of Traditional Chinese Medicine Key Specialist Collaboration Group. Our previous studies also shown that “Ping-Chong-Jiang-Ni formula (PCJNF)” could suppress cell proliferation, migration and invasion in EESCs, and induce apoptosis of EESCs; furthermore, and the suppressed cell proliferation and increased cell apoptosis was dependent on the activation of JNK signaling pathway; however, the decreased cell migration and invasion treated by PCJNF was independent on JNK signaling pathway. Based on these observed results, the current project will combine iTRAQ technique and bioinformatic methods, search for the differentially expressed proteins and signaling pathway networks associated with EESCs treated with “Ping-Chong-Jiang-Ni formula”; finally we will select a representative signaling pathway and detect its potential role in the inhibitory effect of migration and invasion of EESCs treated with PCJNF in vitro and in vivo. These results will help us to understand better the internal link between “adversely ascending qi of thoroughfare vessel-forming blood stasis” in traditional Chinese medicine and “menstrual blood reverse flow-forming lesions” in modern medicine, and provide novel insights into molecular target therapy for endometriosis. In addition, these results will also provide potential modern explanation of the pathogenesis from the viewpoint of traditional Chinese medicine for “adversely ascending qi of thoroughfare vessel and static blood blocking collaterals” in endometriosis.

子宫内膜异位症是妇科疑难慢病。我们前期研究提出该病的新病机“冲气上逆，瘀血阻络”，据此制定治法及“平冲降逆方”方药，其疗效已通过国家中管局重点专科协作组的验证。我们前期研究已证实：“平冲降逆方”能够抑制子宫内膜异位症间质细胞的增殖、迁移和侵袭，并诱导细胞凋亡；其抑制增殖和诱导凋亡的效应均通过JNK信号通路所介导，而其抑制细胞迁移侵袭的效应与JNK信号无关。本项目拟在前期工作基础上，利用iTRAQ技术和生物信息学方法，筛选出该方与细胞迁移、侵袭相关的差异表达蛋白并构建信号通路网络；综合分析后，选择与其密切相关的代表性信号通路，在细胞和整体动物水平分别验证其介导的抑制细胞迁移和侵袭作用。研究结果将有助于更好理解子宫内膜异位症的中医“冲气上逆—成瘀”与现代医学“经血逆流—成病灶”之间的内在联系，为患者分子靶向治疗提供新思路，也将有助于对子宫内膜异位症“冲气上逆，瘀血阻络”的中医病机进行现代诠释。

上一个国自然已证实：“平冲降逆方”能够抑制子宫内膜异位症间质细胞的增殖、迁移和侵袭，并诱导细胞凋亡；其抑制增殖和诱导凋亡的效应均通过JNK信号通路所介导，而其抑制细胞迁移侵袭的效应与JNK信号无关。.本项目拟在前期工作基础上，利用iTRAQ技术和生物信息学方法，筛选出该方与细胞迁移、侵袭相关的差异表达蛋白并构建信号通路网络；综合分析后，选择与其密切相关的代表性信号通路，并验证其介导的抑制细胞迁移和侵袭作用。.在本项目的资助下，成功建立了内异症异位病灶体外细胞模型，经平冲降逆方含药血清（ZY组）和生理盐水含药血清（NS组）干预后，通过定量蛋白质组学技术，对获得的肽段进行质谱分析及数据库检索，在 humAn中共鉴定到蛋白质4984个；ZY组vs NS组的上调差异表达蛋白质有3个，下调差异表达蛋白质9个。经验证，首次在内异症病灶组织中发现TRIP13蛋白、FER等蛋白的阳性表达，其中TRIP13蛋白是介导内异症迁移和侵袭的关键靶标蛋白。研究结果诠释了EMs中医“冲气上逆-成瘀”与现代医学“经血逆流-成病灶”的内在联系，有望为临床治疗应用提供新思路。