基于肝星状细胞自噬及调控通路探讨片仔癀抗肝纤维化的作用机制研究

项目名称： 基于肝星状细胞自噬及调控通路探讨片仔癀抗肝纤维化的作用机制研究

项目编号： No.81503513

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 医药、卫生

项目作者： 郑海音

作者单位： 福建中医药大学

项目金额： 18万元

中文摘要： 肝星状细胞（HSC）活化是各种类型慢性肝损伤发展为肝纤维化的关键环节，而细胞自噬能够调控HSC活化，被认为是抗肝纤维化研究的重要靶点。片仔癀临床上治疗慢性肝损伤疗效明确。前期研究表明片仔癀可明显减轻CCl4肝纤维化模型大鼠肝组织的纤维化；在体外可抑制HSC的增殖活性，并下调miR-155的表达量。本项目拟进一步从动物和细胞实验水平，采用透射电镜、免疫荧光等技术观察片仔癀对体内外HSC活化、细胞自噬的影响，并用免疫组织化学染色、western blot、real-time PCR等方法观察片仔癀对miR-155及下游mTOR/自噬信号通路上关键信号分子表达的调控作用，并构建过表达和抑制表达miR-155的慢病毒载体，建立高表达和低表达miR-155的HSC细胞株进行反向验证，进而探讨片仔癀治疗肝纤维化的分子靶点及调控机制，为片仔癀临床用于抗肝纤维化治疗奠定理论和实验基础。

中文关键词： 肝纤维化；肝星状细胞；自噬；微小RNA；片仔癀

英文摘要： Activation of hepatic stellate cells (HSC) is the key for the development of various types of chronic liver injury to liver fibrosis. In addition, autophagy can regulate HSC activation, so it becomes the important target for anti-fibrosis research. Pien Tze Huang has a protective effect on chronic liver injury. Our previous studies have shown that Pien Tze Huang can reduce hepatic fibrosis in rats induced by carbon tetrachloride, inhibit the proliferation of HSC and down-regulate the expression of miR-155 in vitro. This study intends to further from animal and cell experiment level, using transmission electron microscopy and immunofluorescence techniques to observe the effect of Pien Tze Huang on HSC activation and cell autophagy in vivo and in vitro, and using immunohistochemistry, Western blot, real-time PCR and other methods to observe the regulatory effect of Pien Tze Huang on the expression of miR-155 and key signaling molecules in downstream mTOR/autophagy signaling pathway. Then construction of miR-155 overexpression and knockdown lentiviral expression vector to establish HSC cell lines of high and low miR-155 expression is used to reverse verification, and then to validate the targets and regulation mechanism of Pien Tze Huang. The study will establish a theoretic and experimental foundations for using Pien Tze Huang against liver fibrosis.

英文关键词： hepatic fibrosis;hepatic stellate cell;autophagy;microRNA;Pien Tze Huang