基于网络药理学和实验验证的麻黄细辛附子汤治疗变应性鼻炎的作用机制探讨

Exploring mechanism of action of Mahuang Xixin Fuzi Decoction in treatment of allergic rhinitis based on network pharmacology and experimental verification

目的基于网络药理学和分子对接技术探讨麻黄细辛附子汤治疗变应性鼻炎(AR)的作用机制,并结合体外细胞实验进行验证。方法运用TCMSP平台获取麻黄细辛附子汤的活性成分及作用靶点,通过GeneCards、OMIM、PharmGKB、TTD和DrugBank数据库检索AR疾病靶点。利用STRING平台构建蛋白质相互作用(PPI)网络。通过R语言进行基因本体(GO)和京都基因与基因组百科全书(KEGG)富集分析。利用AutoDock对关键化合物与核心靶基因进行对接验证。体外培养人鼻黏膜上皮细胞(HNEpC),使用酶联免疫吸附(ELISA)、流式细胞术、实时荧光定量PCR(qRTPCR)的方法验证麻黄细辛附子汤对炎症因子、细胞凋亡、部分关键靶点表达的影响。结果经筛选得到麻黄细辛附子汤活性成分62个,治疗AR靶点99个。PPI核心靶点包括前列腺素G/H合成酶2(PTGS2)、丝裂原活化蛋白激酶3(MAPK3)、基质金属蛋白酶-9(MMP-9)等。KEGG富集分析涉及与免疫炎症相关信号通路,与细胞增殖、凋亡相关信号通路,GO富集分析涉及对脂多糖反应等生物过程,细胞因子受体结合、细胞因子活性等分子功能。分子对接结果表明,活性成分与靶基因紧密结合。细胞实验结果显示,与模型组比较,麻黄细辛附子汤可以减少炎症因子白细胞介素-6(IL-6)、白细胞介素-8(IL-8)、肿瘤坏死因子-α(TNF-α)分泌(P<0.05、0.01),抑制细胞凋亡(P<0.01),降低PTGS2、MAPK3 mRNA表达、升高MMP-9 mRNA表达(P<0.05、0.01)。结论麻黄细辛附子汤可以通过多种活性成分作用于多靶点、多通路,调控炎症反应、免疫功能、细胞凋亡等途径发挥治疗AR的作用。

Objective To investigate the mechanism of action of Mahuang Xixin Fuzi Decoction in treatment of allergic rhinitis(AR)based on network pharmacology and molecular docking techniques,and to validate it in combination with in vitro cellular experiments.Methods The TCMSP platform was used to obtain the active ingredients and action targets of Mahuang Xixin Fuzi Decoction,and the AR disease targets were searched by GeneCards,OMIM,PharmGKB,TTD and DrugBank databases.The protein-protein interaction(PPI)network was established by STRING platform.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis were performed by R language.AutoDock was used to verify the docking of key compounds with core target genes.Human nasal epithelial cells(HNEpC)were cultured in vitro,and the effects of Mahuang Xixin Fuzi Decoction on inflammatory factors,apoptosis,and expression of some key targets were verified using ELISA,flow cytometry,and qRT-PCR.Results A total of 62 active components of Mahuang Xixin Fuzi Decoction and 99 therapeutic AR targets were obtained.The core targets of PPI included PTGS2,MAPK3,MMP-9,etc.KEGG enrichment analysis involved signaling pathways related to immune inflammation,cell proliferation and apoptosis,GO enrichment analysis involved biological processes such as response to lipopolysaccharide,cytokine receptor binding,cytokine activity.The molecular docking results showed that the active ingredients were tightly bound to the target genes.The results of cellular experiment showed that compared with the model group,Mahuang Xixin Fuzi Decoction could reduce the secretion of inflammatory factors IL-6,IL-8 and TNF-α(P<0.05,0.01),inhibit apoptosis(P<0.01),reduce PTGS2 and MAPK3 mRNA expression and elevate MMP-9 mRNA expression(P<0.05,0.01).Conclusion Mahuang Xixin Fuzi Decoction can act on multiple targets and pathways through multiple active ingredients to regulate inflammatory response,immune function,and apoptosis to exert therapeutic effects on AR.