基于Notch-PI3K/Akt/mTOR串话调控探讨辛通畅络法干预FSGS足细胞损伤修复的机制研究

The research is in-depth study of previous National Natural Science Foundation of China project which confirmed that effects of XinTong ChangLuo are exact to repair glomerular filtration barrier on FSGS associated with intervention of expression of TGF-β1, reducing the podo-cyte injury, but the underlying mechanism remains to be explained. TGF-β1 signal crosstalk is involved in podocyte epithelial mesenchymal transdifferentiation (EMT) and the autophagy process. In this study, observes the effect of ShenSu II , carrier of XinTongChangLuo to re-gulate podocyte EMT and autophagy on FSGS rat model by stereology, pathology and molecular biology technique and analyse mechanism of ShenSuII with the regulation of Notch-PI3K/Akt/mTOR signal crosstalk by podocyte single culture and double dispersion system. At the same time, based on the theory of Pungency treat kidney, analyse Shen Su II and its disassembled prescription pharmacodynamic characters, related HPLC fingerprints and quality control system has been established, and select drug compatibility. This research establishes regulation of podocyte function based on the XinTongChangLuo method research system and enrich the basic theories and research progress.

本课题系既往国家自然科学基金项目的纵深研究。前期研究证实辛通畅络法对FSGS肾小球滤过屏障修复具确切作用，其干预机制与调控FSGS大鼠肾小球TGF-β1表达，减轻足细胞损伤有关，但深层机制仍有待阐释。已知TGF-β1相关信号串话参与足细胞上皮-间充质细胞转分化（EMT）及自噬进程，本研究拟采用辛通畅络法载体—肾苏Ⅱ干预FSGS大鼠，运用体视学、病理学、分子生物学等技术，观察辛通畅络法对FSGS足细胞病变早期EMT与自噬的干预效应；利用足细胞单培养与两室弥散体系，分析肾苏Ⅱ对EMT与自噬的干预是否与调控TGF-β1相关串话Notch-PI3K/Akt/mTOR有关，探讨辛通畅络法干预FSGS的深层分子机制。同时，依托辛味治肾理论，分析肾苏Ⅱ及其拆方药效特征，初步建立相关HPLC图谱与质控体系，并筛选优化用药配伍。本研究可为序贯建立基于足细胞功能调控的辛通畅络法科研体系丰富理论内涵与研究基础。

足细胞作为增殖能力有限的终末分化细胞，系肾小球滤过屏障的重要组成部分，对维持肾小球滤过屏障具有重要意义，而足细胞病变则是FSGS的重要病理特征。研究多层次开展辛通畅络法复方肾苏Ⅱ对FSGS足细胞病变早期EMT与自噬进程的干预效应分析与机制阐释研究，以期证实“辛通畅络法通过TGF-β1相关串话Notch-PI3K/Akt/mTOR，调控足细胞损伤早期EMT与自噬，实现损伤修复平衡，阻抑FSGS进展”假说的科学性。研究结果表明，辛通畅络法载体复方肾苏Ⅱ可有效调控足细胞损伤修复，其作用机制与调控EMT及自噬信号通路Notch-PI3K/Akt/mTOR有关。但基于中医学术理论思想指导的辛通畅络法载体复方肾苏Ⅱ拆方优势药物组合筛选工作未实现预期科研设想。建立肾苏Ⅱ高效液相色谱指纹图谱以评价其质量。同时，构建肾苏Ⅱ丹参水溶性成分指纹图谱共有模式，显示14个共有峰，指认黄芩苷特征峰。此外，基于本项目相关内容开展的一系列探索性研究，取得部分进展。初步证实辛通畅络法载体复方肾苏Ⅱ尚可抑制足细胞凋亡与肾间质纤维化进程，完成基于PageRank算法的慢性肾脏病中医药学术经验信息挖掘的尝试。为序贯辛通畅络法系列科学研究的实现奠定了前期基础与技术储备。