网络药理学结合谱效关系及分子对接探讨五味甘露抗炎物质基础及作用机制

Molecular mechanism of Tibetan medicine Wuwei Ganlu on anti-inflammatory effect based on network pharmacology combined with Sspectrum effect relationship and molecular docking

目的基于网络药理学结合谱效关系及分子对接,探讨五味甘露抗炎的物质基础及作用机制。方法利用中药系统药理学数据库与分析平台(TCMSP)、Genecards、Pubchem、UniProt、中国学术期刊全文数据库(CNKI)等数据库筛选五味甘露的活性成分及抗炎的作用靶点,利用STRING数据库、Cytoscape3.7软件构建蛋白质-蛋白质相互作用(PPI)网络并筛选核心靶点,利用Metascape数据库进行基因本体(GO)及京都基因与基因组百科全书(KEGG)通路富集分析。将细胞炎症模型与高效液相色谱法相结合分析五味甘露不同极性部位的抗炎活性和药效成分。采用Discovery Studio Lib Dock软件将主要药效成分与核心靶点进行分子对接,并进行可视化分析。结果网络药理学分析共获取五味甘露抗炎成分125个,潜在抗炎靶点251个,得到核心靶点131个,包括肿瘤坏死因子(TNF)、白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)重要靶点。KEGG通路富集分析,共富集到20条相关通路,其中涉及基因较多的包括脂质与动脉粥样硬化和类风湿性关节炎(RA)等通路,诸多通路均涉及NOD样受体热蛋白结构域相关蛋白3(NLRP3)核心靶点。五味甘露的醋酸乙酯相能够抑制炎症相关的NLRP3、IL-1β、TNF的表达,且其中总黄酮含量最高,为(305.0±11.4)mg·g^(-1)。LC-MS图谱分析和分子对接实验显示醋酸乙酯相中山柰酚、杨梅素、异荭草素、异槲皮苷和黄芪苷等黄酮类物质的离子强度远远高于单味药材且其母核结构与NLRP3受体蛋白的7个氨基酸活性位点有显著的相互作用。结论五味甘露抗炎的主要物质为黄酮类,可通过抑制NLRP3炎症小体的活化起到抗炎作用。

Objective To explore the material basis and mechanism of Wuwei Ganlu on anti-inflammatory based on network pharmacology combined with spectrum effect relationship and molecular docking.Methods The active ingredients and antiinflammatory targets of Wuwei Ganlu were obtained using databases such as TCMSP,Genecards,Pubchem,UniProt,and CNKI,then core targets and protein interaction networks were screened by STRING database and Cytoscape 3.7 software.Metascape was used to perform GO and KEGG signal pathway enrichment of core targets.Subsequently,the cellular inflammation model combined with high-performance liquid chromatography was used to analyze the anti-inflammatory activity and pharmacological components of different polar parts of Wuwei Ganlu.Finally,discovery Studio Lib Dock software was applied to connect the main pharmacological components with the core targets for molecular docking and visual analysis.Results Through network pharmacological analysis,a total of 125 anti-inflammatory components of Wuwei Ganlu and 251 potential anti-inflammatory targets,as well as 131 core targets were obtained,including TNF,IL-6,IL-1β.The KEGG pathway enrichment analysis enriched 20 related pathways,including lipid and atherosclerosis,and rheumatoid arthritis pathways,which all involved NLRP3 core targets.In addition,the ethyl acetate phase of Wuwei Ganlu can inhibit the expression of NLRP3,IL-1βand TNF with a total flavonoid content of(305±11.4)mg·g^(-1).LC-MS spectrum analysis and molecular docking experiments showed that the ion strength of flavonoids such as naphthol,myricetin,isoquercetin,isoquercetin,and astragaloside in the ethyl acetate phase was much higher than that of single medicinal herbs,and their mother nucleus structure had significant interactions with the seven amino acid active sites of the NLRP3 receptor protein.Conclusion The main anti-inflammatory substances of Wuwei Ganlu are flavonoids,which can exert antiinflammatory effects by inhibiting the activation of NLRP3 inflammasomes.