四神丸加减方对溃疡性结肠炎模型小鼠结肠组织白细胞介素-6、白细胞介素-10及TLR4/NF-κB信号通路的影响

Effect of Modified Sishen Pills on Interleukin-6,Interleukin-10 and TLR4/NF-κB Signaling Pathway in Colonic Tissue of Mice Model of Ulcerative Colitis

目的:探讨四神丸加减方治疗溃疡性结肠炎的疗效机制。方法:选择30只雌性C57BL/6小鼠为实验对象,将小鼠随机分为正常组、模型组、阳性对照组、四神丸加减方高剂量组(简称高剂量组)和四神丸加减方低剂量组(简称低剂量组)各6只。在适应性喂养1周后,构建葡聚糖硫酸钠诱导的溃疡性结肠炎小鼠模型,并灌胃给药,实验期间每天观察小鼠精神状态并称重。实验结束后,比较5组小鼠疾病活动指数(DAI),结肠组织病理学指标、白细胞介素-6(IL-6)和白细胞介素-10(IL-10)表达水平、Toll样受体4(TLR4)、p65分子量、磷酸化P65(p-p65)、β-actin蛋白表达水平。结果:与正常组比较,模型组小鼠DAI评分、粪便黏稠度评分、便血评分、IL-6水平、结肠组织中TLR4、p65及p-p65表达水平均显著升高(P<0.05),结肠长度、IL-10水平均显著降低(P<0.05)。与模型组比较,阳性对照组和高剂量组小鼠DAI评分均显著下降(P<0.05),而低剂量组小鼠DAI评分无明显变化(P>0.05);高剂量组小鼠结肠长度明显增加(P<0.05),阳性对照组、低剂量组小鼠结肠长度增加不明显(P>0.05);阳性对照组及高、低剂量组小鼠粪便黏稠度评分均明显降低(P<0.05);阳性对照组及高、低剂量组小鼠便血评分无明显变化(P>0.05);高剂量组IL-6水平显著降低(P<0.05),阳性对照组及低剂量组IL-6水平下降不明显(P>0.05);高、低剂量组IL-10水平显著升高(P<0.05),阳性对照组IL-10水平升高不明显(P>0.05);阳性对照组、高剂量组、低剂量组小鼠结肠组织中TLR4、p65和p-p65的表达水平均明显降低(P<0.05)。病理变化上,模型组小鼠结肠组织中杯状细胞明显减少,隐窝结构损伤严重,出现明显的炎症浸润;与模型组比较,阳性对照组、高剂量组小鼠结肠组织上述病理变化改善明显,而低剂量组上述病理病化略有改善。结论:四神丸加减方可在一定程度上改善溃疡性结肠炎小鼠的临床症状及结肠组织病理变化,其作用机制可能与TLR4/NF-κB信号通路有关。

Objective:To explore the curative effect mechanism of modified Sishen Pills for ulcerative colitis.Methods:A total of 30 female C57BL/6 mice were selected as experimental subjects and randomly divided into the normal group,the model group,the positive control group,the high-dose group of modified Sishen pills(the hight-dose group) and the low-dose group of modified Sishen Pills(the low-dose group),with 6 cases in each group.After 1 week of adaptive feeding,a model of ulcerative colitis induced by sodium dextran sulfate was constructed and given by intragastric administration.The mental state of the mice was observed and the mice were weighed every day during the experiment.After the experiment,the Disease Activity Index(DAI),pathological indexes of colon tissue,and the expression levels of interleukin-6(IL-6),interleukin-10(IL-10),Toll-like receptor 4(TLR4),p65 and phosphorylated P65(p-p65),and β-actin protein were compared among the five groups.Results:Compared with those in the normal group,DAI score,fecal viscosity score,blood stool score,IL-6 level,and expression levels of TLR4,p65 and p-p65 in colonic tissue in the model group were significantly increased(P<0.05),and colon length and IL-10 level were significantly decreased(P<0.05).Compared with those in the model group,DAI score in the positive control group and the high-dose group were significantly decreased(P<0.05),and there was no significant change in DAI score in low-dose group(P>0.05);colon length in the high-dose group was significantly increased(P<0.05),and colon length in the positive control group and the low-dose group were not significantly increased(P>0.05);fecal viscosity scores in the positive control group,the high-dose group and the low-dose group were significantly decreased(P<0.05);there was no significant change in blood stool scores in the positive control group,the high-dose group and the lowdose group(P>0.05);IL-6 level in the high-dose group was significantly decreased(P<0.05),and the IL-6 level in the positive control group and the low-dose group did not decrease significantly(P>0.05);IL-10 levels in the high-and low-dose groups were significantly increased(P<0.05),and the level of IL-10 in the positive control group was not significantly increased(P>0.05);the relative expression levels of TLR4,p65 and p-p65 in colonic tissue in the positive control group,the high-dose group and the lowdose group were significantly decreased(P<0.05).In terms of pathological changes,goblet cells in colonic tissue in the model group were significantly reduced,crypt structure was seriously damaged,and there was obvious inflammatory infiltration.Compared with those in the model group,the above pathological changes of colonic tissue in the positive control group and the high-dose group were significantly improved,and the above pathological changes in the low-dose group were slightly improved.Conclusion:Modified Sishen Pills for mice with ulcerative colitis can improve their clinical symptoms and pathological indexes of colon tissue to a certain extent,and its mechanism may be related to TLR4/NF-κB signaling pathway.