青黛中的AhR配体促进IL-22表达抑制炎症性肠病的作用机理研究

Inflammatory bowel disease（IBD）has become a global disease, many studies show that IL-22 is expected to be an effective anti IBD target to repair the damaged intestinal tract and maintain the intestinal mucosal immune barrier. It was reported that the helper CD4+ T cells in the intestine, especially Th17, Th22 cells, and the ILC3 cells in intestinal epithelium were the main cells to produce IL-22 in gut, and AhR was the important transcription factor in these cells. In this study, AhR ligands indigo and indirubin from indigo Naturalis were selected as the research object which had been confirmed could therapy the IBD, the AhR pathway as a starting point, The effect of the ligands on the secretion of IL-22 in intestinal immune cells Th17, Th22 and ILC3 cells were revealed by using the mouse model of DSS induced intestinal inflammation. An in-depth analysis of the effects of indigo and indirubin on the activation of AhR pathway in ILC3 cells to secrete IL-22 by using AhR-/-&RAG1-/- double defect mice. We also analyzed the interaction between AhR and other transcription factors and synergetic factors in the process of stimulating ILC3 to secrete IL-22 by using the methods of gene defect and signal pathway blocking in order to reveal the molecular mechanism of AhR and ligands in the maintenance of intestinal immune homeostasis, and provide a theoretical basis for the rational use of indigo Naturalis to treatment of IBD.

炎症性肠病（IBD）已成为一种全球性疾病，研究表明靶向IL-22有望成为修复受损肠道，维持肠道粘膜免疫屏障的有效抗IBD靶点。肠道中的辅助性CD4+T细胞如Th17、Th22细胞，以及肠道上皮中的固有淋巴细胞（ILC3）是IL-22的主要来源，而AhR是这几类细胞的重要转录调控因子。本研究以对IBD具有治疗效果的青黛中的AhR配体靛蓝、靛玉红为研究对象，以AhR信号通路为切入点，利用DSS诱导小鼠肠道炎症模型，揭示其对肠道免疫细胞Th17、Th22和ILC3细胞分泌IL-22的影响；利用AhR-/-&RAG1-/-双缺陷小鼠深入分析靛蓝和靛玉红对ILC3细胞的AhR通路活化的影响；并综合运用基因缺陷和信号通路靶向阻断等方法，利用分子生物学手段分析AhR在刺激ILC3分泌IL-22过程中与其它转录因子和协同因子的相互作用，揭示AhR及配体在维持肠道免疫稳态中的分子作用机制。

炎症性疾病（IBD）已成为一种全球性疾病，研究表明靶向IL-22有望成为修复受损肠道，维持肠道粘膜免疫屏障的有效抗IBD靶点。肠道中的辅助性CD4+T细胞包括Th17、Th22细胞，以及肠道上皮中的固有淋巴细胞（ILC3）是IL-22的主要来源，而AhR是这几类细胞的重要转录调控因子。本研究以对IBD具有治疗效果的青黛中的AhR配体靛蓝、靛玉红为研究对象，以AhR信号通路为切入点，利用DSS诱导小鼠肠道炎症模型，揭示其对肠道免疫细胞Th17、Th22和ILC3细胞分泌IL-22的影响；本研究原本设计利用AhR-/-&RAG1-/-双缺陷小鼠深入辨析靛蓝和靛玉红的结构差别对ILC3细胞的AhR通路活化的影响。但由于基因缺陷小鼠繁殖的问题，一直没能获得双缺陷小鼠。因而转向研究了靛蓝和靛玉红的结构差别与AhR配体结合能力的关系，以及在体内的代谢情况，同时基因测序分析了靛蓝、靛玉红对小鼠粪便中肠道菌群组成的影响及与IL-22的关系。本项目表明青黛中主要有效成分靛蓝、靛玉红能显著缓解IBD的病程，且与其通过AhR通路参与调节各细胞免疫细胞分泌IL-22密切相关，并揭示AhR及配体在维持肠道免疫稳态中的作用机制。