P2X7受体介导的caspase-1/NLRP3炎症小体活化调控酒精性肝病的作用机制研究

Alcoholic liver disease (ALD) is a result of long-term heavy drinking. There are currently few effective medications available to treat ALD, although researchers are investigating various anti-steatosis and antifibrotic drugs. ALD is mainly characterized by inflammation, excessive accumulation of lipid and activation of hepatic stellate cells (HSCs). P2X7R-mediated inflmmasome signaling plays a critical role in ALD pathogenesis, indicating that P2X7R-inflmmasome may be a new targeting for ALD treatment. We found that P2X7R antagonist reduced the accumulation of lipid in chronic alcohol-feeding mice with hepatic steatosis via suppressing inflammasome activation, and also inhibited inflammasome-mediated activation of HSCs primed by LPS plus ATP. Therefore, we hypothesis P2X7R blockade will inhibit NLRP3 inflammasome activation by blocking assembly of the inflammasome and suppress cellular crosstalk among kupffer cells-hepatocytes-HSCs, eventually reverse inflammatory and fibrotic changes in alcoholic liver fibrosis. Herein, by applying gene knock-down or over-expression cell lines and gene knockout mice, we will investigate the underlying mechanisms of P2X7R-TLR4-mediated inflammasome activation in ALD, and then elucidate cell type-specific role of inflammasome in ALD pathogenesis using cell co-culture system. Thus, this study will provide experimental evidence and therapeutical support for the treatment of ALD.

酒精性肝病(ALD)是长期大量饮酒所致的肝脏疾病，目前尚无特效治疗药物。肝脏内炎症-肝细胞脂肪过度蓄积-肝星状细胞活化是ALD发生发展的关键环节。P2X7R介导的炎症小体信号通路对调控ALD发挥重要作用，为ALD的治疗提供新的干预靶点。我们发现P2X7R拮抗剂通过炎症小体减轻酒精性脂肪肝小鼠的肝脏脂肪蓄积，且基于炎症小体抑制受LPS/ATP联合刺激的肝星状细胞活化。由此提出，若基于P2X7R抑制caspase-1/NLRP3炎症小体的活化，进而切断枯否细胞-肝星状细胞-肝细胞间交叉对话，将改善肝脏炎症并逆转酒精性肝纤维化。基于此，本研究拟利用基因过表达和RNA干扰技术及基因敲除小鼠，阐明P2X7R-TLR4-炎症小体调控酒精性肝病的分子机制；采用共培养体外模型，明确P2X7R-炎症小体信号通路控制酒精性脂肪肝-肝纤维化进程的关键靶细胞及其关键位点，为ALD的防治提供新靶标与理论依据。

酒精性肝纤维化是酒精性肝病向酒精性肝硬化发展的必经阶段，逆转肝纤维化可以有效阻断酒精性肝病恶化为肝硬化，因此，控制肝纤维化进程是防治酒精性肝病的关键环节。肝脏内炎症-肝细胞脂肪过度蓄积-肝星状细胞活化是ALD发生发展的关键环节。P2X7R介导的炎症小体信号通路对调控ALD发挥重要作用，为ALD的治疗提供新的干预靶点。本课题通过体内急性和慢性酒精性脂肪肝动物模型与硫代乙酰胺肝纤维化模型，从体内角度深入研究TLR4-P2X7R介导的炎症小体的活化细胞特异性的调控酒精性肝病的分子机制。另外，采用肝细胞、肝星状细胞或巨噬细胞单独培养，探讨炎性刺激对各肝脏细胞类群内调控P2X7R-炎症小体信号通路的分子机制，然后，巨噬细胞与肝细胞或肝星状细胞共同培养，更好的模拟体内实际作用机制，进一步阐明P2X7R-炎症小体信号通路是枯否细胞-肝细胞-肝星状细胞交叉对话调控酒精性脂肪肝-酒精性肝纤维化的共同关键信号网络。结合体内、体外实验明确P2X7R-炎症小体信号通路为控制肝脏炎症-肝细胞脂肪蓄积-肝星状细胞激活的多靶向调控网络的关键切入点。本研究以P2X7R和TLR4作为研究切入点，确认了P2X7R-炎症小体信号通路与TLR4-NF-κB-炎症小体信号通路的交叉对话及其关键位点，明确活化炎症小体的P2X7R-TLR4信号交叉对话的分子机制，阐明P2X7R-炎症小体信号通路是控制炎症-肝细胞脂肪蓄积-肝星状细胞激活的多靶向调控网络的关键位点，基于P2X7R介导的炎症小体活化调控酒精性肝病发生发展的分子机制方面取得重大突破，为相关酒精性肝病的防治提供新策略与新靶标。