肥胖基因产物瘦素调控肝星状细胞中独特信号分子GIV/Girdin表达的分子机制及其在瘦素促肝纤维化中的作用

Obesity patients are often accompanied by hyperleptinemia and more apt to develop liver fibrosis. Accumulating evidence indicate that leptin is closely associated with liver fibrosis.Hepatic stellate cell (HSC) activation is the key step during the process of liver fibrosis.HSC activation is based on the alterations in the expressions of many genes,which are induced by a various of factors through the corresponding signaling pathways.GIV (Gα-Interacting Vesicle-associated protein,also known as Girdin) is a novel unique transducer.It serves as a common intermediate of a variety of signals pathways including two major classes of receptor-mediated signaling pathways, namely,receptor tyrosine kinase-induced signaling pathways and G protein-coupled receptor-induced signaling pathways.GIV/Girdin is a central hub for pro-fibrogenic signalling networks in HSCs.GIV mRNA was virtually undetectable in normal livers,but elevated 175 fold in those with cirrhosis. GIV promotes HSC activation and liver fibrosis, and correlates with human liver fibrosois. Only recently, we found that leptin significantly induced GIV expression in HSCs in vitro and in vivo. Based on these clues, the present research program aimed to elucidate the complex mechanisms underlying leptin regulation of GIV expression in HSCs and demonstrate the role of GIV in HSC activatin and liver fibrosis induced by leptin in vitro and in vivo (ob/ob mice) by using techniques for protein detection,plasmid construction, promoter mutation, chromatin immune precipitation and other techniques.The results from the researches may have potential implications for clarifying the mechanisms of liver fibrogenesis associated with hyperleptinemia such as in obesity patients.

肥胖症患者常具血清高浓度瘦素及高肝纤维化发生率。肥胖基因产物瘦素与肝纤维化关联。肝星状细胞(HSC)的激活为肝纤维化发生关键，其基础是肝受损产生各种因子作用于HSC受体，通过信号途径改变HSC基因表达。GIV(运输泡相关Gα结合蛋白，也称Girdin)为新独特信号传导分子，其独特在于能介导细胞主要受体包括G蛋白偶联受体及酪氨酸激酶受体下游事件，成为多种多样胞外因子下游通路公共平台，HSC中GIV在诱导肝纤维化相关信号网络中扮演轴心角色。肝硬化患者肝GIV mRNA上升百倍且促HSC激活，与人类肝纤维化密切相关。申请者发现瘦素显著促HSC中GIV表达。该项目旨在据以上基础，通过HSC培养及瘦素缺失的肥胖鼠，基因表达检测，质粒构建，点突变，ChIP等技术，揭示仍未知的瘦素调控HSC中GIV表达机制及GIV在瘦素促HSC激活及肝纤维化中的意义，项目益于阐明肥胖症患者高肝纤维化发生率分子机制。

肥胖症患者易于肝纤维化发生的倾向且常具有高血清瘦素浓度。研究已表明瘦素与肝纤维化关联，肝纤维化受多种受体参与介导发生。GIV (Gα-Interacting Vesicle-associated protein)是一种多模块信号通路转导分子，也是介导Gi控制多种受体下游关键性信号传导通路的鸟嘌呤核苷酸交换因子。该研究的目的是揭示GIV在瘦素导致的肝纤维化中的作用以及分子机制。体内及体外的实验利用了培养激活的肝星状细胞（HSC），无瘦素基因的肥胖小鼠，基因敲低，蛋白表达检测，EMSA 及 ChIP等技术。结果表明在HSCs中瘦素上调GIV，GIV参与瘦素对肝纤维化的促进。其机制：GIV介导瘦素促进金属蛋白酶组织抑制因子TIMP1, 促进两种重要促肝纤维化受体PDGFβ 受体及TGFβ受体。也为瘦素促进Erk1/2， Akt1，及 Smad3信号通路所需。GIV参与瘦素对细胞分裂相关蛋白Cyclin D1，细胞凋亡相关蛋白Caspase-3，基因表达的表观调控分子BrD4， MAT2B，促炎症分子NFkB， 及抑制HSC激活的重要分子PPARgamm的调控。但GIV 降低 Caspase-3 表达在体内不依赖与瘦素。Erk1/2通路及转录因子Egr1 及 c-Jun 与瘦素调控HSC中 GIV表达。瘦素诱导的Erk1/2通路上调Egr1 及 c-Jun活性，促进其在GIV启动子上的结合（– 180 bp 位点 及 – 376 bp位点）。Egr1的敲低减弱瘦素对肝纤维化的促进。在人的肝纤维化标本HSC中GIV 蛋白水平与p-Erk1/2 及 Egr1 同时上调。不平常的信号传导分子GIV与瘦素的促肝纤维化发生联系在一起提示GIV在肥胖症患者的肝纤维化发生中具有意义。研究揭示了肥胖症患者高肝纤维化发生的分子机制。研究结果可有利于治疗血清高瘦素浓度的肥胖症患者肝纤维化的新药设计。