清肠温中方通过miR-675-5p/VDR信号通路调控溃疡性结肠炎Th17/Treg免疫平衡及肠黏膜屏障的机制研究

Effect of Qingchang Wenzhong Decoction on miR-675-5p/VDR Signaling Pathway in Rats with DSS-induced Ulcerative Colitis

目的研究清肠温中方调控DSS诱导溃疡性结肠炎(UC)大鼠Th17/Treg免疫平衡及肠黏膜屏障的作用机制。方法 24只SPF级雄性SD大鼠,按体质量依照随机数字表将其随机分为空白组、模型组、清肠温中方组。空白组自由饮用去离子水,同时予去离子水灌胃;模型组和清肠温中方组自由饮用4.5%DSS溶液制备UC模型,同时模型组给予去离子水,清肠温中方组予清肠温中方灌胃。7 d后麻醉大鼠后取血及结肠组织,应用Real time-PCR法检测结肠miR-675-5p、VDR m RNA、RORγt mRNA、Foxp3 m RNA的表达,ELISA法检测血清IL-10、IL-17的表达,免疫组织化学法检测ZO-1、Occludin的蛋白表达及分布。结果与空白组比较,DSS诱导的UC大鼠结肠mi R-675-5p、RORγt mRNA及血清IL-17的表达较空白组明显升高(P <0.05或P <0.01),VDR mRNA、Foxp3 mRNA、ZO-1、Occludin、血清IL-10的表达较空白组明显降低(P <0.05或P <0.01);清肠温中方干预后,大鼠结肠mi R-675-5p、RORγt mRNA、血清IL-17的表达较模型组明显降低(P <0.05),VDR mRNA、Foxp3 mRNA、ZO-1、Occludin、血清IL-10的表达较模型组明显升高(P <0.05或P <0.01)。结论清肠温中方可能通过降低mi R-675-5p的表达,靶向调控VDR信号通路,从而调控溃疡性结肠炎Th17/Treg免疫平衡、修复肠黏膜屏障损伤,达到治疗UC的目的。

Objective:To explore the mechanism of regulating the immune balance of Th17/Treg and intestinal mucosal barrier in rats with DSS induced ulcerative colitis.Methods:24 SPF male SD rats were randomly divided into three groups according to their body weight:control group,model group and Qingchang Wenzhong Decoction group.Control group:drinking free deionized water,while deionized water was administered.The model group and the Qingchang Wenzhong Decoction group:free drinking 4.5%DSS solution to make UC model,while deionized water was administered in model group and Qingchang Wenzhong Decoction was administered in model group.After 7 days Intervention,blood and colonic tissue were taken from the anesthetized rats.Real time-PCR was used to detect the expression of miR-675-5p,VDR mRNA,RORγt mRNA,Foxp3 mRNA in colon,Elisa to detect the expression of serum IL-10,IL-17,and immunohistochemical method to detect the expression and distribution of ZO-1,Occludin protein.Results:Compared with the control group,the expression of miR-675-5p,RORγt mRNA and serum IL-17 in the colon of UC rats induced by DSS were significantly higher than those in the control group(P<0.05 or P<0.01)and the expression of VDR mRNA,Foxp3 mRNA,ZO-1,Occludin,serum IL-10 was significantly lower than that in the control group(P<0.05 or P<0.01).After the intervention of Qingchang Wenzhong Decoction,The expression of miR-675-5p,RORγt mRNA in colon and serum IL-17 of rats was significantly lower than those in model group(P<0.05)and the expression of VDR mRNA,Foxp3 mRNA,ZO-1,Occludin,serum IL-10 was significantly higher than those in the model group(P<0.05 or P<0.01).Conclusion:Qingchang Wenzhong Decoction may regulate the VDR signaling pathway by decreasing the expression of miR-675-5p,thereby regulating the immune balance of Th17/Treg in ulcerative colitis,repairing the intestinal mucosal barrier injury,and achieving the purpose of treating UC.