人参皂苷Rg1治疗多发性骨髓瘤的网络药理学预测与验证

Network target prediction and mechanism confirmation of ginsenoside Rg1 in treating multiple myeloma

目的:利用网络药理学和体外细胞实验,探究人参皂苷Rg1治疗多发性骨髓瘤(MM)的作用机制。方法:利用TTD、DisGeNet、GeneCards、PharmGKB、PubChem、Super-PRED、PharmMapper预测MM靶标,利用CTD和SymMap数据库预测人参皂苷Rg1靶标,取交集靶点构建蛋白质-蛋白质相互作用网络。基于基因本体(GO)和KEGG-pathway数据库,富集Rg1治疗MM的主要生物功能和信号通路。分子对接验证核心靶点与Rg1之间的结合能力。利用CCK-8、流式细胞术、Western blot实验,分别检测不同剂量人参皂苷Rg1(5、10、20μmol/L)对RPMI 8226细胞的细胞活性、凋亡率、ROS水平及核心作用靶点Caspase3、Bax、Bcl-2、和NF-κB p65的相对表达。结果:筛选得到215个Rg1-MM靶点。GO和KEGG富集分析提示Rg1改善MM的作用机制可能涉及氧化应激、细胞凋亡及炎症相关信号通路。分子对接提示Rg1可能通过调控NFKB1、STAT3、AKT1、MAPK3、CASP3、BCL2等靶点发挥效用。体外实验表明,与对照组比较,Rg1组(5、10、20μmol/L)可显著抑制RPMI 8226细胞增殖(P<0.01)、促进细胞凋亡(P<0.01)和ROS产生(P<0.01),同时抑制NF-κB信号通路的活化(P<0.05)。结论:人参皂苷Rg1对MM有潜在的治疗作用,能够调节RPMI 8226细胞的氧化应激、细胞凋亡和增殖,其作用机制涉及对NF-κB信号通路的调控。

Objective:To explore the mechanism of ginsenoside Rg1 in the treatment of multiple myeloma(MM)by using network pharmacology and in vitro cell experiments.Methods:TTD,DisGeNet,GeneCards,PharmGKB,PubChem,Super-PRED and PharmMapper were used to predict the MM target.Ginsenoside Rg1 target was predicted by CTD and SymMap database.Protein-protein interaction network of MM and Rg1 intersection targets was constructed by STRING database.Gene Ontology(GO)and KEGG-pathway database were used to enrich the main biological functions and signal pathways of Rg1 in treating MM.Molecular docking was used to verify the binding activity of the core target with Rg1.CCK-8,flow cytometry and Western blot experiments were used to detect the cell activity,apoptosis rate,ROS level and the relative expression of core action targets Caspase3,Bax,Bcl-2 and NF-κB p65 in RPMI 8226 cell at different doses of ginsenoside Rg1(5,10 and 20μmol/L).Results:A total of 215 Rg1-MM targets were screened.The enrichment analysis of GO and KEGG pathway suggested that the mechanism of Rg1 in treating MM might involve oxidative stress,apoptosis and inflammation-related signaling pathways.Molecular docking suggested that Rg1 might play a role by regulating NFKB1,STAT3,AKT1,MAPK3,CASP3,BCL2 and other targets.In vitro experiments,compared with the control group,Rg1(5,10 and 20μmol/L)significantly inhibited the proliferation(P<0.01)of RPMI 8226 cell,promoted apoptosis(P<0.01)and ROS production(P<0.01),and inhibited the activation of NF-κB signaling pathway(P<0.05).Conclusion:Ginsenoside Rg1 has a potential therapeutic effect on MM,which can regulate oxidative stress,apoptosis and proliferation of RPMI 8226 cell,and its mechanism involves the regulation of NF-κB signaling pathway.