GRK2/Epac1调控肝星状细胞表型转化在肝纤维化形成中的作用及机制

肝星状细胞（HSC）的表型转化是肝纤维化形成的关键事件。课题组前期研究发现，在肝纤维化大鼠模型中，肝脏G蛋白偶联受体激酶2(GRK2)表达降低，Epac1表达升高。然而，GRK2/Epac1在HSC表型转化中的作用及机制尚不清楚。 我们推测GRK2通过磷酸化或其他机制调节Epac1活性，参与了HSC表型转化及纤维化的形成。本项目采用肝纤维化临床标本，分析GRK2、Epac1与纤维化严重程度的关系。体外研究GRK2如何通过磷酸化调节Epac1活性，影响HSC表型转化；体内应用HSC特异性GRK2或Epac1敲除小鼠，比较其与野生型小鼠肝纤维化形成差异。在此基础上，以GRK2/Epac1为靶点，给予激动剂或抑制剂，考察是否可以抑制肝纤维化的发生。本课题的实施将阐明HSC上GRK2/Epac1在肝纤维化形成中的作用机制，为肝纤维化的病理分子机制提供新的依据，也为肝纤维化治疗的新靶点提供实验基础。

Liver fibrosis is a common end-result of a wide variety of chronic liver diseases following various types of injuries. It is often associated with severe morbidity and significant mortality. In liver, the activation of resident hepatic stellate cells (HSC) into myofibroblasts through the acquisition of a contractile, proliferative, inflammatory, and fibrogenic phenotype is a critical event in fibrogenesis. Activated stellate cells/myofibroblasts are the primary source of extracellular matrix (ECM), whose accumulation leads to the progressive replacement of the liver parenchyma by scar. Currently, clinical reports suggested that advanced liver fibrosis is potentially reversible. Thus, it is essential to develop therapeutic strategies to counteract liver fibrosis.. Exchange protein directly activated by cAMP 1 (Epac1) is a more recently discovered effector of cAMP signaling. Epac1 is a guanine nucleotide exchange factor catalyzing the exchange of GDP for GTP for the Ras-like GTPases Rap1 and Rap2, resulting in their activation. Subcellular localization of Epac1 is dynamic and spatiotemporally regulated. Published work showed that the serine-threonine kinase G protein-coupled receptor kinase 2 (GRK2) inhibits Epac1-mediated pain signaling. Specifically, we showed that hepatic fibrosis is associated with a decrease in GRK2 levels and increase in Epac1 levels in fibrotic liver of rats. However, the role of GRK2/Epac1 in modulation of HSC phenotype transformation, and the mechanism via which GRK2 controls Epac1-mediated signaling in hepatic fibrogenesis remained to be identified.. This study was designed to investigate whether GRK2/Epac1 modulates HSC phenotype transformation in hepatic fibrogenesis and its possible mechanism. To that end, we will choose human samples, which are biopsies from chronic hepatitis patients showing different degrees of fibrosis or control liver samples showing normal histology or minimal change, to determine the possible relationship between GRK2, Epac1 expression and the degree of fibrosis. In vitro studies, the molecular mechanism for which GRK2-mediated controlling Epac1 activity in HSC phenotype transformation will be identified. Meanwhile, mice with a stellate cell-specific knockout of GRK2 or Epac1 were used to compare the severity of hepatic fibrosis with wide type mice in vivo. Furthermore, effect of targeting GRK2/Epac1 on liver fibrosis animal model was assessed to explore the underlying possibilities of therapeutics. Altogether, the present studies will be helpful to discover some new mechanisms underlying the liver fibrosis, and develop a novel therapy target for treatment of hepatic fibrosis.