Tweak-Fn14信号通路促进胆汁淤积肝脏炎性损伤的分子机制研究

Multiple pathologies i.e gallstone obstruction of the bile duct, biliary atresia, pancreas tumors and drug toxicity can cause persisting cholestasis which can lead to liver failure, fibrosis, cirrhosis and death. The accumulation of toxic bile acids and persisting inflammatory liver injury contribute to the progress of cholestasis. However, the molecular mechanism of the liver injury caused by inflammation on cholestasis remains unclear. In our preliminary experiments, we first described that Tweak-Fn14 pathway was activated in human obstructive cholestasis. Meanwhile, the elevated proinflammatory cytokines IL-6 and IL-8 were also observed in human obstructive cholestatic liver, measuring by the gene microarrays. These indicate that Tweank-Fn14 pathway may promote inflammatory liver injury on cholestasis, considering that Tweak-Fn14 pathway mediates the inflammation in rheumatoid arthritis. Moreover, we also found that P38/MAPK was activated in human obstructive cholestatic livers. The phosphorylation of P38 and the production of IL-6 and IL-8 were markedly reduced in HepG2 cells stably infected with Fn14 siRNA virus. All of these results trigger us to formulate a hypothesis: The activation of Tweak-Fn14 pathway stimulates proinflammatory cytokines production by activating P38/MAPK signaling, contributing to the inflammatory liver injury in human obstructive cholestasis. In order to prove this hypothesis, we use different molecular biological techniques, such as Fn14 knockout mice, TaqMan qPCR, western-blot, and human primary hepatocytes culture to study the functional role of Tweak-Fn14 pathway and clarify its molecular mechanism in human obstructive cholestasis, which will be promising for a clue of the clinical therapy and understanding the cholestasis.

胆汁淤积是多因素所致肝损害的常见并发症，易发展为肝纤维化、肝衰竭。肝脏炎性损伤是胆汁淤积发生、发展的关键因素。但是，胆汁淤积肝脏炎性损伤的分子机制尚不明确。我们首次发现：Tweak-Fn14通路在人胆汁淤积肝脏被激活；基因芯片技术也发现炎症因子IL-6等表达增高。而文献报道，Tweak-Fn14在关节炎等可调控炎症反应，这提示该通路在胆汁淤积下也可能促进肝脏炎性损伤。预实验后发现：P38/MAPK在人於胆肝脏被活化，而Fn14 siRNA-HepG2单克隆细胞株中P38/MAPK活性和IL-6等表达均降低。故我们提出假设：胆汁淤积下Tweak-Fn14的活化可激活P38/MAPK信号通路上调肝脏炎症因子表达，从而促进淤胆肝脏的炎性损伤。本课题拟运用基因敲除、原代肝细胞等技术，明确胆汁淤积下Tweak-Fn14通路的功能和其下游调控的分子机制。这将为胆汁淤积临床治疗提供新的治疗靶点和理论依据

胆汁淤积症是一种多因素所致肝损害的常见并发症，慢性持续性的胆汁淤积易造成肝纤维化、肝硬化、肝衰竭和终末期肝病，而肝脏炎性损伤是胆汁淤积发生、发展的关键因素。基于我们前期发现Tweak-Fn14通路在人胆汁淤积肝脏激活的现象，本项目进一步采用小鼠基因敲除、原代肝细胞、电镜等技术，并在临床胆汁淤积和胆汁淤积动物模型中证实Tweak-Fn14 通路在胆汁淤积下存在活化，并揭示胆汁淤积下Tweak-Fn14 的活化可激活NF-kB/ NLRP3 /caspase1/GSDMD信号通路，通过上调肝脏炎症诱导的焦亡，从而促进淤胆肝脏的炎性损伤，同时采用基因敲除小鼠模型进一步证实Fn14 基因缺失减轻肝脏损伤生化指标和减轻焦亡改善肝脏炎症反应和急性肝损伤，对梗阻性胆汁淤积具有潜在的治疗价值。