葛根素治疗冠状动脉支架内再狭窄的核心靶点筛选与细胞学验证  

Screening and cytological validation of core targets of puerarin in the treatment of coronary in-stent restenosis

作　　者：孙云雷 曹月娟[2] 赵振营[3] SUN Yunlei;CAO Yuejuan;ZHAO Zhenying(Graduate School of Tianjin University of Traditional Chinese Medicine,Tianjin 300000,China;不详)

机构地区：[1]天津中医药大学研究生院,天津300000 [2]天津市人民医院心内二科 [3]天津市人民医院药学部

出　　处：《山东医药》2023年第26期10-14,共5页Shandong Medical Journal

基　　金：天津市“131”创新型人才培养工程项目(2016)。

摘　　要：目的基于网络药理学、分子对接的方法筛选葛根素治疗冠状动脉支架内再狭窄(ISR)的核心靶点,并通过细胞学实验验证相关靶点。方法在NCBI公共数据平台基因表达综合数据库(GEO)中下载基因表达谱芯片数据GSE46560,使用R语言软件筛选该基因表达谱的差异基因,与Genecard数据库中ISR相关靶点整合,获得ISR疾病靶点。在TCMSP、SwissTargetPrediction网站下载葛根素的作用靶点,与ISR疾病靶点交集,获得药物—疾病关键靶点;使用String网站和Cytoscape软件分析蛋白—蛋白相互作用网络并筛选出核心靶点;使用AutoDockvina软件将核心靶点与葛根素的三维核心活性成分结构进行分子对接,计算分子对接核能值,评价配体分子与受体蛋白的结合能力。取大鼠胸主动脉血管平滑肌细胞进行培养,将细胞分为空白对照组、AngⅡ组、AngⅡ+葛根素组,采用细胞划痕实验观察各组6、12、24、48 h细胞迁移率;采用Western blotting法检测各组细胞过氧化物酶体增殖物激活受体γ(PPARγ)表达;采用ELISA法检测细胞炎症因子肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)含量。结果共获得ISR疾病相关靶点528个、葛根素作用靶点142个,将二者取交集,获得药物—疾病关键靶点56个。将56个疾病—药物关键靶点输入String11.5网站,获得疾病—药物核心靶点TNF、AKT1、SOD1、VEGFA、ABCB1、CASP3、PTGS2、PPARγ。分子对接显示,各核心靶点蛋白与葛根素中对应的活性化合物均有较好的结合能力。细胞实验表明,与空白对照组比较,AngⅡ组PPARγ蛋白表达降低,TNF-α、IL-6表达升高;与AngⅡ组比较,AngⅡ+葛根素组PPARγ表达升高,TNF-α、IL-6表达降低。结论共筛选得到PPARγ等7个葛根素治疗ISR的核心靶点;经验证,葛根素可能通过激活PPARγ、抑制炎症因子表达从而抑制血管平滑肌细胞增殖,从而达到治疗ISR的目的。Objective Based on the methods of network pharmacology and molecular docking,the core targets of pu⁃erarin in the treatment of coronary in-stent restenosis(ISR)were screened,and the related targets were verified by cytolog⁃ical experiments.Methods The gene expression profile data GSE46560 was downloaded from the NCBI public data plat⁃form gene expression comprehensive database(GEO).The differential genes of the gene expression profile were screened by R language software,and were integrated with ISR-related targets in Genecard database to obtain ISR disease targets.We downloaded the action target of puerarin on TCMSP and SwissTargetPrediction websites and intersected with ISR dis⁃ease target to obtain the drug-disease key target.We used String website and Cytoscape software to analyze the protein-pro⁃tein interaction network and screened the core target;AutoDockvina software was used to dock the core target gene with the three-dimensional core active component structure of puerarin,and we calculated the docking nuclear energy,and evaluat⁃ed the binding ability of ligand molecule and receptor protein.The vascular smooth muscle cells of rat thoracic aorta were cultured and divided into the blank control group,AngⅡgroup,and AngⅡ+puerarin group.The cell migration rates at 6,12,24 and 48 h were observed by Scratch test,and the expression of peroxisome proliferator-activated receptorγ(PPARγ)was detected by Western blotting.The content of tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)was measured by ELISA.Results A total of 528 ISR disease-related targets and 142 puerarin action targets were obtained,and 56 drug-disease key targets were obtained by intersection of the two.Fifty-six disease-drug key target genes were input into String11.5 website to obtain disease-drug core targets TNF,AKT1,SOD1,VEGFA,ABCB1,CASP3,PTGS2,and PPARγ.Molecular docking showed that all the core target proteins had good binding ability to the corresponding active compounds in puerarin.Cell experiments showed that comp

关 键 词：冠脉支架内再狭窄 葛根素 网络药理学 过氧化物酶体增殖物激活受体Γ 血管平滑肌细胞 细胞增殖 

分 类 号：R541.4[医药卫生—心血管疾病]