PNPLA3 I148M基因多态性诱导胆固醇稳态失调促进非酒精性脂肪性肝炎进展的分子机制研究

非酒精性脂肪性肝炎(NASH)是肝病中的关键阶段，目前尚无治疗药物，而游离胆固醇是促进NASH的主要脂毒性分子。PNPLA3基因I148M多态性是目前发现的肝病最强遗传决定因素，但其促进NASH进展的分子机制尚不明确。我们通过转基因小鼠NASH模型发现PNPLA3 I148M可以影响肝胆固醇代谢，加重纤维化和炎症表型，提示其促进NASH发展与体内游离胆固醇聚集有关。为此，本项目拟利用PNPLA3 I148M稳定表达细胞系和动物模型，研究PNPLA3 I148M、游离胆固醇聚集和NASH这三者之间的因果关系和内在联系，确定PNPLA3 I148M通过影响胆固醇代谢途径，而诱导肝纤维化和炎症发生的新分子机制，并建立抑制剂筛选模型，寻找NASH治疗的靶向药物。本研究不仅为PNPLA3 I148M促进NASH分子机制研究提供了新的理论基础和研究方向，也为肝病临床诊治提供新的思路和治疗手段。

Nonalcoholic steatohepatitis (NASH) is a critical stage in the development of nonalcoholic fatty liver disease, which can evolve into cirrhosis and hepatocellular carcinoma (HCC). Unfortunately, there are no FDA-approved medications available for patients with NASH-induced advanced fibrosis. It has been reported recently that hepatic free cholesterol is the main toxic molecule contributed to NAFLD pathogenesis..As a strongest genetic risk factor, PNPLA3 I148M polymorphism has been involved in full spectrum of liver diseases ranging from alcoholic liver disease, non-alcoholic liver disease, NASH, cirrhosis, and hepatocellular carcinoma. It’s been confirmed that PNPLA3 I148M increase the risk of NASH in clinical and epidemiology research, but related underlying mechanism is still unclear. In the previous study, we demonstrated that transgenic (Tg) human PNPLA3 I148M mice can led to a significant increase of liver cholesterol and free cholesterol level while a significantly decrease of expression of genes in the cholesterol metabolism pathway when compared with transgenic human PNPLA3 I148I mice under AMLN diet. Moreover, TgPNPLA3 I148M mice can develop a more severe liver fibrosis and inflammation phenotype. Based on our findings, we will conduct the study focusing on the influence of PNPLA3 I148M in the endogenous pathways of cholesterol metabolism and free cholesterol accumulation in hepatic stellate cells (HSC) or Kupffer cells which isolated from a transgenic mouse model of NASH. A comprehensive proteomics and transcriptomics analysis will be performed to find the difference of expression level of genes involved in cholesterol metabolism of HSC and Kupffer cells of PNPLA3 I148M mouse liver. Furthermore, the impact of PNPLA3 I148M on activation and crosstalk between HSC cells and Kupffer will also be studied in this project. Meanwhile, we will establish high throughput inhibitor screening models of targeted PNPLA3 I148M and to find new drugs for NASH treatment. Such studies will not only clarify a new molecular mechanism of PNPLA3 I148M in the pathogenesis of NASH, but also provide strong support for the development of novel anti-NAFLD drugs.