SCD1通过AMPK通路调节脂质自噬在NAFLD中的作用及其机制研究

Abstract：.Abnormal intrahepatic lipis accumulation is the basis of the development of nonalcoholic fatty liver disease. SCD1 involved in high-fat induced hepatic stestosis, however other current researches about SCD1 focused on the regulation of triglyceride synthesis as well as fatty acid beta-oxidation. Increasing evidence indicates that lilophagy, as a selective way of autophagy, is thought to be a new mechanism in regulating lipid metabolism and shows benificial effects in NAFLD. AMPK , as a cellular energy sensor, plays an important role in the regulation of autophagy, and its activity is controled by SCD1. Our own previous study reavled that the inhibition of SCD1 induced-autophagy lead to cell death in human hepatocellular carcinoma through activation of the AMPK signaling pathway. Thus, we put forward a hypothesis that down-regulation of SCD1 in hepatocytes may improve the development of NAFLD by induced lipophagy, which is mediated by activation of AMPK singaling pathway. To explore the role of SCD1-AMPK singaling pathway in the pathygenesis of NAFLD through the induction of lipophagy, we will build NAFLD model of hepatocytes and rats respectively, and regulate the expression of SCD1 and AMPK separately. Once the hypothesis is corroborated, it could become a novel target for treatment for NAFLD.

肝内脂质异常聚集是NAFLD发生发展的基础。SCD1参与高脂诱导下肝细胞脂肪变性，目前相关机制研究主要涉及调控肝细胞TG合成及脂肪酸氧化。脂质自噬被认为是调节细胞脂代谢和脂质沉积的一种新机制，激活肝细胞内脂质自噬，可使肝细胞免受脂质沉积的损伤，在NAFLD中具有保护性效应。AMPK信号通路参与自噬的调控，同时又受SCD1水平的调控。前期研究发现，抑制SCD1可激活 AMPK信号通路，从而增强自噬促进肝癌细胞凋亡。因此，我们推测SCD1可通过AMPK调节脂质自噬参与NAFLD的发生发展，这一科学假设目前国内外尚无报道。本项目拟构建NAFLD细胞及动物模型，通过增强或抑制肝细胞内及体内SCD1、AMPK，探讨SCD1-AMPK信号通路对脂质自噬的调控作用，以及对肝内脂质沉积的影响，为NAFLD的发病机制和临床治疗提供新的理论依据和治疗靶点。

目的和背景：本研究旨在探讨SCD1是否可通过AMPK调节脂质自噬参与NAFLD的发生发展。 主要研究内容：1）抑制或促进SCD1表达，C57BL/6小鼠原代肝细胞内脂质合成及沉积，SCD1、AMPK和自噬相关蛋白表达及自噬小体形成情况2）联合抑制SCD1表达及AMPK磷酸化，C57BL/6小鼠原代肝细胞内脂质合成及沉积、SCD1、AMPK和自噬相关蛋白表达及自噬小体形成情况。3）SCD1抑制剂及高脂饮食联合喂养C57BL/6小鼠后，肝组织脂质沉积及合成、SCD1、AMPK及自噬相关蛋白表达以及自噬小体形成情况。 重要结果及关键数据：1）高脂情况下，脂质自噬在NAFLD发病早期对细胞起保护性作用，所以SCD1表达降低，AMPK及自噬相关蛋白表达反应性增高，但长期在高脂环境中，SCD1表达会增高，AMPK及自噬相关蛋白表达则会受到抑制，脂质自噬被抑制，形成恶性循环。2）NAFLD肝细胞模型中，抑制SCD1可以减轻肝细胞脂质沉积，过表达SCD1可以加重肝细胞的脂质沉积。抑制SCD1可促进p-AMPK/AMPK表达增加，自噬增强；过表达SCD1可使p-AMPK/AMPK表达降低，自噬减弱。表明脂质可通过SCD1，调节AMPK磷酸化及脂质自噬，从而促进脂质在肝细胞的沉积及合成。3）NAFLD肝细胞模型中，抑制SCD1后，加用AMPK抑制剂可以加重肝细胞脂质沉积和合成，脂质自噬作用减弱；而过表达SCD1后，加用AMPK激动剂可以减轻肝细胞的脂质沉积和合成，脂质自噬增强。说明SCD1可通过AMPK调控脂质自噬，影响肝细胞脂质合成及沉积。 4）在NAFLD动物模型中，SCD1抑制剂能缓解高脂饮食诱导的体重增加及肝功能损害，还能减轻因高脂饮食导致的高血脂水平；同时缓解高脂饮食诱导的脂质在肝脏的沉积。通过检测肝细胞内SCD1、p-AMPK及自噬相关蛋白及自噬体分布，证实SCD1抑制剂能通过增强p-AMPK/AMPK的表达，恢复肝细胞中的脂质自噬流。 科学意义：进一步深入探讨了NAFLD 的发病机制：SCD1可以通过调控AMPK通路来调节脂质自噬，来调节脂质在肝细胞内的沉积和合成。为SCD1抑制剂应用于NAFLD的治疗提供了理论依据，使其在临床治疗中具有一定可行性。