基于网络药理学及体外实验探讨补肺颗粒治疗支气管哮喘的分子机制  

Investigation of the molecular mechanism of Bufei Granules in treating bronchial asthma based on network pharmacology and in vitro experiments

作　　者：张莎苒 周玲羚 李一帆 李晓丹[2] 徐元雯[2] 王虹景 王禹霏 刘伟[2] ZHANG Sharan;ZHOU Lingling;LI Yifan;LI Xiaodan;XU Yuanwen;WANG Hongjing;WANG Yufei;LIU Wei(Graduate School,Tianjin University of Traditional Chinese Medicine,Tianjin301617,China;Department of Respiratory and Critical Care,the Second Affiliated Hospital of Tianjin University of Traditional Chinese Medicine,Tianjin300250,China)

机构地区：[1]天津中医药大学研究生院,天津301617 [2]天津中医药大学第二附属医院呼吸与危重症科,天津300250

出　　处：《中国医药导报》2025年第7期31-38,共8页China Medical Herald

基　　金：天津市教委科研计划项目(2021ZD018);天津市卫生健康委员会中医药重点领域项目(2022010)。

摘　　要：目的 通过网络药理学技术和体外实验探讨补肺颗粒治疗支气管哮喘(以下简称“哮喘”)的作用机制。方法通过TCMSP、Pub Chem、Swiss Target Prediction和Uni Prot数据库筛选补肺颗粒的有效成分及作用靶点;利用Gene Card、OMIM数据库检索哮喘的疾病靶点,选取成分与疾病的交集靶点作为蛋白靶点库,采用STRING获取交集靶点蛋白相互作用的网络,通过Cytoscape软件筛选核心靶点基因;通过DAVID数据库进行基因本体(GO)功能富集分析及京都基因与基因组数据库(KEGG)通路富集分析,微生信平台对结果进行可视化,并构建药物-成分-靶点网络图;利用Auto Dock Tools、Vina及Pymol对药物有效成分与核心靶点进行分子对接。利用CCK-8评价药物对细胞活性的影响,并将人类支气管平滑肌细胞分为空白组和给药组(100μg/ml),采用Western blot法、RT-q PCR、酶联免疫吸附试验法检测PDK1、Akt、PDE4D、c AMP表达水平。结果 补肺颗粒靶点897个,哮喘靶点1 321个,两者的交集靶点238个。GO富集分析得到生物过程729项、细胞组成76项、分子功能169项。KEGG通路主要包括PI3K-Akt信号通路、c AMP信号通路、MAPK信号通路等。关键靶点包括Akt1、TNF、PTGS2等,关键成分包括槲皮素、山柰酚、木犀草素等。分子对接显示,Akt1和PDE4D与槲皮素、山柰酚、木犀草素均具有较好的结合活性。细胞实验显示,补肺颗粒浓度为100μg/ml时,细胞存活率较0μg/ml时降低(P<0.01),给药组PDK1和PDE4D的m RNA表达量低于空白组,Akt的m RNA表达量高于空白组(P<0.05);给药组p-PDK1和p-PDE4D蛋白表达水平低于空白组,p-Akt的m RNA表达量高于空白组(P<0.05);给药组c AMP表达高于空白组(P<0.01)。结论 补肺颗粒可能通过槲皮素、山柰酚、木犀草素等成分,作用于Akt、PDE4D等靶点,调节PI3K-Akt、c AMP等信号通路,从而发挥治疗哮喘的作用。Objective To explore the mechanism of Bufei Granules in treating bronchial asthma(“asthma”for short)through network pharmacology and in vitro experiments.Methods Active components and targets were screened using TCMSP,PubChem,Swiss Target Prediction,and UniProt.Disease targets of asthma were obtained from GeneCard and OMIM databases.Intersection targets of ingredients and diseases were selected as the protein target library,and the network protein interaction of intersection tragets was obtained by STRING.DAVID was employed for gene ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses the WeChat platform visualized the results and constructed a drug-component-target network diagram,and molecular docking was performed using AutoDock,Vina,and Pymol.CCK-8 was used to evaluate the effect of drugs on cell activity,and human bronchial smooth muscle cells were divided into blank group and drug administration group(100μg/ml).Western blot,RT-qPCR,and enzyme linked immunosorbent assay were used to detect the expression levels of PDK1,Akt,PDE4D,and cAMP.Results There were 897 targets of Bufei Granules,1321 targets of asthma and 238 targets of their intersection.GO enrichment analysis obtained 729 biological processes,76 cell compositions,and 169 molecular functions.KEGG pathway includes PI3K-Akt,cAMP, and MAPK signaling, etc. Key targets include Akt1, TNF, PTGS2, and key components include quercetin, kaempferol, luteolin, etc. Molecular docking showed that Akt1 and PDE4D had good binding activity with quercetin, kaempferol, and luteolin. The cell experiment showed that when the concentration of Bufei Granules was 100 μg / ml, the cell survival rate was lower than that of 0 μg/ml (P<0.01). The mRNA expression of PDK1 and PDE4D in the administration group was lower than that in the blank group, and the mRNA expression of Akt was higher than that in the blank group (P<0.05). The protein expression of p-PDK1 and p-PDE4D in administration group was lower than that in blank group, and the mRNA e

关 键 词：PDK1/Akt/PDE4D信号通路 补肺颗粒 支气管哮喘 网络药理学 分子对接 实验验证 

分 类 号：R285.5[医药卫生—中药学]