基于网络药理学和实验验证探讨白藜芦醇对髓系白血病细胞的作用机制

Exploring the mechanism of action of resveratrol on myeloid leukemia cells based on network pharmacology and experimental validation

目的:通过各种药理学公共数据库平台和实验验证探讨白藜芦醇(Res)对髓系白血病细胞的作用机制。方法:运用R语言软件(4.0)的Venny包和clusterprofiler包分别获取在TCMSP、PubChem、Swiss Target Prediction公共平台上Res和髓系白血病的相关基因的共同靶点基因以及两者共同靶点基因可能富集的信号通路和细胞组成、分子功能和生物过程。从String数据库导出靶点基因相关文件并通过Cytoscape 3.7.2软件筛选关键靶点蛋白,建立Res—关键靶点—信号通路—髓系白血病网络模型。对关键靶点蛋白与Res进行分子对接。分别采用Western blotting法、CCK-8法和流式细胞术检测不同浓度Res(50μmol/L、100μmol/L)对慢性髓系白血病K562细胞Ras蛋白表达、细胞增殖和细胞凋亡的影响。结果:共取得Res抗髓系白血病共同靶点14个,富集分析获得122条通路(P<0.05),其中富集靶点较多的主要信号通路有Ras信号通路(Ras signaling pathway)、Neurotrophin信号通路(Neurotrophin signaling pathway)等,得到22个分子功能、13个细胞组成、89个生物过程。Ras蛋白与Res在THR-74、GLN-150、GLU-143之间形成结合能最低的氢键。Res能够下调K562细胞Ras蛋白表达,抑制细胞增殖,促进凋亡,且100μmol/L Res作用更为显著(均P<0.05)。结论:Res可能通过作用于Ras靶点,介导Ras信号通路,对髓系白血病的治疗发挥积极作用,同时可能影响白血病细胞的增殖和凋亡。

Objective:To investigate the mechanism of action of resveratrol(Res)on myeloid leukemia cells through various public pharmacological databases and experimental validation.Methods:The Venny package and clusterprofiler package of R language software(4.0)were used to obtain the common target genes of Res and myeloid leukemia-related genes respectively,as well as the possible enriched signaling pathways,cellular compositions,molecular functions,and biological processes from the common target genes on TCMSP,PubChem,and Swiss Target Prediction public platforms.The target gene-related files were exported from the String database and the key target proteins were screened by Cytoscape 3.7.2 software to establish the Res-key target-signaling pathway-myeloid leukemia network model.Molecular docking of key target proteins to Res was performed.The effects of different concentrations of Res(50μmol/L,100μmol/L)on Ras protein expression,cell proliferation and apoptosis of chronic myeloid leukemia K562 cells were detected respectively by Western blotting,CCK-8 assay,and flow cytometry.Results:A total of 14 common targets of Res against myeloid leukemia were obtained,and 122 pathways were obtained by enrichment analysis(P<0.05),among which the main signaling pathways with more enriched targets were Ras signaling pathway,Neurotrophin signaling pathway,etc.and 22 molecular functions,13 cell compositions,and 89 biological processes were obtained.Ras protein and Res formed the hydrogen bonds with the lowest energy among THR-74,GLN-150 and GLU-143.Res was able to down-regulate Ras protein expression,inhibit cell proliferation and promote apoptosis in K562 cells,and the effect of 100μmol/L Res was more significant(all P<0.05).Conclusion:Res may play a positive role in the treatment of myeloid leukemia by acting on Ras targets and by mediating the Ras signaling pathway,meanwhile,it possibly affects the proliferation and apoptosis of leukemic cells.