基于IL-6/JAK/STAT3/SOCS3及EGF/EGFR信号通路研究彝药美洲大蠊调控溃疡性结肠炎黏膜免疫及修复的机制

On the basis of our continuous series researches for decades on the medicinal insect Periplaneta americana, anew bioactive extract Ento-A with notable therapeutic effects towards ulcerative colitis (UC) was obtained using the new purification process andthe guidance of in vitro and in vivo bioactivity assays.. Previous studies explored that Ento-A possessed significant therapeutic effects on acute, chronic and immunological UC in rats, and the bioactive extract is mainly composed of polypeptides and benefits to the expression of epidermal growth factor. The proposed study aims to further analyze the influences of Ento-A on the expressions of intestinal mucosal immune, repair, and inflammatory related cytokines of 2,4,6-trinitrobenzene sulfonic acid-induced UC in ratsthrough its effects on IL-6/JAK/STAT3/SOCS3 and EGF/EGFR signaling pathwaysin the mucosal injury repairing process; and to probe the mechanism of action of Ento-A on intestinal mucosal immunomodulatory and repair in UC. The results of this study are expected to provide a theoretical basis for the applications of more efficient pharmaceutical technology of colonic localization and targeting preparations towards the bioactive extract and improving the clinical efficacy of Ento-A in the treatment of UC, as well as to provide scientific basis for better development of Yunnan's Yi nationality characteristic insect medicine.

本项目在课题组数十年对药用昆虫美洲大蠊系列研究的基础上，根据体内外药效活性指导，使用新工艺从美洲大蠊中精制了对溃疡性结肠炎有显著治疗作用的活性部位Ento-A。前期研究显示：该活性部位对大鼠急、慢性及免疫性溃疡性结肠炎有显著疗效，Ento-A以多肽类物质为主要成分且有助于表皮生长因子的表达。现拟进一步通过对2,4,6-三硝基苯磺酸诱导的UC大鼠肠黏膜损伤修复过程中IL-6/JAK/STAT3/SOCS3及EGF/EGFR信号通路的作用，分析其对肠黏膜免疫、损伤修复及相关的炎性细胞因子表达的影响；探讨Ento-A对溃疡性结肠炎黏膜免疫及修复的可能机制。本研究结果预期可为该活性部位应用更高效的结肠定位靶向制剂技术、提高临床的UC治疗效果提供理论基础，为更好的应用开发云南彝族药物提供科学依据。

溃疡性结肠炎（UC）是以黏膜上皮细胞凋亡和炎症浸润为病理基础的免疫功能紊乱性疾病。目前临床以抗炎为主的治疗方案并不能得到满意的结果，究其原因是损伤黏膜的愈合质量不高，导致了UC的反复发作。大量的研究报道和临床实践表明，美洲大蠊对消化系炎性疾病的炎症和黏膜溃疡有显著的疗效。然而，美洲大蠊对UC的作用机制鲜有报道。因此，项目研究旨在探讨美洲大蠊活性部位对溃疡性结肠炎大鼠IL-6/JAK2/STAT3炎症通路和EGF/EGFR修复通路的作用机制，解析美洲大蠊活性部位的物质基础，通过建立体外细胞模型筛选具有抗炎、抗氧化应激和促进结肠上皮细胞增殖、迁移的功效物质，并深入阐述相关功效物质抗UC的作用机制。本次研究内容及结果如下：（1）美洲大蠊提取物PAWE能够通过调控CD4+ T细胞亚群的分化平衡，影响Th1/Th2和Th17/Treg细胞分泌细胞因子，从而减轻UC大鼠的炎症状态，同时改善VEGF、bFGF和PDGF的表达水平，促进大鼠结肠受损黏膜愈合；（2）PAWE通过抑制IL-6/JAK2/STAT3的信号传导，调节EGF与EGFR间的表达平衡，抑制结肠黏膜的过度增生，提高肠黏膜的愈合质量；（3）采用LC-MS/MS和nano LC-ESI-MS/MS等现代质谱技术和相关数据库检索比对，最终检测到PAWE中小分子化合物1355个，多肽类物质913条；（4）PAWE抗UC的作用机制可能与其富含的活性氨基酸促进NRF2核转位，调控下游抗氧化应激蛋白GCLM、NQO1、HO-1和GPX2的表达有关；（5）PAWE中的多肽可以通过调控EGFR及下游的PI3K/AKT/mTOR通路有效促进结肠上皮细胞增殖、迁移。