白头翁汤通过调节T淋巴细胞平衡治疗溃疡性结肠炎小鼠的机制研究  

Study on the Mechanism of Baitouweng Decoction in the Treatment of Mice with Ulcerative Colitis by Regulating T-lymphocyte Homeostasis

作　　者：潘思敏 黄梦芬 王春丽 张美玲 周若愚 王青 罗霞[2] 周联[2] 侯江涛[3] 陈斌[3] PAN Simin;HUANG Mengfen;WANG Chunli;ZHANG Meiling;ZHOU Ruoyu;WANG Qing;LUO Xia;ZHOU Lian;HOU Jiangtao;CHEN Bin(The First Clinical Medical School,Guangzhou University of Chinese Medicine,Guangzhou 510405 Guangdong,China;School of Pharmaceutical Sciences,Guangzhou University of Chinese Medicine,Guangzhou 510006 Guangdong,China;Department of Gastroenterology and Hepatology,The First Affiliated Hospital of Guangzhou University of Chinese Medicine,Guangzhou 510405 Guangdong,China)

机构地区：[1]广州中医药大学第一临床医学院,广东广州510405 [2]广州中医药大学中药学院,广东广州510006 [3]广州中医药大学第一附属医院脾胃病科,广东广州510405

出　　处：《中药新药与临床药理》2023年第4期433-442,共10页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：国家自然科学基金面上项目(82074197);广州中医药大学第一附属医院创新强院资助项目(2019ⅡT25,2019ZD01);广州中医药大学一流学科建设项目(A3-0402-20-415-007);广东省研究生教育创新计划项目(2020JGXM031);广州市科技计划项目(202102010454)。

摘　　要：目的探讨白头翁汤(Baitouweng Decoction)通过调节T淋巴细胞平衡治疗溃疡性结肠炎(UC)小鼠的机制。方法将60只C57BL/6小鼠随机分为正常组、模型组、美沙拉秦组(500 mg·kg^(-1))及白头翁汤低、中、高剂量组(5、10、15 g·kg^(-1)),每组10只。除正常组外,其他各组小鼠均采用自由饮用3%葡聚糖硫酸钠(DSS)溶液,连续7 d,诱导建立UC小鼠模型;给药组小鼠造模同时按上述剂量灌胃给药(20 mL·kg^(-1)),正常组和模型组给予等体积蒸馏水,每日1次,连续给药10 d。实验期间记录小鼠体质量变化,观察粪便性状,检测并记录小鼠便血情况,进行疾病活动指数(DAI)评分;测量结肠长度及脾脏、胸腺湿质量,计算脾脏及胸腺指数;采用HE染色法观察结肠组织病理变化,并进行病理学评分;采用阿尔新蓝染色法观察杯状细胞黏蛋白的表达;采用流式细胞术检测小鼠肠系膜淋巴结及脾脏中辅助性T细胞1(Th1)、辅助性T细胞17(Th17)及调节性T细胞(Treg)细胞的比例;采用ELISA法检测小鼠结肠组织中炎性细胞因子肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、IL-6、γ干扰素(IFN-γ)、IL-17A、转化生长因子β(TGF-β)的表达;采用免疫荧光法检测结肠组织紧密连接蛋白ZO-1、Occludin的表达。结果与正常组比较,模型组小鼠的体质量显著降低(P<0.01),DAI评分显著升高(P<0.01);脾脏指数显著上升(P<0.01),胸腺指数显著下降(P<0.01),结肠长度显著缩短(P<0.01);结肠组织出现明显溃疡,上皮细胞大量脱落,腺体排列紊乱,黏膜层和黏膜下层有大量炎性细胞浸润,病理组织评分显著升高(P<0.01);肠系膜淋巴结及脾脏组织中CD4^(+)IFN-γ^(+)Th1、CD4^(+)IL-17A^(+)Th17细胞占比显著升高(P<0.05,P<0.01),而CD25^(+)FOXP3^(+)Treg细胞占比显著降低(P<0.01);结肠组织中TNF-α、IL-1β、IL-6、IL-17A和IFN-γ水平显著升高(P<0.01),TGF-β水平显著降低(P<0.01);结肠组织中的黏蛋白含量及紧密连接�Objective To investigate the mechanism of Baitouweng Decoction in treating mice with ulcerative colitis(UC)by regulating T-lymphocyte homeostasis.Methods Sixty C57BL/6 mice were randomly divided into the normal group,the model group,the Mesalazine group(500 mg·kg^(-1)),and the low-,medium-and high-dose groups(5,10 and 15 g·kg^(-1))of Baitouweng Decoction,with 10 mice in each group.The mice in all groups except the normal group were induced to establish the UC mouse model by drinking 3%DSS solution ad libitum for 7 consecutive days.The mice in the drug administration group were given the above dose(20 mL·kg^(-1))by gavage at the same time,and the normal and model groups were given an equal volume of distilled water once daily for 10 days.During the experiment,changes in body mass of the mice were recorded,faecal properties were observed,and blood in the faeces was detected and recorded.The mice were scored on disease activity index(DAI);the length of colon and the wet mass of spleen and thymus were measured and the spleen(thymus)index was calculated;the intestinal histopathological changes were observed by HE staining and the pathological score was calculated;the expression of cuprocyte mucin was observed by Alcian blue staining;the flow cytometry was used to detect proportions of helper T cells 1(Th1)and helper T cells 17(Th17)in the mesenteric lymph nodes and spleen of mice;the expressions of tumor necrosis factorα(TNF-α),interleukin 1β(IL-1β),IL-6,interferonγ(IFN-γ),IL-17A and transforming growth factorβ(TGF-β)were measured by ELISA in mouse colonic tissue.The expressions of ZO-1 and Occludin were detected by immunofluorescence method.Results Compared with normal group,body mass of mice in model group was significantly decreased(P<0.01),DAI score was significantly increased(P<0.01);spleen index was significantly increased(P<0.01),thymus index was significantly decreased(P<0.01),colon length was significantly shortened(P<0.01);there were obvious ulcers in the colon tissue,a large number of epithelial

关 键 词：白头翁汤 溃疡性结肠炎 辅助性T细胞1 辅助性T细胞17 调节性T细胞 肠上皮损伤 黏蛋白 紧密连接蛋白 小鼠 

分 类 号：R285.5[医药卫生—中药学]