基于Notch信号通路研究白头翁汤对溃疡性结肠炎小鼠肠黏液屏障的保护作用  

Protective effect of Baitouweng Decoction on intestinal mucus barrier in mice with ulcerative colitis based on Notch signaling pathway

作　　者：刘伟[1,3] 刘又前 蒋翠花 张东建 方雨夕 龚旻 李寅庆 王甜甜 张健[1,2,3] LIU Wei;LIU You-qian;JIANG Cui-hua;ZHANG Dong-jian;FANG Yu-xi;GONG Min;LI Yin-qing;WANG Tian-tian;ZHANG Jian(School of Third Clinical Medicine,Nanjing University of Chinese Medicine,Nanjing 210028,China;Nanjing Lishui District Hospital of Traditional Chinese Medicine,Nanjing 211200,China;Affiliated Hospital of Integrated Traditional Chinese and Western Medicine,Nanjing University of Chinese Medicine,Nanjing 210028,China;School of Traditional Chinese Pharmacy,China Pharmaceutical University,Nanjing 210009,China;HeBei Yuzhilin Pharmaceutical Co.,Ltd.,Shijiazhuang 050000,China)

机构地区：[1]南京中医药大学第三临床医学院,江苏南京210028 [2]南京市溧水区中医院,江苏南京211200 [3]南京中医药大学附属中西医结合医院,江苏南京210028 [4]中国药科大学中药学院,江苏南京210009 [5]河北御芝林药业有限公司,河北石家庄050000

出　　处：《中草药》2023年第16期5257-5266,共10页Chinese Traditional and Herbal Drugs

基　　金：南京市卫生科技发展专项资金项目(YKK22241)。

摘　　要：目的探讨白头翁汤对葡聚糖硫酸钠(dextran sodium sulfate,DSS)诱导的溃疡性结肠炎(ulcerative colitis,UC)小鼠肠黏液屏障的保护作用以及对Notch通路的影响。方法C57BL/6小鼠随机分为对照组、模型组、美沙拉嗪(50 mg/kg)组和白头翁汤低、高剂量(7.4、14.8 g/kg)组,对照组小鼠自由饮用纯净水,其余各组连续8 d自由饮用2.5%DSS溶液诱导UC模型,造模同时各给药组ig相应药物。每日记录小鼠体质量、粪便稠度以及血便情况;采用苏木素-伊红(HE)染色观察结肠组织病理变化;采用AB-PAS染色观察结肠组织黏蛋白2(mucin 2,MUC2)的分泌情况;ELISA法检测结肠组织中髓过氧化物酶(myeloperoxidase,MPO)活性及促炎因子白细胞介素-6(interleukin-6,IL-6)、IL-1β、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)和抗炎因子IL-10的含量;免疫组化法检测结肠组织Ki67的表达情况;免疫荧光法测定结肠组织富含亮氨酸重复序列的G蛋白偶联受体5(leucine-rich repeat-containing G-protein-coupled receptor 5,Lgr5^(+))肠道干细胞(intestinal stem cells,ISCs)阳性细胞数;Western blotting测定结肠组织Notch-1、Atoh1、Hes1和MUC2蛋白表达。结果与模型组比较,白头翁汤能够缓解DSS诱导的UC小鼠体质量降低和疾病活动指数(disease activity index,DAI)升高(P<0.05、0.01、0.001),缓解结肠组织的缩短、水肿以及病理学损伤(P<0.01、0.001),减少炎性细胞浸润、隐窝的破坏与杯状细胞的丢失,降低结肠组织MPO活性和IL-1β、TNF-α水平(P<0.05、0.01),升高IL-10水平(P<0.05),增加结肠组织Ki67阳性细胞数(P<0.01),促进MUC2的分泌和Lgr5^(+)ISCs的阳性细胞数,并下调Notch和Hes-1蛋白表达水平(P<0.05、0.001),上调Atoh1和MUC2蛋白表达水平(P<0.05、0.01)。结论白头翁汤能够降低UC小鼠结肠组织促炎细胞因子水平,增加Lgr5^(+)ISCs的增殖和分化,保护隐窝和杯状细胞结构,促进MUC-2分泌,其作用可能与调控Notch-1/Atoh1/Hes1通路密Objective To explore the protective effect of Baitouweng Decoction(白头翁汤,BTWD)on intestinal mucus barrier in mice with ulcerative colitis(UC)induced by dextran sodium sulfate(DSS)and its effect on Notch pathway.Methods C57BL/6 mice were randomly divided into control group,model group,mesalazine(50 mg/kg)group and BTWD low-,high-dose(7.4,14.8 g/kg)groups.The mice in control group were free to drink pure water,and the other groups were free to drink 2.5% DSS solution for 8 d to induce UC model.At the same time,each group was given corresponding drugs.The body weight,fecal consistency and bloody stool of mice were recorded every day.HE staining was used to observe the pathological changes of colon tissue.The secretion of mucin 2(MUC2)in colon tissue was observed by AB-PAS staining.The activity of myeloperoxidase(MPO)and contents of pro-inflammatory factors interleukin-6(IL-6),IL-1β,tumor necrosis factor-α(TNF-α)and anti-inflammatory factor IL-10 in colon tissue were detected by ELISA.The expression of Ki67 in colon tissue was detected by immunohistochemistry.The number of positive cells of leucine-rich repeat-containing G-protein-coupled receptor 5(Lgr5^(+))intestinal stem cells(ISCs)in colon tissue was determined by immunofluorescence.Western blotting was used to detect Notch-1,Atoh1,Hes1 and MUC2 protein expressions in colon tissue.Results Compared with model group,BTWD alleviated the decrease of body weight and increase of disease activity index(DAI)in UC mice induced by DSS(P<0.05,0.01,0.001),alleviated the shortening,edema and pathological damage of colon tissue(P<0.01,0.001),reduced the infiltration of inflammatory cells,destruction of crypts and goblet cells,decreased MPO activity and IL-1β,TNF-α levels in colon tissue(P<0.05,0.01),increased IL-10 level(P<0.05),increased the number of Ki67 positive cells in colon tissue(P<0.01),promoted the secretion of MUC2 and the number of Lgr5^(+)ISCs positive cells,down-regulated Notch and Hes-1 protein expressions(P<0.05,0.001),up-regulated Atoh1 and MUC2 pr

关 键 词：白头翁汤 溃疡性结肠炎 肠道干细胞 黏液屏障 Notch通路 白头翁皂苷B4 黄柏碱 小檗碱 表小檗碱 黄连碱 巴马汀 秦皮甲素 秦皮乙素 

分 类 号：R285.5[医药卫生—中药学]