补肾活血方治疗特发性肺间质纤维化的网络药理学研究与实验验证

Network Pharmacology Analysis of Bushen Huoxue Formula in the Treatment of Idiopathic Pulmonary Fibrosis and Experimental Verification

目的:基于网络药理学系统挖掘补肾活血方治疗特发性肺间质纤维化(IPF)的分子作用机制。方法:利用数据库寻找补肾活血方活性成分的作用靶点和疾病靶点,构建蛋白质-蛋白质相互作用(PPI)网络;进行基因本体(GO)功能与京都基因和基因组百科全书(KEGG)通路富集分析;构建生物网络,并利用分子对接技术对关键靶点进行验证;比较吡非尼酮和尼达尼布与该方活性成分在共同靶点及通路上的结合力。进一步采用体外实验验证。结果:通过筛选得到胱天蛋白酶3(CASP3)、肿瘤坏死因子(TNF)、Jun激酶(JNK)等50个核心靶点,富集通路有80条,包括白细胞介素-17(IL-17)信号通路等,分子对接构象良好的活性成分有汉黄芩素、千层纸素等。此外,在吡非尼酮、尼达尼布发挥作用的关键靶点上,补肾活血方中的白藜芦醇等活性成分与这些靶点结合,表现出了与之类似的结合力。体外实验证明,补肾活血方通过抑制IL-17A,激活细胞自噬,进一步降解胶原蛋白起到抗纤维化作用,验证了部分网络药理学预测结果。结论:补肾活血方中的活性成分靶点及作用通路在对炎症反应、免疫、新陈代谢等方面表现出显著富集,该方的抗肺纤维化作用可能与抑制IL-17A的表达,激活IL-17A介导的细胞自噬通路,降解胶原蛋白有关。

Objective:To explore the molecular mechanisms underlying the therapeutic effect of Bushen Huoxue Formula in the treatment of idiopathic pulmonary fibrosis(IPF)using network pharmacology.Methods:The targets of active ingredients of Bushen Huoxue Formula and disease targets were identified by searching databases.A protein-protein interaction(PPI)network was constructed,and gene ontology(GO)functional and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analyses were performed.A biological network was constructed,and molecular docking was conducted to validate the key targets.The binding affinities of pirfenidone and nintedanib to the active ingredients and shared targets and pathways were compared.Further in vitro experiments were conducted for validation.Results:Fifty core targets,including caspase-3(CASP3),tumor necrosis factor(TNF),and c-Jun N-terminal kinase(JNK),were identified.Eighty pathways were enriched,including the interleukin-17(IL-17)signaling pathway.Active ingredients such as wogonin and oroxylin A exhibited good molecular docking conformations.Furthermore,the active ingredients in Bushen Huoxue Formula,such as resveratrol,showed similar binding affinities to the key targets that had effect with pirfenidone and nintedanib.In vitro experiments demonstrated that Bushen Huoxue Formula exerted anti-fibrotic effects by inhibiting IL-17A,activating autophagy,and further degrading collagen.These findings validated some of the predictions made by network pharmacology.Conclusion:The targets of active ingredients and pathways in Bushen Huoxue Formula showed significant enrichment in inflammatory response,immunity,metabolism,etc.The anti-fibrotic effect of Bushen Huoxue Formula may be attributed to the inhibition of IL-17A expression,activation of IL-17A-mediated autophagy pathways,and degradation of collagen.