基于网络药理学和实验验证探究黄精增强免疫功能的作用机制

Network Pharmacology and Experimental Verification Reveal the Mechanism of Immunity Enhancement by Polygonati Rhizoma

目的:探究黄精增强免疫功能的作用机制。方法:使用中药系统药理学数据库与分析平台(TCMSP)收集黄精活性成分,通过SwissTargetPrediction数据库获得对应靶点基因。使用GeneCards、DrugBank数据库筛选免疫抑制(IS)相关靶点基因,并通过Venny数据库获得与黄精靶点基因的交集基因。使用String数据库构建蛋白质-蛋白质相互作用(PPI)网络,使用Cytoscape软件构建“黄精-有效成分-靶基因-免疫抑制”网络,利用Metascape数据库对交集基因进行基因本体(GO)富集分析以及京都基因和基因组百科全书(KEGG)富集分析,使用分子对接对有效成分和靶点进行验证。体外细胞试验采用脂多糖(LPS)诱导的RAW264.7细胞模型,通过MTT法检测不同浓度黄精提取物对细胞的毒性反应;采用Griess法检测黄精提取物对细胞中一氧化氮(NO)的影响。结果:经过筛选,ESR1、SRC、IGF1R、PTGS1、PTGS2等均参与了多个关键生物过程和关键通路。GO富集表明,细胞活化、炎症反应、细胞对应激的反应等为主要生物过程;KEGG富集表明,糖尿病并发症中的AGE-RAGE信号通路、AMPK、核因子κB信号通路等是关键通路。分子对接结果显示有效成分与核心靶点对接结果良好。体外试验结果显示,与LPS组比较,50,200,800μg/mL的黄精提取物组RAW264.7细胞中NO生成量均显著降低(均P<0.01)。结论:本研究初步预测了黄精增强免疫功能的ESR1、SRC等作用靶点和AMPK、核因子κB等信号通路,并结合Griess法表明黄精可通过调控炎症模型RAW264.7细胞NO的释放来发挥免疫作用。

Objective:To explore the mechanism of immunity enhancement by Polygonati Rhizoma.Methods:The active components of Polygonati Rhizoma were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP),and SwissTargetPrediction was used to predict the corresponding target genes.GeneCards and DrugBank were employed to screen out the target genes of immunosuppression(IS).The Venn diagram was established to obtain the common genes shared by Polygonati Rhizoma and IS.String was employed to construct the protein-protein interaction network,and Cytoscape was used to construct the “Polygonati Rhizoma-active component-target gene-IS” network.Metascape was used to conduct gene ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analyses on the common genes.Molecular docking was adopted to verify the binding between active components and targets.The RAW264.7 cell model induced by lipopolysaccharide(LPS) was established.The methyl thiazolyl tetrazolium(MTT) method was used to examine the toxicity of Polygonati Rhizoma extracts at different concentrations to cells.The Griess method was used to test the effect of Polygonati Rhizoma on nitric oxide(NO) in cells.Results:ESR1,SRC,IGF1R,PTGS1,and PTGS2 were predicted to be involved in key biological processes and pathways.GO enrichment showed that cell activation,inflammatory response,and cell response to stress were the main biological processes involved.KEGG enrichment showed that the AGE-RAGE signaling pathway in diabetic complications,AMPK,and nuclear factor-kappa B(NF-κB) signaling pathways were the critical pathways.Molecular docking results showed strong binding affinity between active components and core targets.Compared with the LPS group,the Polygonati Rhizoma extract groups(50,200,and 800 μg/mL) showed reduced production of NO in the RAW264.7 cells(P<0.01).Conclusion:This study preliminarily predicted the targets such as ESR1 and SRC and signaling pathways such as AMPK and NF-κB of Polygonati Rhizoma in enhancing immunity.Polygonati Rhizoma can enhance immunity by regulating the release of NO in RAW264.7 cells.