内质网应激IRE1－XBP1S通路在高糖引起肾脏及系膜细胞发生氧化应激及损伤中的机制研究

Oxidative stress plays key role in diabetes-induced renal injury. The dowstream mechanism of oxidative stress in renal injury has been extansively investigated: overactivation of intrarenal RAS, renal glomerular endothelial cell damage, mesangial cell dysfunction and proinflammaroty factor releases are all related to the high glucose-induced oxidative stress. However, the upstream mechanism of oxidative stress is unclear. In our primary study, the data showed that: 1) supression in IRE1-XBP1S pathway was observed in both kidney of diabetic rats and high glucose-treated mesangial cells; 2) thanscriptional factor CHOP was directly regulated by XBP1S; 3) there is bingding site on the promoter regieon of p47phox and gp91phox; 4) overexpression of XBP1S reversed the high glucose-induced ROS generation and mesangial cell proliferation. Based on the preliminary data, we hypothesized that: hyperglycemia induced supression in IRE1-XBP1S pathway, then dereased the expression of CHOP, which inturn increase the expressions of p47phox and gp91phox, rsult in increase in NADPH oxidase activity and oxidative stress. The proposal plans to test the hypothesis on diabetic rats and isolated renal mesangial cell.

氧化应激（oxidative stress）发生在糖尿病引起肾损害中起关键作用。其下游机制已被充分证实：氧化应激与肾局部RAS激活、系膜细胞功能异常、小管上皮细胞表型转变、炎症因子释放等均密切相关，然而，糖尿病导致氧化应激的上游机制却不清楚。我们在预实验中发现：1) 高糖导致肾脏及系膜细胞出现内质网应激（ERS），ERS中IRE1－XBP1S通路受到抑制；2) XBP1S能直接调控核转录因子CHOP(C/EBP homologous protein, CHOP)；3) NADPH氧化酶p47和gp91亚基Promoter区域有CHOP结合位点；4) 过表达XBP1S能抑制高糖引起的氧化应激。基于上述结果提出假说：高糖导致ERS，其中IRE1－XBP1S通路受到抑制，通过核转录因子CHOP调控NADPH亚单位表达升高，氧化酶活性增强，氧化应激发生和肾损伤。本课题拟在整体和离体上验证这一假说。

随着我国糖尿病发病率的快速升高，糖尿病导致的肾脏损伤已经成为我国慢性肾损伤发生的重要原因。肾小球硬化和肾间质脏纤维化是糖尿病肾病的重要特征。本课题研究结果显示，糖尿病时，持续的高糖环境导致细胞代谢发生改变，细胞代谢改变促发的大量蛋白合成增加导致细胞内质网应激的发生，内质网应激中IRE1/XPB1通路被激活，剪切型XBP1（XBP1S）减少，作为核转录因子，受其直接调控的下游靶基因CHOP表达水平下降，导致肾小球系膜抗氧化能力下降，细胞发生氧化应激，进而坏死性凋亡增加，加剧糖尿病导致的肾损伤。肾素-血管紧张素系统（RAS）的过度激活在糖尿病肾病的发生发展中起到至关重要的作用，ROS水平的升高与RAS过度激活有关，转激活因子SDNp102水平的升高在肾脏组织RAS过度激活中起作用，SDNp102直接参与了AT1等RAS系统多个成员的表达上调，用活性氧清除剂NAC可以缓解纤维化进展。RAS过度激活通过TGF-b加剧纤维化和肾损伤，本课题研究还显示，重编程因子c-Myc通过调控intergrin av的表达参与TGF-b激活的调控过程和纤维化的进展，炎症反应导致的肾脏组织过度修复是纤维化发生的关键，TGF-b可通过促进肾脏组织炎性反应的发生，导致肾组织过度修复，形成纤维疤痕。本课题研究成果还显示Caspase-11参与了肾脏固有细胞产生的多个炎症因子，包括IL-b，IL-18的激活过程，并通过直接切割Caspase-11促进肾脏小管上皮细胞的凋亡。本课题研究成果为认识糖尿病肾病的发展机制和纤维化发生机制提供了新的实验依据。