PDIA5通过缓解内质网应激调节肝脏糖异生的机制研究

Endoplasmic reticulum stress is involved in the occurrence and development of various diseases. It is one of the common pathophysiological mechanisms of obesity and type 2 diabetes. Protein disulfide isomerase 5 (PDIA5), a disulfide isomerase located in the endoplasmic reticulum (ER), can not only catalyze protein folding (through forming the appropriate disulfide bond) and thiol-disulfide interchange reactions, but also regulate the ER stress. However, there's no related researches on the role of PDIA5 playing in the glucose metabolism hitherto. For the first time, our preliminary study found that the expression of PDIA5 was significantly decreased in non-alcoholic fatty liver disease (NAFLD). Moreover, hepatic overexpression of PDIA5 could improve glucose metabolism by inhibiting hepatic glucose output, and alleviating ER stress significantly at the same time. The current study intended to take high-fat diet (HFD)-induced obese mice and db/db diabetic mice as experimental subjects, with adeno-associated virus (AAV) injection to overexpress PDIA5, aiming at 1) detecting the pathway of PDIA5 to regulate gluconeogenesis and ER stress signaling pathway; 2) clarifying the molecular mechanism of PDIA5 on inhibiting gluconeogenesis by reducing ER stress through explore the interaction between PDIA5 and proteins that are critical in ER stress pathways; 3) finally, verifying the key role of PDIA5 in gluconeogenesis and ER stress with HFD-induced PDIA5 knockout mice. The goal of this study is to determine whether PDIA5 is a pivotal link between ER stress and gluconeogenesis, and whether PDIA5 can serve as a potential intervention target for inhibiting hepatic gluconeogenesis, therefore, clarifying the pathogenesis of hepatic insulin resistance and providing molecular biological basis for drug therapeutic targets.

内质网应激参与多种疾病的发生与发展，是肥胖和2型糖尿病的共同病理生理机制之一。PDIA5是一个定位于细胞内质网的二硫键异构酶，能使蛋白正确折叠，并可能调控内质网应激，但在糖代谢中的作用尚未见报道。我们的前期研究首次发现PDIA5在脂肪肝中低表达，而肝脏过表达PDIA5后能通过抑制肝糖输出而改善糖代谢，同时明显缓解内质网应激。本项目拟在肥胖及糖尿病小鼠中，予腺相关病毒过表达PDIA5，检测其对葡萄糖异生及内质网应激的信号通路的调控；分子水平探讨PDIA5对内质网应激关键蛋白的可能作用，明确PDIA5如何通过缓解内质网应激而抑制葡萄糖异生的分子机制；最后利用PDIA5敲除小鼠并高脂诱导，验证PDIA5在葡萄糖异生和内质网应激中的关键作用。这一研究旨在明确PDIA5是否为连接内质网应激和糖异生的关键酶，能否作为抑制肝脏糖异生的潜在干预靶点，并为阐明肝脏胰岛素抵抗的发病机制提供分子生物学依据。

内质网应激是肥胖和2型糖尿病的共同病理生理机制之一。蛋白二硫键异构酶 5（PDIA5）定位于细胞内质网，可能调控内质网应激，但在糖代谢中的作用机制尚不明确。本研究旨在通过肝脏过表达小鼠模型中研究PDIA5对糖代谢的影响，同时在肝原代细胞中过表达PDIA5对机制进行研究。我们的研究结果发现，在高脂诱导的小鼠肝脏中过表达PDIA5，能够显著改善糖耐量和胰岛素敏感性，同时影响肝糖异生关键基因的mRNA的水平，调节糖异生。在肝原代细胞中的研究结果发现，通过感染腺病毒过表达PDIA5后，肝原代细胞产糖水平下降，且糖异生关键酶mRNA表达水平下降，同时影响了激活转录因子-6（ATF6）的入核和降解，可能影响到CREB调节的转录共激活蛋白2（CRTC2）在促进糖异生方面的作用。研究结果提示PDIA5可能为连接内质网应激和糖异生的一个分子，或可作为抑制肝脏糖异生的潜在干预靶点。同时，在人群中的研究结果发现，以空腹糖异生增加为主要机制的黎明现象在糖代谢紊乱的早期就已经出现，提示糖异生在代谢性疾病尤其是糖尿病的发病过程中可能起着重要作用。