摘要
目的:通过网络药理学方法、分子对接技术和体外实验,探讨虎杖干预病毒性肺炎的可能作用机制。方法:通过TCMSP平台筛选中药虎杖的活性成分及其作用靶点,从GeneCards数据库筛选病毒性肺炎的相关靶点,取交集得到潜在作用靶点;通过STRING数据库结合Cytoscape 3.7.2软件对潜在作用靶点进行交集靶点蛋白互作(PPI)分析和拓扑学分析,得到虎杖核心成分和病毒性肺炎关键靶点。采用DAVID数据库对交集靶点进行GO功能分析和KEGG通路富集分析,结合Cytoscape 3.7.2软件构建“虎杖-活性成分-靶点-通路”调控网络。采用AutoDock Vina、OpenBabel、PyMOL等软件对虎杖活性成分和病毒性肺炎关键靶点进行分子对接。以脂多糖(LPS)诱导的急性肺损伤人Ⅱ型肺泡上皮细胞A549建立急性肺损伤模型,对网络药理学的研究结果进行验证。结果:共获得虎杖10个活性成分,对应188个作用靶点,病毒性肺炎靶点1216个,取药物与疾病交集靶点93个;槲皮素、木犀草素、毒扁豆次碱、β-谷甾醇等可能是虎杖的核心成分;丝氨酸/苏氨激酶1(Akt1)、血管内皮生长因子A(VEGFA)、肿瘤坏死因子(TNF)、白介素-6(IL6)、白介素-1β(IL1β)、JUN激酶(JUN)、前列腺素G/H合成酶2(PTGS2)、基质金属蛋白酶-9(MMP-9)、半脱氨酸蛋白酶-3(CASP3)等蛋白可能是虎杖干预病毒性肺炎的核心靶点;GO功能分析和KEGG通路富集分析发现,靶点蛋白在细胞外基质、线粒体、核浆、胞质等中通过调控细胞因子、生长因子、蛋白、蛋白激酶以及TNF、PI3K-Akt、Toll样受体等信号通路发挥作用。分子对接发现,虎杖5种核心成分与TNF、Akt1、CASP3、IL1β、JUN、TP53、VEGFA靶点均有强烈的结合能力,结合能<-7.0 kcal/mol,与IL6、PTGS2均有较好的结合能力,结合能<-5.0 kcal/mol,与MMP-9有一定的结合能力,结合能<-4.25 kcal/mol。体外实验结果表明,虎杖可以降低模型组TNF-α、IL-1β、IL-6的表达,与空白组比较,差异有统计学意义(P<0.01,P<0.05),可以抑制Toll样受体4(TLR4)、核因子-κB(NF-κB)表达,差异有统计学意义(P<0.05)。结论:虎杖可能通过调控TNF-α、IL-1β、IL-6、TLR4、NF-κB等靶点干预病毒性肺炎,发挥治疗作用。
Objective:To explore the possible mechanism of Huzhang in the intervention of viral pneumonia by network pharmacology,molecular docking technology and in vitro experiment.Methods:The active ingredients and their targets of action of Chinese medicine Huzhang were screened by TCMSP platform,and the related targets of viral pneumonia were screened from GeneCards database.Then the potential targets were obtained by intersection.By using STRING database combined with Cytoscape 3.7.2 software,protein-protein interaction(PPI)analysis of intersection targets and topological analysis were performed on potential targets,and the core components of Huzhang and key targets of viral pneumonia were obtained.GO function analysis and KEGG pathway enrichment analysis were performed on intersection targets by using DAVID database.The“Huzhang-active ingredient-target-pathway”regulatory network was constructed by combining Cytoscape 3.7.2 software.AutoDock Vina,OpenBabel,PyMOL and other software were used to perform molecular docking between the active components of Huzhang and the key targets of viral pneumonia.Acute lung injury model was established by lipopolysaccharide(LPS)-induced acute lung injury in human typeⅡalveolar epithelial cell A549,and the results of network pharmacology were verified.Results:A total of 10 active ingredients of Huzhang were obtained,corresponding to 188 targets of action and 1216 targets of viral pneumonia,and 93 intersection targets of drugs and diseases were taken.Quercetin,luteolin,physostigmine,β-sitosterol might be the core components of Huzhang.Serine/threonine kinase 1(Akt1),vascular endothelial growth factor A(VEGFA),tumor necrosis factor(TNF),interleukin-6(IL6),interleukin-1β(IL1β)JUN kinase(JUN),prostaglandin G/H synthetase 2(PTGS2),matrix metalloproteinase-9(MMP-9)and a cysteine protease-3(CASP3)might be the core targets of Huzhang in the intervention of viral pneumonia.GO functional analysis and KEGG pathway enrichment analysis showed that the target proteins played a role by regulating cytokines,growth factors,proteins,protein kinases,TNF,PI3K-Akt,Toll-like receptors and other signaling pathways in extracellular matrix,mitochondria,nucleoplasm,and cytoplasm.Molecular docking showed that the five core components of Huzhang had strong binding ability with TNF,Akt1,CASP3,IL1β,JUN,TP53 and VEGFA with binding energy<-7.0 kcal/mol,and had good binding ability with IL6 and PTGS2 with binding energy<-5.0 kcal/mol,and had a certain binding ability with MMP-9 with binding energy<-4.25 kcal/mol.The results of in vitro experiments showed that Huzhang could reduce the expressions of TNF-α,IL-1βand IL-6 in the model group,with the statistical difference(P<0.01,P<0.05),and could inhibit the expressions of Toll-like receptor 4(TLR4)and nuclear factor-κB(NF-κB),with the statistical difference(P<0.05).Conclusion:Huzhang may play a therapeutic role in the intervention of viral pneumonia by regulating targets such as TNF-α,IL-1β,IL-6,TLR4 and NF-κB.
作者
王志晓
史婉丽
岳宝森
赵峰
张炜华
Wang Zhixiao;Shi Wanli;Yue Baosen;Zhao Feng;Zhang Weihua(Xi′an Hospital of Traditional Chinese Medicine,Xi′an Shaanxi 710000)
出处
《山西中医药大学学报》
2024年第4期421-430,共10页
Journal of Shanxi University of Chinese Medicine
基金
陕西省卫生健康委员会课题(2021A001)
西安市卫生健康委员会课题(2021yb01)
陕西中医药大学课题(2021FS15)。
关键词
病毒性肺炎
网络药理学
分子对接
虎杖
体外实验
viral pneumonia
network pharmacology
molecular docking
Huzhang
in vitro experiment
