基于网络药理学和动物实验探讨黄连解毒汤通过Bcl-2/Bax/Caspase-3信号通路抗结直肠腺瘤作用机制  

The Mechanism of Huanglian Jiedu Decoction(黄连解毒汤)Against Colorectal Adenoma Via Bcl-2/Bax/Caspase-3 Pathway Based on Network Pharmacology and Animal Experiments

作　　者：张艺凡 贾苏杰 刘静远 郎晓猛[2,3] 康欣 赵源[2,3] 刘龙辉 刘悦[1] 翟文静 胡博乾 刘建平 ZHANG Yifan;JIA Sujie;LIU Jingyuan;LANG Xiaomeng;KANG Xin;ZHAO Yuan;LIU Longhui;LIU Yue;ZHAI Wenjing;HU Boqian;LIU Jianping(Hebei University of Chinese Medicine,Shijiazhuan Hebei 050091,China;Hebei Provincial Hospital of Traditional Chinese Medicine,Shijiazhuan Hebei 050011,China;Key Laboratory of Integrated Chinese and Western Medicine for Gastroenterology Research,Shijiazhuan Hebei 050011,China)

机构地区：[1]河北中医药大学,河北石家庄050091 [2]河北省中医院,河北石家庄050011 [3]河北省中西医结合胃肠病研究重点实验室,河北石家庄050011

出　　处：《中医药导报》2024年第11期32-40,47,共10页Guiding Journal of Traditional Chinese Medicine and Pharmacy

基　　金：国家中医临床研究基地建设项目(国中医药办科技函〔2018〕131号);河北省自然科学基金项目(H2022423326);河北省中西医结合胃肠病研究重点实验室(SZX2021019);河北中医药大学研究生创新项目(XCXZZSS2024052)。

摘　　要：目的:研究黄连解毒汤通过调控Bcl-2/Bax/Caspase-3信号通路发挥抗结直肠腺瘤(CRA)的作用机制。方法:通过中药系统药理学数据库与分析平台(TCMSP)获取黄连解毒汤的活性成分及作用靶点;通过在线人类孟德尔遗传病数据库(OMIM)和人类基因组数据库(GeneCard)获取CRA疾病靶点;通过Venny 2.1软件获得黄连解毒汤与CRA的交集靶点;通过String数据库和Cytoscape 3.9.1软件绘制蛋白互作网络,并筛选黄连解毒汤治疗CRA的核心作用靶点;通过DAVID数据库对交集靶点进行基因本体(GO)富集分析和京都基因与基因组百科全书(KEGG)富集分析;通过Auto Dock Tools 1.5.6软件将前5位药物活性成分与前5位关键靶点进行分子对接验证。采用随机数字表法将60只小鼠分为正常组、模型组、黄连解毒汤低剂量组、黄连解毒汤中剂量组、黄连解毒汤高剂量组、阿司匹林组,每组10只。采用氧化偶氮甲烷(AOM)/葡聚糖硫酸钠(DSS)联合诱导小鼠CRA模型,同时各组予相应药物进行灌胃干预9周。苏木精-伊红染色法(HE)观察小鼠肠组织的病理变化;原位末端转移酶标记法(TUNEL)观察肠组织细胞凋亡;实时荧光定量聚合酶链式反应(Realtime PCR)检测腺瘤组织B淋巴细胞瘤-2(Bcl-2)mRNA、Bcl-2相关X蛋白(Bax)mRNA、细胞色素C(Cyt C)mRNA、胱天蛋白酶-9(Caspase-9)mRNA、胱天蛋白酶-3(Caspase-3)mRNA表达水平;蛋白免疫印迹法(Western blotting)法检测腺瘤组织Bcl-2、Bax、Cyt C、Caspase-9、Caspase-3蛋白的表达水平。结果:网络药理学共筛选得到黄连解毒汤80个活性成分,药物-疾病交集靶点170个;通过药物-成分-靶点-疾病调控网络的构建,筛选出槲皮素(quercetin)、β-谷甾醇(beta-sitosterol)、豆甾醇(Stigmasterol)、黄连素(berberine)、汉黄芩素(wogonin)等核心活性成分;通过构建蛋白互作网络,筛选出AKT1、JUN、HSP90AA1、CASP3、IL-6等关键靶点;GO与KEGG富集分析发现黄连解毒汤可能通�Objective:To study the effect of Huanglian Jiedu decoction on colorectal adenoma(CRA)by regulating Bcl-2/Bax/Caspase-3 pathway.Methods:The active ingredients and targets of Huanglian Jiedu decoction were obtained through TCMSP.CRA disease targets were obtained through the OMIM and GeneCard.The intersection target of Huanglian Jiedu decoction and CRA was obtained by Venny 2.1 software.The protein interaction network was mapped by String database and Cytoscape3.9.1 software,and the core anti-CRA target of Huanglian Jiadu decoction was screened.GO analysis and KEGG enrichment analysis were performed for the intersection targets through DAVID database.The top 5 key targets were verified by molecular docking with the top 5 active ingredients by Auto Dock Tools 1.5.6 software.Totally 60 mice were categorized into normal group,model group,Huanglian Jiedu decoction high-dose group(high-dose group),Huanglian Jiedu decoction medium-dose group(medium-dose group),Huanglian Jiedu decoction low-dose group(low-dose group)and aspirin group,with each group containing 10 mice.The CRA model of mice was induced by the combination of AOM and DSS,and each group was given corresponding drugs for 9 weeks of gastric lavage intervention.The pathological changes of intestinal tissue were observed by hematoxylin-eosin staining(HE).TUNEL was used to observe the apoptosis of intestinal tissue cells in each group.Realtime PCRwas used to detect Bcl-2 mRNA,Bax mRNA,Cyt C mRNA,Caspase-9 mRNA and Caspase-3 mRNA expression levels.Western blotinng was used to detect Bcl-2,Bax,Cyt C,Caspase-9 and Caspase-3 protein expression levels.Results:Totally 80 active ingredients and 170 drug-disease intersection targets of Huanglian Jiedu Decoction were obtained through network pharmacological screening.The core components of quercetin,β-sitosterol,stigmasterol,berberine and wogonin were selected through the construction of drug-composition-target-disease regulatory network.Key targets such as AKT1,JUN,HSP90AA1,CASP3 and IL-6 were screened by constructing pro

关 键 词：结直肠腺瘤 黄连解毒汤 Bcl-2/Bax/Caspase-3信号通路 细胞凋亡 网络药理学 分子对接 动物实验 小鼠 

分 类 号：R285.5[医药卫生—中药学]