基于p38 MARK/MMP-9信号通路探讨自拟复方愈溃汤对UC大鼠肠道炎症、氧化应激及屏障功能的影响

Exploration of Effects of Compound Yukui Decoction on Intestinal Inflammation,Oxidative Stress and Barrier Function in UC Rats Based on p38 MARK/MMP-9 Signaling Pathway

目的探讨自拟复方愈溃汤通过p38促分裂原活化的蛋白激酶(MAPK)/基质金属蛋白酶9(MMP-9)通路对溃疡性结肠炎(UC)大鼠肠道炎症、氧化应激及屏障功能的影响。方法选取雄性Wistar大鼠75只,采用随机数字表法分为对照组、模型组、低、中、高剂量汤药组,每组15只。除对照组外,其余各组均以2,4,6-三硝基苯磺酸/乙醇致炎法建立UC大鼠模型。其中,对照组和模型组以0.9%氯化钠溶液灌胃,低、中、高剂量汤药组分别以自拟复方愈溃汤4.2 g/(kg·d)、8.4 g/(kg·d)、16.8 g/(kg·d)灌胃。比较各组大鼠的一般情况,结肠组织疾病活动指数(DAI)评分和组织病理评分、炎症因子[肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)和γ干扰素(INF-γ)]、氧化应激指标[超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱甘肽(GSH)]、肠黏膜屏障功能蛋白[闭锁蛋白(Occludin)、闭锁小带蛋白-1(ZO-1)]及p-p38 MAPK/p38 MAPK、MMP-9蛋白表达的变化。结果低、中、高剂量汤药组DAI评分和组织病理评分均低于模型组[(2.55±0.50)分、(1.50±0.32)分、(0.82±0.25)分比(3.25±0.65)分,(5.15±1.14)分、(3.00±0.78)分、(2.00±0.65)分比(6.33±1.30)分],且低、中、高剂量汤药组DAI评分和组织病理评分呈剂量依赖性降低趋势(P<0.05)。低、中、高剂量汤药组TNF-α、IL-6、IFN-γ水平均低于模型组,且低、中、高剂量汤药组TNF-α、IL-6、IFN-γ水平呈剂量依赖性降低趋势(P<0.05)。低、中、高剂量汤药组SOD、GSH水平均高于模型组,MDA水平均低于模型组;且低、中、高剂量汤药组SOD、GSH水平呈剂量依赖性升高趋势,MDA水平呈剂量依赖性降低趋势(P<0.05)。低、中、高剂量汤药组Occludin、ZO-1表达均高于模型组,p-p38 MAPK/p38 MAPK与MMP-9表达均低于模型组;且低、中、高剂量汤药组Occludin、ZO-1表达呈剂量依赖性升高趋势,p-p38 MAPK/p38 MAPK与MMP-9表达呈剂量依赖性降低趋势(P<0.05)。结论自拟复方愈溃汤能通过抑制p38 MARK信号通路的活化和MMP-9蛋白的表达,降低UC大鼠结肠组织炎症因子水平,减轻氧化应激反应,改善肠黏膜上皮屏障功能。

Objective To investigate the effect of compound Yukui decoction on intestinal inflammation,oxidative stress and barrier function in ulcerative colitis(UC)rats via the p38 mitogen-activated protein kinase(MAPK)/matrix metalloproteinase-9(MMP-9)pathway.Methods A total of 75 male Wistar rats were selected and divided into a control group,a model group,a low,a medium and a high dose decoction group by random number table method,with 15 rats in each group.Except for the control group,all the other groups established UC rat models using 2,4,6-trinitro-benzene-sulfonic acid/ethanol induced inflammation method.The control group and the model group were given intragastric administration of 0.9%sodium chloride solution.The low,medium and high dosedecoction groups were gavaged with self-made compound Yukui decoction 4.2 g/(kg·d),8.4 g/(kg·d)and 16.8 g/(kg·d),respectively.The general condition,disease activity index(DAI)score and histopathological score,the expression of inflammatory factors[tumor necrosis factor-α(TNF-α),interleukin-6(IL-6),interferon-γ(INF-γ)],oxidative stress indicators[superoxide dismutase(SOD),malondialdehyde(MDA),glutathione(GSH)],colonic mucosal barrier function proteins[occludin,zonula occluden-1(ZO-1)]and p-p38 MAPK/p38 MAPK and MMP-9 proteins expression changes of each group were compared.Results The DAI score and histopathological score of the low,medium,and high dose decoction groups were lower than those of the model group(2.55±0.50,1.50±0.32,0.82±0.25 vs 3.25±0.65,5.15±1.14,3.00±0.78,2.00±0.65 vs 6.33±1.30),and the DAI score and histopathological score of the low,medium,and high-dose decoction groups showed a dose-dependent decreasing trend(P<0.05).The levels of TNF-α,IL-6,IFN-γin the low,medium,and high dose decoction groups are all lower than the model group,and the levels of TNF-α,IL-6,IFN-γin the low,medium,and high dose decoction groups showed a dose-dependent decreasing trend(P<0.05).The levels of SOD and GSH in the low,medium,and high dose decoction groups were higher than those in the model group,while the levels of MDA were lower than that in the model group;and the levels of SOD and GSH in the low,medium,and high dose decoction groups showed a dose-dependent increasing trend,while the levels of MDA showed a dose-dependent decreasing trend(P<0.05).The expression levels of occludin and ZO-1 in the low,medium,and high dose decoction groups were higher than those in the model group,while the expression levels of p-p38 MAPK/p38 MAPK and MMP-9 were lower than those in the model group;the expression of occludin and ZO-1 in the low,medium,and high dose decoction groups showed a dose-dependent increasing trend,while the expression of p-p38 MAPK/p38 MAPK and MMP-9 showed a dose-dependent decreasing trend(P<0.05).Conclusion The compound Yukui decoction can inhibit the activation of p38 MARK signal pathway and the expression of MMP-9 protein,reduce the level of inflammatory factors in colon tissue of UC rats,relieve oxidative stress response,and improve the function of intestinal mucosal epithelial barrier.