溃结改良方改善DSS诱导的UC小鼠肠道炎症反应的机制研究
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摘要
目的从效应T细胞活化的源头Treg、Th17细胞入手,探讨溃结改良方(Kuijie Gailiang Prescription,KGP)调节Th17和Treg细胞的平衡转化机制,揭示溃结改良方治疗溃疡性结肠炎(Ulcerative colitis,UC)的深层次作用机理及作用靶点。方法采用3.5%DSS诱导复制UC小鼠模型,并随机分为正常组、模型组、5-ASA组和溃结改良方低、高剂量组,每组8只。观察各组小鼠体质量、大便黏稠度及出血情况,计算疾病活动指数;结肠组织HE染色,观察远端结肠组织病理形态学变化;流式细胞术检测小鼠脾脏淋巴细胞Treg、Th17细胞水平。结果溃结改良方对UC小鼠具有治疗效应,其疾病活动指数明显降低(P<0.05),结肠病理改变得到改善。与正常组相比,模型组CD4~+CD25~+Foxp3~+Treg细胞、IL-10、Foxp3、Smad3下调,CD3~+CD4~+IL-17A~+Th17细胞、IL-17A、RORγt、STAT3蛋白上调,差异具有统计学意义(P<0.01);溃结改良方治疗后,CD4~+CD25~+Foxp3~+Treg细胞、IL-10、Foxp3、p-Smad3上调,CD3~+CD4~+IL-17A~+Th17细胞、IL-17A、RORγt、p-STAT3下降,与模型组比较差异具有统计学意义(P<0.01)。结论溃结改良方可能通过调控UC小鼠Treg/Th17免疫平衡起到治疗UC的作用。
OBJECTIVE To investigate the mechanism of balance transformation of Th17 and Treg by Kuijie Gailiang Prescription(KGP) and to investigate the role of KGP in the treatment of ulcerative colitis(UC) from the source of effector T cell activation Treg and Th17 cells. Deep-level mechanism of action and targets. METHODS The UC mouse model was induced by 3.5% DSS and randomly divided into normal group, model group, 5-ASA group and low and high dose KGP group, with 8 rats in each group. Observe the body mass, stool consistency and hemorrhage of each group, calculate the index of disease activity, observe the pathological changes of colon tissue by HE staining of colon tissue, and detect the levels of Treg and Th17 cells in mouse spleen by flow cytometry. RESULTS KGP had a therapeutic effect on UC mice. The disease activity index was significantly reduced(P<0.05), and the pathological changes of colon tissue were improved. Compared with the normal group, the levels of CD4~+CD25~+Foxp3~+ Treg, IL-10, Foxp3, and Smad3 were down-regulated, and the levels of CD3~+CD4~+IL-17 A~+Th17, IL-17 A, RORγt, and STAT3 were up-regulated in the model group(P<0.01); after KGP treatment, The levels of CD4~+CD25~+Foxp3~+ Treg, IL-10, Foxp3, and p-Smad3 were up-regulated, and the levels of CD3~+CD4~+IL-17 A~+Th17, IL-17 A, RORγt, and p-STAT3 were decreased. There was a statistically significant difference compared with the model group(P<0.01). CONCLUSION KGP may play a role in the treatment of UC by regulating Treg/Th17 immune balance in UC mice.
OBJECTIVE To investigate the mechanism of balance transformation of Th17 and Treg by Kuijie Gailiang Prescription(KGP) and to investigate the role of KGP in the treatment of ulcerative colitis(UC) from the source of effector T cell activation Treg and Th17 cells. Deep-level mechanism of action and targets. METHODS The UC mouse model was induced by 3.5% DSS and randomly divided into normal group, model group, 5-ASA group and low and high dose KGP group, with 8 rats in each group. Observe the body mass, stool consistency and hemorrhage of each group, calculate the index of disease activity, observe the pathological changes of colon tissue by HE staining of colon tissue, and detect the levels of Treg and Th17 cells in mouse spleen by flow cytometry. RESULTS KGP had a therapeutic effect on UC mice. The disease activity index was significantly reduced(P<0.05), and the pathological changes of colon tissue were improved. Compared with the normal group, the levels of CD4~+CD25~+Foxp3~+ Treg, IL-10, Foxp3, and Smad3 were down-regulated, and the levels of CD3~+CD4~+IL-17 A~+Th17, IL-17 A, RORγt, and STAT3 were up-regulated in the model group(P<0.01); after KGP treatment, The levels of CD4~+CD25~+Foxp3~+ Treg, IL-10, Foxp3, and p-Smad3 were up-regulated, and the levels of CD3~+CD4~+IL-17 A~+Th17, IL-17 A, RORγt, and p-STAT3 were decreased. There was a statistically significant difference compared with the model group(P<0.01). CONCLUSION KGP may play a role in the treatment of UC by regulating Treg/Th17 immune balance in UC mice.
引文
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[15] 张小艳,杨雯,张英,等.1,25-(OH)2D3和全反式视黄酸联合应用对溃疡性结肠炎小鼠TREG细胞Foxp3表达的影响[J].营养学报,2016,38(2):157-161.
[16] FONSECA-CAMARILLO G,FURUZAWA-CARBALLEDA J,YAMAMOTO-FURUSHO J K.Interleukin 35 (IL-35) and IL-37:Intestinal and peripheral expression by T and B regulatory cells in patients with Inflammatory Bowel Disease[J].Cytokine,2015,75(2):389-402.
[17] YAN Y,ZHAO N,HE X,et al.Mesenchymal stem cell expression of interleukin-35 protects against ulcerative colitis by suppressing mucosal immune responses[J].Cytotherapy,2018,20(7):911-918.
[2] BARBI J,PARDOLL D,PAN F.Metabolic control of the Treg/Th17 axis[J].Immunol Rev,2013,252(1):52-77.
[3] 陆琴,章蓓,张苏闽,等.溃结灌肠液治疗溃疡性结肠炎临床疗效回顾性多因素分析[J].时珍国医国药,2018,29(4):893-899.
[4] 宋亚玲,王红梅,倪付勇,等.金银花中酚酸类成分及其抗炎活性研究[J].中草药,2015,46(4):490-495.
[5] 俞浩,方艳夕,毛斌斌,等.地榆炮制前后水提物抗炎效果研究[J].中药材,2014,37(1):34-37.
[6] 时松,黄振,罗熠,等.白及多糖对小鼠溃疡性结肠炎的治疗作用[J].中国药科大学学报,2012,43(6):535-540.
[7] SABATINO AD,LENTI MV,GIUFFRIDA P,et al.New insights into immune mechanisms underlying autoimmune diseases of the gastrointestinal tract[J].Autoimmun Rev,2015,14(12):1161-1169.
[8] EASTAFFLEUNG N,MABARRACK N,BARBOUR A,et al.Foxp3+ regulatory T cells,Th17 effector cells,and cytokine environment in inflammatory bowel disease.[J].J Clin Immunol,2010,30(1):80-89.
[9] JIANG W,SU J,ZHANG X,et al.Elevated levels of Th17 cells and Th17-related cytokines are associated with disease activity in patients with inflammatory bowel disease[J].Inflamm Res,2014,63(11):943-950.
[10] 黄晓丽,张新,费先艳,等.普拉梭菌上清对葡聚糖酸钠诱导的溃疡性结肠炎小鼠Th17细胞及IL-17A的影响[J].中南大学学报(医学版),2015,40(12):1320-1326.
[11] WANG Y,WANG LL,YANG HY,et al.Interleukin-21 is associated with the severity of psoriasis vulgaris through promoting CD4+ T cells to differentiate into Th17 cells[J].Am J Transl Res,2016,8(7):3188.
[12] GE J,ZHANG X,LIU J,et al.Elevated expression of interleukin-21 and its correlation to T-cell subpopulation in patients with ulcerative colitis[J].Cent Eur J Immunol,2015,40(3):331-336.
[13] YU J,HE S,LIU P,et al.Interleukin21 promotes the development of ulcerative colitis and regulates the proliferation and secretion of follicular T helper cells in the colitides microenvironment[J].Mol Med Rep,2015,11(2):1049.
[14] WANG Y,LIU XP,ZHAO ZB,et al.Expression of CD4+ forehead box P3 (FOXP3)+ regulatory T cells in inflammatory bowel disease[J].J Dig Dis,2011,12(4):286-294.
[15] 张小艳,杨雯,张英,等.1,25-(OH)2D3和全反式视黄酸联合应用对溃疡性结肠炎小鼠TREG细胞Foxp3表达的影响[J].营养学报,2016,38(2):157-161.
[16] FONSECA-CAMARILLO G,FURUZAWA-CARBALLEDA J,YAMAMOTO-FURUSHO J K.Interleukin 35 (IL-35) and IL-37:Intestinal and peripheral expression by T and B regulatory cells in patients with Inflammatory Bowel Disease[J].Cytokine,2015,75(2):389-402.
[17] YAN Y,ZHAO N,HE X,et al.Mesenchymal stem cell expression of interleukin-35 protects against ulcerative colitis by suppressing mucosal immune responses[J].Cytotherapy,2018,20(7):911-918.