基于自噬-炎症小体通路探讨中药菝葜治疗慢性盆腔炎的分子作用机制

Chronic pelvic inflammatory disease (CPID) is a common disease that seriously threatens the health of women, but the mechanism is unknown and there is no specific drug treatment. Inflammatory response is a key link in the development of CPID. It is regulated by autophagy related factors to form the autophagic inflammatory corpuscle inflammatory feedback axis. But its mechanism in CPID has not been reported. Traditional Chinese medicine Smilax has a good therapeutic effect on CPID, the previous studies have found that VPS34 and NLRP3 related to autophagy - inflammasome signaling pathway and the treatment mechanism of Smilax, there may be a potential target for the treatment of CDIP, and it is also an important breakthrough to clarify the therapeutic effect of active ingredients of traditional Chinese medicine. This project starting from the experiments in vivo and in vitro, verify the therapeutic effect of traditional Chinese medicine by endometrial cells inflammation model and CPID rat model, to test the influence on autophagy - inflammasome signaling pathway. Besides, The CRISPR/Cas9 technique is used to verify the VPS34 gene in Chinese medicine to investigate the molecular mechanisms of autophagy - inflammasome pathway and the role of traditional Chinese medicine therapy in CPID.

慢性盆腔炎疾病（CPID）是严重威胁女性健康的常见病，但机制不明，尚无特异性药物治疗。炎症异常反应是CPID发展的核心环节，受自噬相关因子调节形成“自噬-炎症小体-炎症反馈轴”，其在CPID中的作用机制尚无相关报道。中药菝葜对CPID有良好的治疗作用，课题组前期研究发现，VPS34与NLRP3相关的自噬-炎症小体信号通路与菝葜的治疗作用密切相关，有可能是治疗CDIP的潜在靶点群，也是阐明菝葜活性成分治疗作用的重要突破口。本课题拟从体内和体外实验出发，验证菝葜对CPID模型大鼠、子宫内膜细胞炎症模型的治疗作用及对自噬-炎症小体信号通路的影响，同时采用CRISPR/Cas9技术反向验证VPS34自噬基因在中药作用中的地位；探讨自噬-炎症小体通路在CPID中的作用及中药治疗的分子机制。

慢性盆腔炎可能引起输卵管损伤、盆腔粘连及纤维化，是女性生殖道病变引发后遗症的关键因素，且发病率呈逐年上升趋势，严重影响人民生活质量及心理健康，目前尚无特异性治疗药物。慢性盆腔炎的“单方成药”治疗药物菝葜，其总提物已开发为金刚藤胶囊作为临床治疗药物，课题组前期研究发现，菝葜黄酮提取物（FSCR）对相较金刚藤胶囊，在动物模型中表现出对CPID有更好的治疗作用。本项目在此基础上深入研究自噬-NLRP3炎症小体信号通路在CPID发生发展和治疗中的作用及调控机制。本项目通过CPID动物模型和细胞模型发现：① FSCR驱动CPID模型大鼠子宫内膜组织巨噬细胞重编程。② FSCR促进CPID模型大鼠子宫内膜组织自噬-NLRP3炎症小体信号通路恢复平衡。③ 分子对接模拟发现FCSR中五种黄酮类成分均与VPS34具有较好的结合潜力。④ 大鼠子宫内膜原位敲降VPS34，CPID模型大鼠的子宫内膜病理损伤更为严重，提示自噬-NLRP3炎症小体信号通路可能是CPID发生发展和治疗的突破点。⑤ 体外模型研究发现，敲降VPS34，FSCR抑制原代大鼠子宫内膜细胞炎症因子释放作用消失。综上所述，本项目首次研究NLRP3炎症小体信号通路和VPS34介导的自噬传导通路在CPID发生发展中的作用机理，为CPID防治提供依据；首次发现VPS34是菝葜提取物调控自噬-NLRP3炎症小体信号通路治疗CPID的潜在靶点，为研发新的CPID治疗药物奠定基础。