乳酸菌调控内质网应激在肠粘膜屏障损伤修复中的机制研究

The intestinal mucosal barrier injury is closely related to the occurrence and development of many diseases, and the probiotic repair of intestinal epithelial cells (IEC) damage plays a crucial role in the maintenance of intestinal barrier function. However, the probiotic repair mechanism of IEC damage is unclear. Our previous experimental study found that the probiotic repair mechanism of IEC damage was coupled with the alteration of Grp78 and IRE1 expression and NF-κB-mediated proinflammatory cytokines expression, suggesting that endoplasmic reticulum stress (ERS)-mediated intracellular signaling pathway involved in the probiotic repair mechanism of IEC damage. To this end, from the molecular, cellular, organizations, and animals levels, the regulation of relations between the ERS and NF-κB signaling pathway were evaluated In the subject of human intestinal epithelial cell line Caco-2, IL-10-/ - IEC and IL-10-/ - mouse model and clinical IBD intestinal specimens,through the intervention of many intermediate factors as TLR ,p38 MAPK, IRE1,Grp78, NF-kappa B and so on, eventually to clarify the role of TLR-p38 MAPK-ERS (IRE1/XBP1)-NF-κB regulatory network in the specific probiotic repair mechanism of IEC damage.In this respect, it has important theoretical and practical value for promoting the understanding of the pathogenesis and prevention of diseases associated with intestinal barrier injury.

肠黏膜屏障损伤与诸多疾病发生发展密切相关，而肠上皮细胞（IEC）损伤的益生菌修复在维护肠屏障功能方面发挥着至关重要作用，但有关IEC损伤益生菌修复机制尚不清楚。我们前期实验发现IEC损伤益生菌修复伴有Grp78和IRE1表达活性降低以及NF-κB介导的促炎因子表达下调，提示内质网应激（ERS）介导的胞内信号通路参与了IEC损伤益生菌修复机制。为此，本课题通过人肠上皮细胞系Caco-2、IL-10-/- IEC及IL-10-/-小鼠模型和临床IBD肠标本，从分子、细胞、组织以及动物整体水平分别评价ERS和NF-κB信号通路间的调控关系，通过干预TLR、p38 MAPK、IRE1、Grp78、NF-κB等多个中间环节，阐明TLR- p38 MAPK- ERS（IRE1/XBP1）-NF-κB调控网络在IEC损伤益生菌修复中分子机制，对推进肠粘膜屏障损伤相关疾病的防治具有重要理论和实际参考价值。

肠黏膜屏障功能损伤参与了多种疾病的发生发展，其中包括炎症性肠病（IBD），且至今尚缺乏有效治疗手段，目前研究证实益生菌对IBD肠上皮细胞（IEC）的损伤有保护作用，但其具体作用机制尚未阐明，结合我们前期实验发现IEC损伤益生菌修复伴有Grp78和IRE1表达活性降低以及NF-κB介导的促炎因子表达下调，提示内质网应激（ERS）介导的胞内信号通路参与了IEC损伤益生菌修复机制。为此，本课题通过人肠上皮细胞系Caco-2、IL-10-/- IEC及IL-10-/-小鼠模型和临床IBD肠标本，从分子、细胞、组织以及动物整体水平分别评价ERS和NF-κB信号通路间的调控关系，试图阐明ERS相关调控网络在IEC损伤益生菌修复中分子机制。研究结果表明，植物乳杆菌能显著减轻IEC损伤，增强肠黏膜屏障功能，缓解肠道炎症表现，这与内质网应激反应的调节密切相关，其具体作用机制主要包括：一是通过IRE1-NFκB信号通路调控内质网应激抑制促炎细胞因子的表达，二是通过PERK-elF2α-Chop信号通路调控内质网应激抑制细胞的凋亡。另对肠道菌群的宏基因组检测及功能分析发现，植物乳杆菌干预后能显著改变炎症状态下肠道菌群构成，显著降低Firmicutes/Bacteroidetes比率，提高与“细胞周期调控、细胞分裂、氨基酸的转运和代谢、碳水化合物的运输和代谢、转录、复制、重组和修复”等有关的基因含量，且首次发现B.salanitronis与炎症性肠病密切相关，以上研究揭示植物乳杆菌具有调节肠道菌群及影响机体代谢的作用。本研究结果更具用临床实用参考价值，本研究对IEC损伤益生菌修复的新的作用靶点阐明，将有利于从分子水平寻求新的治疗思路，对推进肠上皮屏障功能障碍发病机制的认识和相关疾病的防治具有重要的意义。