HSPA12A基因沉默对非酒精性脂肪肝病的保护作用及机制研究：对“miR-29/IFN-r/STAT1/巨噬细胞M1极化”信号的负调节

The Morbidity of Non-alcoholic fatty liver disease (NAFLD) is higher than 14% in China. Evidence demonstrates that excessive inflammation mediated by M1 macrophage plays critical roles in the development and progression of NAFLD, while “miR-29/IFN-r/STAT1” signaling axis is important for M1 polarization of macrophage..HSPA12A is a novel heat shock protein which biological role is barely known. During the last NSFC (81571290) studies, we observed that HSPA12A was upregulated in the hepatocytes of NAFLD patients. Interestingly, knockout of HSPA12A expression in hepatocytes attenuates the high fat diet-induced liver fat accumulation and dysfunction (characters of NAFLD) in mice. The pilot data suggest that this protective action of HSPA12A silence could be mediated by the increase of hepatocyte-derived exosomes containing miR-29, which in turn suppresses the macrophage “miR-29/IFN-r/STAT1/M1 polarization” signaling-mediated inflammation..To test this possibility, we will comprohensively evaluate the protection of HSPA12A silence from NAFLD using hepatocyte HSPA12A transgenic and knockout mice. We will also investigate whether the protection of hepatocyte HSPA12A silence against NAFLD is achieved by the suppression of “miR-29/IFN-r/STAT1/macrophage M1 polarization” signaling using both mice and cell culture models. The results will provide novel clues for identifying potential targets of NAFLD based on the inter-cellular interactions of “hepatocyte - macrophage”.

M1型巨噬细胞介导的炎症反应是非酒精性脂肪肝病(NAFLD)的关键病理机制，而“miR-29/IFN-r/STAT1”信号对巨噬细胞M1极化至关重要。.HSPA12A是功能尚知之甚少的热休克蛋白。在前1项国家自然基金研究中，我们发现：HSPA12A在NAFLD病人肝细胞表达增加，而HSPA12A基因沉默则减轻高脂诱导的NAFLD，且该作用可能是通过增加肝细胞exosome负载miR-29、抑制巨噬细胞“IFN-r/STAT1/M1极化”信号介导的炎症反应而实现的。.本项目将在前期基础上，采用肝细胞HSPA12A转基因鼠、基因敲除鼠，综合评价HSPA12A基因沉默对NAFLD的保护作用，并结合细胞模型，阐明 “miR-29/IFN-r/STAT1/巨噬细胞M1极化”信号系统在其中的作用，以明确其保护机制。研究结果将从“肝细胞-巨噬细胞”相互影响的角度，为防治NAFLD提供新的思路。

非酒精性脂肪肝炎(Nonalcoholic steatohepatitis, NASH)是世界性发病率最高的慢性肝脏疾病。巨噬细胞介导的炎症反应在NASH病理进程中具有重要作用。然而，目前尚缺乏理想控制巨噬细胞炎性活化、从而治疗NASH的相关药物。热休克蛋白A12A (Heat shock protein 12A,HSPA12A)是新发现的HSP70家族新成员。本研究中，我们发现NASH患者的肝脏中HSPA12A表达增加、血清中HSPA12A水平升高。值得关注的是，HSPA12A基因敲除(Hspa12a-/-)减轻高脂食物(high-fat diet, HFD)诱导的肝脏脂肪堆积和肝功能损害。HFD所致的肝脏巨噬细胞M1激化、炎症反应，也被在HSPA12A基因敲除所缓解。通过功能获得、功能缺失实验，我们发现肝脏细胞新合成脂肪受巨噬细胞HSPA12A的旁分泌调控。进一步实验表明，HSPA12A与2型丙酮酸激酶(M2 isoform of pyruvate kinase,PKM2)直接相互作用并促进其向细胞核移位，进而促进巨噬细胞M1激化、分泌炎症因子，最终通过旁分泌形式刺激肝细胞从头合成脂肪。综上所述，我们的结果表明，HSPA12A是一个新的巨噬细胞M1激化、NASH发生发展的调节因子，该作用是通过与PKM2直接作用、并促进其向细胞核转位而实现的。因此，抑制巨噬细胞HSPA12A可能对NASH具有潜在治疗价值。