从"ERK/MAPK"通路探讨益气健脾药膳保护化疗大鼠胃肠黏膜屏障的机制

The high incidence can be found in the chemotherapy-induced gastrointestinal mucositis (GIM)in clinic.The targeted therapy of GIM has currently become the research hotspot worldwide. Previous researches have shown that the MAPK signal pathway, ERK and its receptor signaling pathways are closely related to the pathogenesis of gastrointestinal damage and the repair process afterwards. Based on the previous clinical and laboratory studies, the popular traditional Chinese health-protection food,known as " Qi Yao Ji Jin Gruel", has the a significant effect on replenishing Qi to invigorate the spleen , regulating stomach and promoting digestion.Meanwhile, this particular diet can also relieve chemotherapy-induced gastrointestinal symptoms, regulate the immunologic function of intestinal tract and the levels of gastrointestinal hormones, and maintain the expression of tight junction-related proteins in rats with chemotherapy-induced GIM. On the basis of previous researches,our team used the rats models of chemotherapy-induced GIM and colon cancer cells of Caco-2 as the research platform,based on "ERK/MAPK" signaling pathways,techniques such as tumor cells culture in vitro,Western Blot will be performed to investigate the changes of the tight junction associated protein and immune factors in the intestinal epithelium, ERK1/2, P-ERK1/2, GRB2, Sos, Ras, Raf, MEK expression,which is used to elucidate the molecular mechanism and action target in preventing and treating GIM,so as to provide a scientific basis for the application of health-protection food in replenishing Qi to invigorate the spleen after chemotherapy.

化疗所导致的胃肠黏膜屏障损伤发病率高，其靶向治疗成为当今的研究热点。研究发现MAPK信号通路、ERK及其受体信号传递途径与胃肠黏膜损伤病理机制及修复过程密切相关。本课题组前期研究证实，药膳"芪药鸡金粥"具有益气健脾，和胃消食之功效，能有效减轻化疗后胃肠道反应，调节化疗后胃肠黏膜损伤大鼠肠道免疫及胃肠激素水平,维持肠道机械屏障紧密连接相关蛋白的表达。本项目在前期研究基础上以化疗后GIM大鼠模型及结肠癌细胞Caco-2为研究平台，基于"ERK/MAPK"通路，运用细胞培养、Western Blot 等技术，通过观察肠上皮紧密连接相关蛋白、肠上皮相关免疫因子以及细胞外信号调节蛋白激酶（ERK1/2、P-ERK1/2），GRB2、Sos、Ras、Raf、MEK等蛋白表达的变化，阐明"芪药鸡金粥"修复GIM的分子机制与作用靶点，为益气健脾药膳在化疗患者中的应用提供科学依据。

化疗所导致的胃肠黏膜屏障损伤发病率高，其靶向治疗成为当今的研究热点。研究发现MAPK信号通路、ERK及其受体信号传递途径与胃肠黏膜损伤病理机制及修复过程密切相关。有关“芪药鸡金粥”在缓解化疗后胃肠道反应的临床疗效已获得证实。本研究将在前期研究的基础上从ERK/MAPK通路角度，观察芪药鸡金粥对化疗所致胃肠黏膜损伤大鼠屏障功能及通路蛋白与肠道紧密连接蛋白等的影响，进而探讨芪药鸡金粥的作用机制。实验分为体内和体外实验，体内实验通过建立GIM大鼠模型，检测 “芪药鸡金粥”对化疗后大鼠肠道免疫因子、紧密连接相关蛋白（Claudin-1、Occludin、JAM-1和Z0-1）及通路相关蛋白（ERK1/2、P-ERK1/2、MBP、MAP-2）表达水平；体外实验通过“芪药鸡金粥”药物血清干预体外培养人结肠癌Caco-2 细胞，检测紧密连接相关蛋白（ claudin-1 、Occludin 、JAM-1 和Z0-1 ）和通路相关蛋白（ERK1/2、P-ERK1/2、MBP、MAP-2）的表达与活化；阐明“芪药鸡金粥”对肠上皮细胞“ERK／MAPK”信号转导通路的作用靶点。研究结果表明，芪药鸡金粥可上调肠道机械屏障紧密连接蛋白尤其是Occludin蛋白的表达，有利于肠黏膜CD3+、CD4+T淋巴细胞增生，改善肠黏膜T淋巴细胞亚群的比例，进而维持肠道机械屏障和生物屏障的功能，降低化疗后肠道内毒素和D-乳酸含量，减少肠道细菌移位，并在一定程度上预防及减轻化疗所致的胃肠道黏膜损伤反应。本研究从分子蛋白角度验证了芪药鸡金粥在化疗后胃肠黏膜损伤中的治疗作用，为中医药膳资源开发的及推广应用提供了证据支撑，为中医护理研究提供指导和新的方向。