内质网应激-自噬通路在烧伤后肠黏膜紧密连接屏障损害中的作用与机制

The disruption of intestinal mucosa barrier plays a pivotal role in the pathophysiological changes after severe burn injury, however the underlying mechanisms are still unclear. Based on the published data and our previous studies demonstrating that the changes of tight junction (TJ) proteins in intestinal epithelial cells are involved in the intestinal epithelial barrier dysfunction following severe burns, we take the lead in hypothesizing that burn injury may trigger the endoplasmic reticulum stress (ERS) in intestinal epithelial cells, which in turn induces the autophagy and then leads to the disruption of intestinal mucosal TJ barrier and the increase of permeability by causing the changes of expression and/or localization of TJ proteins. Therefore, the current proposed research project is intended to study the changes of ERS, autophagy, TJ structure, TJ proteins, TJ barrier and permeability in intestinal epithelial cells after severe burns both in vivo and in vitro, and then investigate the effects of ERS-autophagy pathway on TJ structure, TJ proteins, TJ barrier and permeability by the means of inducing and inhibiting both ERS and autophagy in vivo and in vitro respectively, so as to determine the exact roles of ERS-autophagy pathway in the disruption of intestinal mucosal TJ barrier induced by severe burn injury and the underlying molecular mechanisms. It is expected that these studies will have positive effects on further understanding the molecular mechanisms of the disrupted intestinal mucosal TJ barrier following severe burn injury, and without any doubt have big referential significance to the study of other barriers.

肠黏膜屏障损害在严重烧伤病理生理变化中至关重要，但发生机制仍未完全明了。在既往基金研究中发现肠上皮细胞紧密连接(TJ)蛋白变化参与严重烧伤后肠上皮屏障损害发生的基础上，根据文献资料并结合动物预实验结果，率先提出烧伤后肠黏膜上皮细胞发生内质网应激(ERS)，并通过诱导自噬而影响TJ蛋白表达和/或分布的改变，导致肠黏膜TJ屏障损害及通透性增加的新假说。拟通过体内外实验研究严重烧伤后肠上皮细胞ERS、自噬、TJ结构、TJ蛋白、TJ屏障及通透性变化的关系，并分别采用诱导及抑制ERS与自噬等干预手段，明确严重烧伤后ERS-自噬通路对TJ 结构、TJ 蛋白、TJ屏障及通透性等的影响，从而率先阐明严重烧伤后肠上皮细胞ERS-自噬通路活化机制及其在肠黏膜TJ屏障损害中的调控作用与分子机制。对进一步阐明严重烧伤后肠黏膜TJ屏障损害及通透性增加的分子机制具有重要学术价值，对其它屏障损害的研究也有重要借鉴意义。

严重烧伤后肠黏膜屏障功能紊乱的发生机制仍不完全清楚。本项目提出严重烧伤后肠黏膜上皮细胞发生内质网应激(ERS)，并通过诱导细胞自噬而影响紧密连接（TJ）蛋白表达和/或分布的改变，导致肠黏膜TJ屏障损害及通透性增加的新假说。本项目通过体内外实验研究了严重烧伤及缺氧状态下肠上皮细胞ERS、细胞自噬、TJ蛋白、TJ屏障及通透性的变化，还分别采用诱导及抑制ERS与细胞自噬等干预手段，研究了严重烧伤及缺氧状态下ERS-自噬通路活化对肠上皮细胞TJ 蛋白、TJ屏障及通透性等的影响，以期明确严重烧伤及缺氧状态下肠上皮细胞ERS-自噬通路活化在肠上皮细胞TJ屏障损害中的调控作用与分子机制。项目的主要研究结果表明：严重烧伤早期及缺氧状态下肠上皮TJ屏障功能明显受损及通透性增加，并伴随有肠上皮细胞TJ蛋白ZO-1、Occludin与Claudin-1表达减少及分布的改变；同时，肠上皮细胞ERS明显活化，细胞自噬水平增强。结果还表明，诱导肠上皮细胞ERS活化，能加重严重烧伤或缺氧引起的肠上皮细胞自噬、屏障损害、通透性增加、TJ蛋白ZO-1、Occludin与Claudin-1表达减少及分布的改变；而抑制ERS则能减轻严重烧伤或缺氧引起的肠上皮细胞自噬、屏障损害、通透性增加、TJ蛋白ZO-1、Occludin与Claudin-1表达减少及分布的改变。此外，诱导肠上皮细胞自噬，也能加重严重烧伤或缺氧引起的肠上皮细胞自噬、屏障损害、通透性增加、TJ蛋白ZO-1、Occludin与Claudin-1表达减少及分布的改变；而抑制肠上皮细胞自噬则也能减轻严重烧伤或缺氧引起的肠上皮细胞自噬、屏障损害、通透性增加、TJ蛋白ZO-1、Occludin与Claudin-1表达减少及分布的改变。本项目的这些结果提示，严重烧伤与缺氧能诱导肠上皮细胞ERS活化，从而增强细胞自噬水平，进一步引起TJ蛋白ZO-1、Occludin与Claudin-1表达减少及分布改变，导致肠上皮细胞TJ屏障功能损害及通透性增加。这些结论初步明确了肠上皮细胞ERS-自噬通路活化在严重烧伤早期肠上皮TJ屏障功能损害中的作用与分子机制，对进一步深入阐明严重烧伤早期肠上皮屏障功能损害的分子机制具有重要意义。