基于网络药理学、分子对接及实验验证探讨益气通脉散抗脑缺血再灌注损伤的作用机制  

Study on the Mechanism of Yiqi Tongmai Powder Against Cerebral Ischemia-Reperfusion Injury Based on Network Pharmacology,Molecular Docking and Experimental Verification

作　　者：宋鹏鹏 郭燕可 关东升[2] 马鸣[2] 崔应麟 SONG Pengpeng;GUO Yanke;GUAN Dongsheng;MA Ming;CUI Yinglin(Henan University of Traditional Chinese Medicine,Zhengzhou 450046,China;Henan Provincial Hospital of Traditional Chinese Medicine,Zhengzhou 450002 Henan,China;Henan Key Laboratory of Traditional Chinese Medicine for Prevention and Treatment of Cardiovascular and Cerebrovascular Diseases,Zhengzhou 450002 Henan,China)

机构地区：[1]河南中医药大学,河南郑州450046 [2]河南省中医院,河南郑州450002 [3]河南省中医药防治心脑血管病重点实验室,河南郑州450002

出　　处：《中药新药与临床药理》2024年第7期1016-1027,共12页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：国家自然科学基金项目(81573919,81673943);国家中医药管理局中医药科学技术研究专项课题(GZY-KJS-2021-017);河南省中医药科学研究专项重点项目(20-21ZY2205,2022ZYZD07);河南省科技攻关项目(212102311123)。

摘　　要：目的基于网络药理学、分子对接及实验验证探讨益气通脉散(人参、丹参、三七、水蛭、土鳖虫、大黄)抗脑缺血再灌注损伤(CIRI)的作用机制。方法(1)通过TCMSP、TCMID、ETCM数据库筛选益气通脉散各中药活性成分及其作用靶点;通过GeneCards、OMIM、TTD疾病数据库筛选CIRI疾病相关靶点;通过Venny 2.1在线平台获得上述靶点的交集靶点,即益气通脉散治疗CIRI的潜在作用靶点。通过Cytoscape 3.7.1软件构建“药物-活性成分-靶点”网络,并筛选出益气通脉散治疗CIRI的潜在活性成分。通过STRING 11.0数据库进行潜在作用靶点的蛋白互作(PPI)分析,筛选核心靶点。通过Metascape数据库对潜在作用靶点进行GO功能及KEGG通路富集分析。采用AutoDockTools 1.5.7软件对关键活性成分与核心靶点进行分子对接验证。(2)采用大脑中动脉阻塞再灌注(MCAO/R)术建立CIRI大鼠模型。将SD大鼠随机分为:假手术组、模型组、益气通脉散低剂量组(0.27 g·kg^(-1))、益气通脉散高剂量组(1.08 g·kg^(-1))、尼莫地平组(30 mg·kg^(-1)),每组10只。造模前3 d开始预给药,每日1次,连续7 d。采用改良神经功能缺损程度评分(mNSS)法对MCAO/R大鼠的神经功能缺损进行评估;TTC染色法检测脑梗死体积;尼氏染色法观察脑组织神经元受损情况;ELISA法检测大鼠血清炎症因子及氧化应激相关指标;TUNEL染色法检测脑组织细胞凋亡情况;Western Blot法检测脑组织中Bax、Bcl-2蛋白表达水平。结果(1)共得到益气通脉散的46个有效活性成分,178个益气通脉散治疗CIRI的潜在作用靶点。分析出槲皮素、毛地黄黄酮、山柰酚、缬氨酸、尿嘧啶等关键活性成分;TNF、IL-6、STAT3、VEGFA、AKT1、IL-1β、CASP3、TP53、MAPK3、EGFR等核心靶点;潜在作用靶点参与炎症反应、氧化应激、细胞增殖分化、细胞凋亡等生物过程,涉及cAMP、NF-κB、PI3K-Akt等信号通路。主要活性成分槲皮素、�Objective To investigate the mechanism of Yiqi Tongmai Powder(Ginseng Radix et Rhizoma,Salviae Miltiorrhizae Radix et Rhizoma,Notoginseng Radix et Rhizoma,Hirudo,Eupolyphaga Steleophaga,Rhei Radix et Rhizoma)against cerebral ischemia-reperfusion injury(CIRI)based on network pharmacology,molecular docking and experimental verification.Methods(1)The active components of Yiqi Tongmai Powder and their action targets were screened by TCMSP,TCMID and ETCM databases,the disease related targets of CIRI were screened by GeneCards,OMIM and TTD disease databases,and the intersection targets of the above targets were obtained through Venny 2.1 online platform,that is,the potential targets of Yiqi Tongmai Powder in the treatment of CIRI.The“drugs-active components-targets”network was constructed by Cytoscape 3.7.1 software,and the potential active components of Yiqi Tongmai Powder in the treatment of CIRI were screened.Protein-protein interaction(PPI)analysis of potential targets was carried out by STRING 11.0 database to screen core targets.The GO function and KEGG pathway enrichment of potential targets were analyzed by Metascape database.AutoDockTools 1.5.7 software was used to verify the molecular docking between the key active components and the core targets.(2)The rat model of CIRI was established by middle cerebral artery occlusion and reperfusion(MCAO/R).SD rats were randomly divided into sham operation group,model group,Yiqi Tongmai Powder low-dose group(0.27 g·kg^(-1)),Yiqi Tongmai Powder high-dose group(1.08 g·kg^(-1))and Nimodipine group(30 mg·kg^(-1)),with 10 rats in each group.Pre-administration began three days before the establishment of the model,once a day for seven days.The neurological deficit of MCAO/R rats was evaluated by modified neurological deficit score(mNSS).The volume of cerebral infarction was measured by TTC staining.Nissl staining was used to observe the damage of neurons in brain tissue.ELISA method was used to detect serum inflammatory factors and oxidative stress related indexes.TUNEL

关 键 词：益气通脉散 脑缺血再灌注损伤 网络药理学 分子对接 炎症反应 氧化应激 细胞凋亡 大鼠 

分 类 号：R285.5[医药卫生—中药学] R857.3[医药卫生—中医学]