基于Th17/Treg免疫平衡研究黄芩醇提物治疗溃疡性结肠炎的作用机制

Exploration of mechanism of ethanol extract of Scutellaria baicalensis Georgi in treating ulcerative colitis based on Th17/Treg immune balance

目的:探究黄芩醇提物调节免疫平衡治疗溃疡性结肠炎(ulcerative colitis,UC)的作用及机制。方法:将24只C57BL/6小鼠随机分为正常组、模型组、美沙拉嗪组(500 mg·kg^(-1))、黄芩醇提物(400 mg·kg^(-1))组,每组6只。采用2.5%葡聚糖硫酸钠(dextran sulfate sodium,DSS)的饮用水建立UC小鼠模型。每天观察各组小鼠体重及一般情况,d 6计算疾病活动指数(DAI);测量结肠长度;采用流式细胞术检测肠系膜淋巴结和脾脏中辅助性T细胞17(T helper 17,Th17)和调节性T细胞(regulatory T cells,Treg)比例;苏木精-伊红染色观察结肠组织损伤程度;酶联免疫吸附法检测结肠白细胞介素-6(interleukin,IL-6),IL-10和IL-1β的含量;蛋白免疫印迹法检测结肠Toll样受体4(Toll-like receptor 4,TLR4)、核因子κB p65(nuclear factor-kappa B,NF-κB p65)、p-NF-κB p65及核因子κB抑制蛋白-α(I kappaB-alpha,IκB-α)的表达水平;气相色谱-质谱联用检测短链脂肪酸的含量;高内涵成像技术观察RAW264.7细胞核转位情况。结果:黄芩醇提物可改善DSS诱导的小鼠结肠炎症状,恢复肠系膜淋巴结和脾脏中Th17/Treg免疫平衡,降低结肠组织中IL-1β和IL-6水平、提高IL-10表达水平,降低TLR4,NF-κB p65和p-NF-κB p65表达水平,提高IκB-α表达水平,提高肠道短链脂肪酸含量,抑制脂多糖诱导引起的核转位。结论:黄芩醇提物可以改善UC小鼠的症状,其机制可能是通过改善肠道稳态,抑制TLR4/NF-κB通路以恢复Th17/Treg免疫平衡。

Objective:To explore the effect and mechanism of alcohol extract of Scutellaria baicalensis Georgi on immune balance in treating ulcerative colitis(UC).Methods:24 C57BL/6 mice were randomly divided into control group,model group,mesalazine group,and Scutellaria baicalensis Georgi alcohol extract group(400 mg·kg^(-1),n=6 for each group).A mouse model of UC was established by drinking water containing 2.5%dextran sulfate sodium(DSS).The body weight and general condition of the mice in each group were observed every day,and the disease activity index(DAI)was calculated on the 6th day;the length of the colon was measured;the proportion of T helper 17(Th17)/regulatory T cells(Treg cells)in the mesenteric lymph nodes and spleen was detected by flow cytometry;hematoxylin-eosin staining was used to observe the degree of colon tissue damage;the levels of interleukin-6(IL-6),IL-10 and IL-1βin colon were detected by enzyme-linked immunosorbent assay;the expressions of Toll-like receptor 4(TLR4),nuclear factor-kappa B(NF-κB p65),p-NF-κB p65,and I kappaB-alpha(IκB-α)in colon were detected by Western blot;the main short chain fatty acids were detected by gas chromatography-mass spectrometry;high-content imaging was used to observe the nuclear translocation of RAW264.7 cells.Results:The alcohol extract of Scutellaria baicalensis Georgi improved the symptoms of colitis induced by DSS in mice,restored the Th17/Treg immune balance in mesenteric lymph nodes and spleen,reduced the levels of IL-1βand IL-6 in colon tissue,and increased the expression level of IL-10.It decreased the expressions of TLR4,NF-κB p65 and p-NF-κB p65,increased the expression of IκB-α,increased the content of intestinal short-chain fatty acids,and inhibited the nuclear translocation induced by LPS.Conclusion:The alcohol extract of Scutellaria baicalensis Georgi can improve the symptoms of UC mice,and the mechanism may be to restore Th17/Treg immune balance by improving intestinal homeostasis and inhibiting TLR4/NF-κB pathway.