虎杖调控miR-200a介导Keap1-Nrf2通路干预果糖诱导NAFLD机制研究

Non-alcoholic fatty liver disease (NAFLD) is increasingly serious. Lack of understanding of the key link for NAFLD pathogenesis, so far there are no effective therapeutic measures. In the clinic of TCM, Rhizoma Polygoni Cuspidati has its special advantage for the treatment of NAFLD, however, its underlying molecular mechanisms are incompletely understood. Fructose-induced NAFLD is associated with oxidative stress. miR-200a activates antioxidant transcription factor Nrf2 by mediating Keap1. Our previous studies found that fructose induced hepatocyte ROS-TXNIP oxidantive stress pathway to mediate inflammatory signaling and lipid metabolism disorder, and down-regulated nuclear Nrf2 protein levels. Moreover, polydatin, a main constituent of Rhizoma Polygoni Cuspidati, was preliminarily confirmed to restore fructose-induced NAFLD, as well as miR-200a down-regulation, oxidative stress and inflammation. This project will continue to investigate new way that miR-200a down-expression mediate Keap1-Nrf2 pathway to suppress antioxidant enzymes-related gene expression in hepatocytes, which, in turn, induce ROS-TXNIP and subsequently to cause inflammation and lipid metabolism disorder，as well as an important role of miR-200a in fructose-induced NAFLD. Furthermore, this project will explore new mechanisms and pharmacological targets for therapeutic intervention of water extracts of Rhizoma Polygoni Cuspidati and polydation on NAFLD via regulating miR-200a-mediated Keap1-Nrf2 pathway in hepatocytes. These results will sopport the hypothesis, and provide theoretical and experimental evidences for the treatment of NAFLD by Rhizoma Polygoni Cuspidati in clinic, as well as the promotion of the translational medicine research.

非酒精性脂肪肝NAFLD危害日益严重，因对其发生发展关键环节缺乏了解，迄今尚无有效治疗措施。虎杖临床治疗NAFLD有优势，但作用机理尚不明确。果糖引起NAFLD与氧化应激密切相关。miR-200a可调控Keap1激活抗氧化应激转录因子Nrf2。我们前期工作发现果糖诱导肝细胞ROS-TXNIP氧化应激通路介导炎症信号激活和脂代谢紊乱，下调Nrf2核蛋白水平；证实虎杖主要成分虎杖苷可逆转果糖诱导NAFLD、miR-200a低表达、氧化应激和炎症。本项目拟进一步研究miR-200a低表达介导Keap1-Nrf2通路抑制抗氧化酶等基因表达，诱导ROS-TXNIP促进炎症和脂代谢紊乱新途径，及miR-200a所起重要作用；探询虎杖水提取物和虎杖苷调控miR-200a表达介导Keap1-Nrf2通路干预NAFLD新机制、新靶点。佐证工作假说，为虎杖治疗NAFLD及相关转化医学研究提供理论和实验基础。

目前对日益危害严重的NAFLD发病关键环节仍缺乏了解，揭示其病理机制和发现有效治疗药物则甚为重要。中药虎杖临床治疗NAFLD有优势，但其作用机制尚需探索。基于前期工作，本项目阐述了高果糖诱导代谢综合征肝脏氧化应激、炎症与脂质蓄积新途径：高果糖下调miR-200a而提高Keap1，破坏Nrf2抗氧化信号通路，促进ROS蓄积与TXNIP过表达以激活NLRP3炎症小体，提高IL-1β水平，干扰脂代谢相关蛋白表达，而致氧化应激、炎症与脂质蓄积。NAFLD可加剧肝纤维化进程，为此本项目拓展研究并发现高果糖促进代谢综合征肝纤维化新机制：高果糖增加肝细胞ZEB1入核以降低miR-200a和miR-203表达，提高Survivin以激活TGF-β1/Smads信号，诱导肝细胞EMT形成，促进肝纤维化。在证实虎杖减轻代谢综合征肝损伤基础上，基于上述新病机的发现，揭示了虎杖苷干预代谢综合征肝脏氧化应激、炎症与脂质蓄积新机制、新靶点：虎杖苷提高miR-200a修复Keap1/Nrf2通路损伤，阻断ROS-TXNIP信号激活NLRP3炎症小体，调节脂代谢相关蛋白表达，从而减轻氧化应激、炎症与脂质蓄积；阐述了虎杖苷改善肝纤维化新途径：虎杖苷抑制肝细胞ZEB1入核以提高miR-200a和miR-203，遏制Survivin激活TGF-β1/Smads信号，逆转肝细胞EMT而缓解肝纤维化。这些结果佐证了本项目所提出的工作假说，为虎杖治疗NAFLD与肝纤维化及其相关转化医学研究提供理论和实验基础。发表研究论文5篇，受邀撰写综述2篇。负责人入选Elsevier2016-2018年中国高被引学者榜单(“药理、毒理和药剂学”项)。部分研究成果获2019年度高等学校科学研究优秀成果奖（科学技术）自然科学一等奖。