基于TGF-β/Smad/MAPK诱导的上皮-间质转化研究黄芪甲苷促进UC黏膜屏障修复机制

Reduced intestinal mucosal barrier function plays an important role in the pathogenesis of ulcerative colitis (UC), therapies that focused on intestinal mucosal barrier repair have been considered as a vital strategy for UC treatment. Epithelinal-to-mesenchymal transition (EMT) is the necessary step in mucosal barrier repair, while the over-activity in EMT can induce the decrease of mucosal barrier function. Transforming growth factor β (TGF-β) is the key important inducer of EMT, it regulates EMT via Smad classical pathway and non-Smad pathways. It is reported that there was an over-activation of EMT and up-regulation of TGF-β/Smad/MAPK signal in UC patients. Our previous study showed that astragaloside had therapeutic effect on UC rats, and the underlying mechanism has close correlation with promoting mucosal barrier repair and inhibiting EMT over-activation. Based on above evidences, we hypothesize that astragaloside may enhance mucosal barrier repair in UC rats by inhibiting EMT over-activation via TGF-β/Smad/MAPK signaling pathways. To test this hypothesis, we will firstly investigate the effect of astragaloside on mucosal barrier repair, EMT, and TGF-β/Smad/MAPK signal protein expression in UC rats. Meanwhile, the effect and the underlying mechanism of astragaloside on EMT and TGF-β/Smad/MAPK signal pathways will also be investigated using murine intestinal epithelial cell line. Clarify the effect and mechanism of EMT on intestinal mucosal barrier function of UC may provide a novel target for UC therapy, and identify the effect and molecular mechanism of astragaloside on intestinal mucosal barrier repair in UC may provide useful information for the clinical usage of astragaloside in UC treatment.

黏膜屏障功能降低是溃疡性结肠炎（UC）发病的关键环节，促进黏膜屏障修复对UC治疗至关重要。上皮-间质转化（EMT）是黏膜修复必需环节，其过度活化可致黏膜屏障功能下降。转化生长因子β（TGF-β）是EMT必需的诱导因子，经由Smad与非Smad通路调控EMT。文献报道，UC存在EMT过度活化及TGF-β/Smad/MAPK信号上调；课题组发现黄芪甲苷可促进黏膜修复和抑制EMT过度活化。基于此，我们提出“黄芪甲苷通过TGF-β/Smad/MAPK抑制EMT过度活化并促进UC黏膜屏障修复”的设想。研究拟用UC模型研究黄芪甲苷对黏膜屏障修复、EMT及TGF-β/Smad/MAPK信号影响；并用肠上皮细胞研究黄芪甲苷对EMT及TGF-β/Smad/MAPK信号蛋白的影响及机制。阐明EMT对UC黏膜屏障的影响机制，有望为UC治疗提供新靶点；明确黄芪甲苷促进肠黏膜修复的分子机制可为其临床应用提供依据。

黏膜屏障功能降低是导致溃疡性结肠炎（UC）发病的关键环节之一，促进黏膜屏障修复对于UC治疗至关重要。上皮细胞-间质转化（EMT）是黏膜屏障修复的必需环节，TGF-β/Smad/MAPK信号通路是诱导EMT的主要信号通路，EMT的过度活化与肠道黏膜屏障功能降低有关。前期研究表明黄芪甲苷可改善溃疡性结肠炎模型大鼠肠道黏膜屏障的功能，但其作用机制尚不清楚。本项目通过反复灌肠50%TNBS乙醇溶液，建立溃疡性结肠炎肠道纤维化模型，探究黄芪甲苷改善溃疡性结肠炎肠道黏膜屏障的作用机制。研究发现反复炎症刺激可引起肠道EMT的改变，肠道TGF-β/Smad/MAPK信号通路激活，肠道ZO-1和Occludin蛋白表达降低；黄芪甲苷可抑制肠道TGF-β/Smad/MAPK信号通路，增加肠道E-cadherin、β-catenin、ZO-1和Occludin蛋白表达，降低肠道N-cadherin、Vimentin及Snail蛋白表达。上述结果表明黄芪甲苷通过抑制肠道EMT过程改善肠道黏膜屏障功能。细胞实验发现， TGF-β可诱导IEC-6细胞发生上皮细胞-间质转化，降低肠道黏膜屏障功能，黄芪甲苷可抑制TGF-β诱导的上皮细胞-间质转化过程，抑制TGF-β/Smad/MAPK信号通路，改善黏膜屏障。通过项目的实施，初步明确了黄芪甲苷抑制EMT过程改善黏膜屏障的作用机制，并为后续深入研究改善肠道黏膜屏障的机制奠定了基础。