基于网络药理学与实验验证探讨加味半夏厚朴汤防治食管癌的作用机制  

Mechanism of Modified Banxia Houpo Decoction in Prevention and Treatment of Esophageal Cancer Based on Network Pharmacology and Experimental Verification

作　　者：陈晓乐[1] 李永亮[1] 李明彦 李振华[1] 张勤生 CHEN Xiaole;LI Yongliang;LI Mingyan;LI Zhenhua;ZHANG Qinsheng(The Second Affiliated Hospital to Henan University of Chinese Medicine/Henan Provincial Hospital of Chinese Medicine,Zhengzhou Henan China 450002)

机构地区：[1]河南中医药大学第二附属医院/河南省中医院,河南郑州450002

出　　处：《中医学报》2024年第10期2191-2202,共12页Acta Chinese Medicine

基　　金：国家中医临床研究基地科研专项项目(2021JDZX2004);河南省科技厅科技攻关项目(222102310240,242102310551);河南省中医药学科领军人才培养项目{豫卫中医函[2021]8号}。

摘　　要：目的:运用网络药理学和分子对接预测加味半夏厚朴汤(modified Banxia Houpo decoction,MBHD)治疗食管鳞状上皮内瘤变(esophageal squamous intraepithelial neoplasia,ESIN)的潜在靶点及信号通路,并通过体内外实验加以验证。方法:检索MBHD和ESIN的靶点,并获取两者的交集靶点。构建蛋白质-蛋白质相互作用(protein-protein interaction,PPI)网络,通过聚类分析筛选MBHD治疗ESIN的核心靶点。针对核心靶点作京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)富集分析。构建“中药-成分-通路-靶点”网络,通过拓扑分析筛选MBHD治疗ESIN的关键靶点和关键成分,并进行分子对接验证。KYSE-150细胞分为对照组和MBHD组,采用克隆形成实验检测集落形成情况,流式细胞术检测细胞凋亡情况,Western blot、qRT-PCR法检测肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、受体相互作用蛋白激酶1(receptor-interacting protein kinase 1,RIPK1)、受体相互作用蛋白激酶3(receptor-interacting protein kinase 3,RIPK3)、混合系列蛋白激酶样结构域(mixed lineage kinase domain like protein,MLKL)表达水平。将15只小鼠平均分为正常组、模型组和治疗组,除正常组外,其余小鼠建立食管癌模型。治疗组灌胃MBHD,连续20周。采用Western blot、qRT-PCR法检测TNF-α、RIPK1、RIPK3、MLKL表达水平。结果:MBHD靶点896个,ESIN靶点1262个,两者的交集靶点279个。KEGG通路主要包括PI3K/Akt信号通路、C型凝集素受体信号通路、IL-17信号通路、T细胞受体信号通路等。关键靶点包括过氧化物酶体增殖物激活受体-γ(peroxisome proliferator-activated receptor gamma,PPARG)、丝裂原活化蛋白激酶3(mitogen-activated protein kinase 3,MAPK3)、丝裂原活化蛋白激酶14(mitogen-activated protein kinase 14,MAPK14)等,关键成分包括木犀草素、黄芩素、厚朴新酚等。分子对接显示,TNF与黄芩素、木犀草素均具有较好的结合活性。细胞Objective:To use network pharmacology and molecular docking to predict the potential targets and signaling pathways of modified Banxia Houpo Decoction(MBHD)in treatment of esophageal squamous intraepithelial neoplasia(ESIN),and to verify them through in vitro and in vitro experiments.Methods:The targets of MBHD and ESIN were retrieved,and the intersecting targets of the two were obtained.A protein-protein interaction(PPI)network was constructed,and the core targets of MBHD in the treatment of ESIN were screened by cluster analysis.Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis was performed for core targets.The"TCM-Component-Pathway-Target"network was constructed,and the key targets and key components of MBHD treatment for ESIN were screened through topological analysis,and the molecular docking was verified.KYSE-150 cells were divided into control group and MBHD group,colony formation assay was used to detect colony formation,flow cytometry was used to detect apoptosis,and tumor necrosis factor-α(TNF-α)and receptor-interacting protein kinase 1(RIPK1)were detected by Western blot and qRT-PCR,receptor-interacting protein kinase 3(RIPK3),and mixed lineage kinase domain like protein(MLKL)expression levels.Fifteen mice were divided into normal group,model group and treatment group,and in addition to the normal group,the rest of the mice were established as esophageal cancer models.The treatment group was given intragastric MBHD for 20 consecutive weeks.Western blot and qRT-PCR were used to detect the expression levels of TNF-α,RIPK1,RIPK3 and MLKL.Results:There were 896 MBHD targets,1262 ESIN targets,and 279 intersecting targets.The KEGG pathway mainly includes PI3K-Akt signaling pathway,C-type lectin receptor signaling pathway,IL-17 signaling pathway,and T cell receptor signaling pathway.Key targets include peroxisome proliferator-activated receptorγgamma(PPARG),mitogen-activated protein kinase 3(MAPK3),and mitogen-activated protein kinase 14 kinase 14,MAPK14),etc.,key components include luteol

关 键 词：加味半夏厚朴汤 食管鳞状上皮内瘤变 食管癌 RIPK1/RIPK3/MLKL信号通路 网络药理学 分子对接 小鼠 细胞 毛德西 

分 类 号：R285.5[医药卫生—中药学]