胰岛素抵抗（IR）在轻微型肝性脑病（MHE）发病中的作用及分子机制研究

Higher prevalence of minimal hepatic encephalopathy (MHE) in cirrhosis patients has been taken more and more seriously. The pathogenesis and the therapy method of MHE have to be resolved. MHE is characterized by cognitive disorder. Some studies have reported that insulin resistance (IR) was closely related with the impairment of cognitive function. In our previous study, we found IR in both of MHE patients and MHE rats, and the intimate relationship of cognitive decline with IR in MHE rats. Therefore, the study aims to confirm that IR is one of the essential factors for pathogenesis of MHE, to determine that insulin–mediated classical signaling pathway ―InsR (insulin receptor)/IRS (insulin receptor substrate)/PI3K (phosphatidylinositol 3- kinase)/AKT (protein kinase B) has the effect on downstream cognitive signaling pathway ―NMDAR (N-methyl-D-aspartic acid receptor)/CaMKⅣ(calmodulin kinase Ⅳ)/CREB (cAMP-response element binding protein), and to clarify that the inactivation of insulin -mediated cognitive signaling pathway (InsR/IRS/PI3K/AKT/NMDAR/CaMKⅣ/CREB) is involved in the pathogenesis of MHE, by using MHE rat/mice models and primary cultured neurons models which were intervened by insulin, insulin sensitizer, antibody, antagonist, agonist, short hairpin RNA (shRNA) or gene overexpression plasmid etc., and using techniques such as Morris water maze, Y maze, electrophysiology test, Q/RT-PCR, immunoblotting, or coimmunoprecipitation etc.. Furthermore, our study may provide new theory basis of pathogenesis and therapy of MHE.

轻微型肝性脑病（MHE）在肝硬化患者中的高患病率日益受到重视，其复杂的发病机制及有效的治疗措施亟待解决。MHE主要表现为认知障碍。有研究报道胰岛素抵抗（IR）与认知功能的受损密切相关。课题组前期研究发现MHE患者或大鼠中均存在IR，以及MHE大鼠中的IR与其认知障碍的发生具有显著相关性。为了进一步明确IR是MHE发病的主要因素之一，探索胰岛素介导的经典信号通路（InsR/IRS/PI3K/AKT）对下游认知信号通路（NMDAR/CaMKⅣ/CREB）的影响，阐明IR参与MHE发病的分子机制与脑内胰岛素介导的认知信号通路失活相关；本课题拟采用胰岛素、胰岛素增敏剂、抗体、激动剂、抑制剂、shRNA、过表达质粒等干预MHE大鼠/小鼠模型或原代神经元模型，运用Morris水迷宫、Y迷宫、电生理测试、Q/RT-PCR、免疫印迹、免疫共沉淀等技术进行研究；为MHE发病机理及其治疗提供新的理论依据。

本项目旨在探讨轻型肝性脑病（MHE）发病的具体分子机制。我们采用抗体、激动剂、抑制剂、shRNA、过表达质粒等干预MHE大鼠/小鼠模型或原代神经元模型，运用Morris水迷宫、Y迷宫、电生理测试、Q/RT-PCR、免疫印迹、免疫共沉淀等技术进行研究；发现1）IR是MHE发病的主要因素之一；2）IR参与MHE发病的分子机制与脑内胰岛素信号通路介导的下游认知信号通路（InsR/IRS/PI3K/Akt/NMDAR/CaMK Ⅳ/CREB）相关的研究；3）冬凌草甲素改善胰岛素抵抗（IR）介导的MHE认知障碍；4）硫氢化钠改善多巴胺介导的MHE认知障碍；5）多巴胺通过介导胆固醇超负荷诱导MHE认知障碍；6）黄芩苷（baicalin，BAI）改善MHE大鼠认知障碍；7）DA通过引发谷氨酸（Glu）蓄积诱导MHE发病；8）DA通过抑制Shh（Sonic Hedgehog）信号通路活性参与MHE发病；9）MHE发病与神经元来源的Wnt5a表达受抑有关；本项目所取得的成果为MHE的发病机理提供更新的理论解释，并为MHE的治疗提供更全面的理论依据及相关靶标。