芍药汤联合美沙拉嗪调控TRL4-ERK1/2-NF-κB信号通路改善溃疡性结肠炎的作用机制研究  

Study on the mechanism of Shaoyao decoction combined with mesalazine regulate TRL4-ERK1/2-NF-κB signal pathway in improving ulcerative colitis

作　　者：张薇[1] 王晓戎[2] 徐婧熙 盛红艳 杨朋 陈晓 李先伟[3] Zhang Wei;Wang Xiaorong;Xu Jingxi;ShengHongyan;Yang Peng;Chen Xiao;Li Xianwei(Department of gastroenterology,affiliated Wuhu hospital of Traditional Chinese Medicine affiliated to Anhui college of Traditional Chinese Medicine,Wuhu,Anhui,241000,China;Basic teaching and research department of traditional Chinese Medicine,Anhui College of Traditional Chinese Medicine,Wuhu,Anhui,241000,China;Department of pharmacology,Wannan Medical College,Wuhu,Anhui,241002,China)

机构地区：[1]安徽中医药高等专科学校附属芜湖市中医医院脾胃病2科,安徽芜湖241000 [2]安徽中医药高等专科学校中医教研室,安徽芜湖241000 [3]皖南医学院药理学教研室,安徽芜湖241002

出　　处：《齐齐哈尔医学院学报》2022年第8期701-707,共7页Journal of Qiqihar Medical University

基　　金：安徽省卫生健康委科研重点项目(AHWJ2021a033);2019年度安徽高校自然科学研究项目(KJ2019A1084);安徽省质量工程项目示范基层教学组织(中医教研室)(2020sjsfjxzz256)。

摘　　要：目的 观察芍药汤(SYD)联合美沙拉嗪(MES)对溃疡性结肠炎(UC)大鼠的保护作用机制是否与其抑制Toll样受体4(TRL4)介导的细胞外信号调节蛋白激酶1/2(ERK1/2)、核转录因子-κB(NF-κB)信号通路的激活有关。方法 60只Sprague Dawley大鼠随机分为正常组(NG)、模型组(MG)、SYD 20 ml·kg^(-1)组、MES 0.2 g·kg^(-1)组及SYD 20 ml·kg^(-1)+MES 0.2 g·kg^(-1)组,每组各12只。5%葡聚糖硫酸钠溶液连续喂养7天制备UC大鼠模型,造模后连续灌胃给药14天,给药后第7天和第14天进行大鼠疾病活动指数(DAI)评分。ELISA检测大鼠血浆炎症介质白介素-1β(IL-1β)、白介素-6(IL-6)和炎症小体NLR家族Pyrin域蛋白3(NLRP3)的含量。HE观察结肠病理变化,免疫组化法检测结肠黏蛋白2(MUC2)蛋白表达。qPCR检测结肠MUC2、NLRP3、IL-1β和IL-6 mRNA的表达。Western Blot检测结肠NLRP3、IL-1β、IL-6、TRL4、髓样分化因子88(MyD88)、磷酸化ERK1/2(p-ERK1/2)、磷酸化核因子κB抑制因子α(p-ⅠκBα)和磷酸化NF-κB p65(p-NF-κB p65)蛋白表达及细胞核内NF-κB p65水平。结果 与MG相比,SYD组及MES组肠黏膜病理损伤明显减轻、DAI评分显著降低、TRL4和MyD88的蛋白表达及ERK1/2、ⅠκBα和NF-κB p65磷酸化水平明显降低,同时NF-κB p65核转移显著减少、炎症因子IL-1β、IL-6和NLRP3的水平明显降低、MUC2蛋白的表达明显升高(P<0.05、P<0.01)。但与SYD组或MES组相比,SYD 20 ml·kg^(-1)+MES 0.2 g·kg^(-1)组肠黏膜损伤程度、DAI评分及NLRP3、IL-1β和IL-6的表达水平进一步降低,MUC2蛋白表达进一步升高,同时TRL4、MyD88的蛋白表达及ERK1/2、ⅠκBα和NF-κB p65磷酸化水平及NF-κB p65的核转移程度进一步降低(P<0.05)。结论 芍药汤联合美沙拉嗪对UC的疗效都优于单用美沙拉嗪或芍药汤,其机制可能与其抑制TRL4受体介导的ERK1/2、NF-κB信号通路活化,进而抑制炎症因子NLRP3、IL-1β和IL-6的表达、上调黏蛋白-2的表达有关。Objective To observe the effect of Shaoyao decoction(SYD) combined with mesalazine(MES) on ulcerative colitis(UC) and explore whether the anti-inflammatory mechanism is related to the inhibition of Toll-like receptor 4(TRL4), extracellular signal-regulated protein kinase 1/2(ERK1/2) and nuclear factor-kappa B(NF-κB) signal pathway activation.Methods 60 SD rats were randomly divided into normal group(NG), model group(MG), SYD 20 mL·kg^(-1)group, MES 0.2 g·kg^(-1)group and SYD 20 mL·kg^(-1)+MES 0.2 g·kg^(-1)group, 12 rats in each group. UC rat model was induced by feeding 5% dextran sodium sulfate solution for 7 days, the drug was administered by gavage for 2 weeks after the success of the model. Disease activity index(DAI) of rats was scored on the 7 th and 14 th day after drug administration. The levels of NLR Family Pyrin Domain Containing Protein 3(NLRP3), IL-1β and IL-6 in plasma were determined by ELISA. The pathological changes of colon were observed by HE staining and the expression of mucin 2(MUC2) was observed by immunohistochemistry. The mRNA levels of MUC2, NLRP3, IL-1β and IL-6 in colon were detected by qPCR. The protein levels of NLRP3, IL-1β, IL-6, TRL4, myeloid differentiation factor 88 (MyD88),p-ERK1/2,p-IκBα,p-NF-κB p65( cytoplasm) and NF-κB p65( nucleus) in colon were detected by Western blotting. Results Compared with MG,SYD or MES group could reduce DAI score,improvepathological injury of intestinal mucosa and up-regulate the expression of MUC2 protein. SYD or MES group also could reduce the expression of NLRP3,IL-1β and IL-6,and also could down-regulate the protein expression of TRL4 and MyD88,inhibit ERK1/2,IκBα,NF-κB p65 phosphorylation and reduce NF-κB p65nuclear transfer. Compared with SYD or MES group,the DAI score,pathological injury of intestinal mucosa and the expression of NLRP3,IL-1β and IL-6 were further decreased and the expression of MUC2 protein was further increased in SYD 20 m L· kg^(-1)+MES 0.2 g· kg^(-1)group. And the protein expression of TRL4,MyD88,and E

关 键 词：芍药汤 美沙拉嗪 溃疡性结肠炎 TRL4 ERK1/2 NF-ΚB 

分 类 号：R966[医药卫生—药理学]