基于网络药理学、分子对接及实验验证探讨黄柏治疗痛风的作用机制

Mechanism of Phellodendri Chinensis Cortex in the treatment of gout based on network pharmacology,molecular docking and experimental validation

采用网络药理学、分子对接和体外细胞实验探讨黄柏抗痛风(gout)的物质基础与潜在作用机制。首先通过TCMSP数据库获得黄柏主要活性成分及其对应作用靶点信息;通过GeneCards、OMIM、TTD数据库获得痛风相关疾病靶点;将黄柏有效成分对应靶点与痛风靶点取交集,借助STRING平台及Cytoscape3.9.0软件,绘制交集基因蛋白互作(PPI)网络图;利用基因注释与分析平台(Metascape)数据库对核心靶点进行基因本体(GO)功能及京都基因与基因组百科全书(KEGG)通路富集分析,通过微生信云平台对富集结果可视化;借助AutoDock Tools软件对核心成分及关键靶点基因进行分子对接,并对核心化学成分抗痛风炎症作用进行实验验证。共筛选出25个黄柏抗痛风活性成分和70个关键交集靶点,PPI网络分析获得5个关键靶点包括蛋白激酶B1(AKT1)、肿瘤坏死因子(TNF)、过氧化物酶体增生激活受体γ(PPARγ)、白介素6(IL-6)、前列腺素内过氧化物合酶(PTGS 2);GO功能和KEGG通路富集显示,黄柏作用于细胞迁移的正向调控、细胞分化的负调控、炎症反应等生物学过程,调控PI3K-Akt、MAPK等信号通路,进而发挥抗痛风作用。分子对接结果显示,黄柏的5个主要活性成分与关键靶点间存在分子结合位点且结合能较强,均小于-5 kcal/mol;体外实验显示核心化学成分对尿酸钠诱导的炎症反应有较好的抑制作用,本研究初步揭示了黄柏具有多种潜在的抗痛风活性成分,其作用机理可能是通过作用于多靶点和多通路来实现的。

To investigate the potential pharmacodynamic material basis and mechanism of Phellodendri Chinensis Cortex(PCC)in the treatment of gout by combining network pharmacology prediction,molecular docking validation and experimental validation.The active ingredients and action targets of PCC were obtained through TCMSP databases,gout-related disease targets were obtained from GeneCards,OMIM and TTD databases.The corresponding targets of the active ingredients of PCC were intersected with the gout targets,and the protein-protein interaction(PPI)network of the intersected genes was mapped with the help of STRING platform and Cytoscape 3.9.0 software.The analysis of gene ontology(GO)function and Kyoto gene and gene targets(KEGG)pathway enrichment were performed by using String and MetaScape databases and visually presented through the platform of bioinformatics.Furthermore,molecular docking technology was performed to validate the binding pattern and affinity between the key ingredients and the crucial targets by using AutoDock Tools software.A total of 25 active ingredients and 70 potential key targets for the treatment of gout was screened in PCC.The enrichment of GO function and KEGG pathway showed that PCC might positively regulate cell migration,negatively regulate cell differentiation,inflammatory response,positively regulate cell adhesion,protein phosphorylation,DNA transcription and other biological processes.The most crucial biotargets of PCC against gout were protein kinase B1(AKT1),tumor necrosis factor(TNF),peroxisome proliferative activated receptor gamma(PPARγ),interleukin-6(IL-6),prostaglandin-endoperoxide synthase 2(PTGS2),and KEGG enrichment analysis revealed that PCC possessed anti-gout activity by regulating PI3K-Akt signaling pathway,MAPK signaling pathway.The molecular docking results showed that the binding energy between the key biotargets and the five potential active components were much less than-5 kcal/mol.In vitro experiments showed that the core chemical components exhibited potent inhibitory effect on the inflammatory response induced by sodium urate crystal.This study initially revealed PCC has a variety of potential anti-gout active components,and its mechanism may be achieved by modulating multiple biotargets and multiple signal transduction pathways.