CD73/Adora2b信号通路调节IEMT干预实验性结肠炎肠纤维化的机制研究

目的：慢性炎症诱导的肠上皮间质转化（IEMT）是肠纤维化的关键因素，CD73和腺苷代谢广泛参与肠道炎症和免疫调节。本项目探讨肠纤维化时CD73/Adora2b信号通路调节IEMT的作用机制 。方法：建立TNBS诱导实验性结肠炎肠纤维化小鼠模型，检测结肠组织胶原蛋白，纤维化相关细胞因子 TGF-β 、IL-17A等，EMT标志物，细胞间连接蛋白，CD73/Adora2b腺苷通路相关蛋白，电镜观察结肠上皮细胞间紧密连接变化；分离培养结肠上皮细胞，FACS检测CD73和Adora2b表达，转染CD73/Adora2b特异性SiRNA质粒，体外观察CD73/Adora2b通路对IEMT的调节。利用CD73-/-和（或）Adora2b-/-小鼠及细胞重建实验证实其干预作用。意义：从CD73/腺苷代谢调节IEMT的全新角度探索肠纤维化的免疫学发病机制，为寻找干预肠纤维化的新靶点提供理论和实验依据。

Aim: Intestinal epithelial-to-mesenchymal transition (IEMT) induced by chronic inflammation is one of the key factor in the pathogenesis of intestinal fibrosis. CD73 participates in the intestinal inflammation and immune regulation through adenosine receptors induced pathway. Our aim is to investigate the role of IEMT regulated by CD73/Adora2b signaling pathway in the pathogenesis of intestinal fibrosis.. Methods: Intestinal fibrosis animal model was established in mice with experimental colitis induced through clyster with TNBS+ethyl alcohol.. The expression of extracellular matrix like collegen, fibrosis-associated cytokines including CTGF、TGF-β、TNF-α、IFN-γ、IL-17A, the biomarkers of IEMT, intercellular junction protein, ATP/cAMP and CD73/Adora2b signling pathway-associated proteins in colonic tissue were detected. The changes of the intercellular junction was observed under electric microscope. Intestinal epithelial cells were enriched, cultured and transinfected with CD73 and or Adora2b-specific SiRNA. Wheather and how CD73/Adora2b regulated IEMT were observed at the cellular and molecular level in vitro. CD73-/- and/or Adora2b-/- mice and epithelial cellular restruction were used to evaluate the role of CD73/Adora2b regulated IEMT in the intestinal fibrosis in vivo. Significance: The current project will contribute to illuminate the precise immune pathogenesis of intestinal fibrosis from the perspective of CD73/Adora2b regulated IEMT, and find the potential target for the effective treatment of this disorder.