IRE1α/XBP1s-GRP78-Beclin1诱导小管上皮细胞自噬在肾脏缺血再灌注损伤中的作用及机制

Endoplasmic Reticulum stress is one of the mechanisms of acute renal injury induced by ischemia reperfusion(I/R). Excessive endoplasmic Reticulum stress caused by renal injury can make XBP1 cleave into active XBP1s. We found that XBP1s can promote autophagy to play a protective role in renal I/R injury, but the mechanism is still unknown. In this paper, GFP-LC3 fusion protein tracer and transmission electron microscope were used to evaluate autophagy level of renal tubular epithelial cells in I/R model in vivo and vitro. The interaction between XBP1s and GRP78 protein was determined by immunoco-precipitation technique. By administering an endoplasmic reticulum stress inhibitor, local injection of IRE1α siRNA, XBP1s siRNA, and GRP78 siRNA to kidney, we verified that IRE1α/XBP1s-GRP78-Beclin1 reduces renal I/R injury by inducing autophagy in tubular epithelial cells.The findings of this study will provide a novel insight into the mechanism of renal ischemia-reperfusion injury and lay a foundation for the future development of new treatment modalities.

内质网应激是缺血再灌注（I/R）诱导急性肾损伤的机制之一。肾损伤引起的过度内质网应激能使XBP1剪切成活性XBP1s。我们发现XBP1s可以促进自噬在肾脏I/R损伤发挥保护作用，但机制仍未明。本课题拟在体内外I/R模型，应用GFP-LC3融合蛋白示踪、透射电镜等方法，评价小管上皮细胞自噬水平；利用免疫共沉淀技术，明确XBP1s与GRP78的蛋白蛋白相互作用；通过给予小鼠腹腔注射内质网应激抑制剂、肾脏局部点注射IRE1α-siRNA、XBP1s-siRNA、GRP78-siRNA明确IRE1α/XBP1s-GRP78-Beclin1通过诱导小管上皮细胞自噬减轻肾脏I/R损伤，为阐明肾脏I/R损伤的机制提供全新的视角，为探讨新的治疗手段提供重要的线索。

缺血再灌注（I/R）损伤是大脑，心脏和肾脏等靶器官损伤的常见原因，由I/R引起的肾损伤是已知的急性肾损伤的重要原因。内质网应激（ERS）是I/R诱导的急性肾损伤的机制之一。X-盒结合蛋白1s（XBP1s）是ERS的关键介质，在大多数情况下，XBP1s在调节多细胞生物功能和参与信号转导基因的表达、氧化还原稳态、细胞生长和分化、碳水化合物代谢方面充当内质网的保护因子。肾损伤时，过度内质网应激可使XBP1 mRNA的切割和随后剪接翻译成剪切的XBP1s，XBP1s通过诱导自噬囊泡形成和Beclin1、LC3-βII的表达在内皮细胞中触发自噬反应。本研究发现，肾脏缺血再灌注损伤后，过度的ERS激活IRE1α，使活性的XBP1s表达增加，转染XBP1s过表达质粒使XBP1过表达后，促进GRP78 的表达，增强自噬，减轻肾损伤；NaHS预处理可通过抑制自噬减轻肾脏缺血再灌注损伤，并且IRE1抑制剂STF-083010干预后，小鼠肾脏缺血再灌注损伤减轻，从而为阐明肾脏I/R损伤的机制提供全新的视角，为降低肾脏I/R损伤急性死亡率和预防急性肾损伤进展为慢性肾衰竭寻找新的治疗策略。