基于网络药理学和体外实验探讨荷叶碱治疗弥漫大B细胞淋巴瘤的作用机制  

Exploring the mechanism of Nuciferine in the treatment of diffuse large B-cell lymphoma based on network pharmacology and in-vitro experiments

作　　者：周晓丽 郭倩茜 胡顺凤 ZHOU Xiaoli;GUO Qianqian;HU Shunfeng(Department of Hematology,Shandong Provincial Hospital Affiliated to Shandong First Medical University,Jinan 250021,China;Qilu Medical College of Shandong University,Jinan 250012,China)

机构地区：[1]山东第一医科大学附属省立医院血液科,山东济南250021 [2]山东大学齐鲁医学院,山东济南250012

出　　处：《山东第一医科大学(山东省医学科学院)学报》2025年第1期24-32,共9页Journal of Shandong First Medical University & Shandong Academy of Medical Sciences

基　　金：国家自然科学基金(82400231);山东省泰山学者工程;中国博士后科学基金(2023M741506);山东省自然科学基金(ZR2023QH193)。

摘　　要：目的探讨荷叶碱治疗弥漫大B细胞淋巴瘤(diffuse large B-cell lymphoma,DLBCL)的核心靶点及作用机制。方法基于GeneCards数据库筛选DLBCL相关的靶基因,通过中药系统药理学数据库与分析平台(traditional Chinese medicine systems pharmacology,TCMSP),SwissTargetPrediction和PharmMapper网站检索荷叶碱相关的靶基因,取交集后进行富集分析。基于STRING平台进行蛋白质互作分析,利用Cytoscape筛选核心靶点并开展分子对接。细胞增殖实验检测荷叶碱处理后DLBCL细胞的增殖水平,流式细胞学实验检测加药后DLBCL细胞的周期阻滞水平和凋亡水平,Western blot检测其相关蛋白的表达水平。结果筛选出1422个与DLBCL相关的靶基因和247个与荷叶碱相关的靶基因,取交集后共得到50个潜在基因。基因本体论(gene ontology,GO)富集分析、京都基因与基因百科全书(Kyoto encyclopedia of genes and genomes,KEGG)通路分析显示,潜在基因主要富集于细胞增殖、细胞周期、细胞凋亡等信号通路。随后构建“药物-靶点-通路-疾病”的网络关系图。分子对接提示荷叶碱与核心靶点具有良好的结合活性,通过qRT-PCR进一步验证荷叶碱的表达调控作用。CCK-8实验表明,荷叶碱可以抑制DLBCL细胞的增殖,且抑制作用呈浓度依赖性和时间依赖性。流式细胞学实验发现,荷叶碱以浓度依赖性的方式诱导DLBCL细胞在G2/M期阻滞,细胞凋亡率增加,且具有浓度依赖性。Western blot结果表明,荷叶碱可降低DLBCL细胞中c-MYC、Cyclin B1和CDK1的表达水平,升高cleaved-PARP和cleaved-caspase-3的表达水平。结论荷叶碱可以通过调控细胞增殖、细胞凋亡、细胞周期等生物学过程,进而抑制DLBCL的发生、发展,为进一步探究荷叶碱治疗DLBCL的分子机制和临床应用提供了科学依据。Objective:To explore key targets and related mechanisms of Nuciferine in the treatment of diffuse large B-cell lymphoma(DLBCL),network pharmacology,molecular docking techniques and in-vitro experiments are performed.Methods:Related target genes of DLBCL were screened based on GeneCards database.traditional Chinese medicine systems pharmacology(TCMSP)database,SwissTargetPrediction and PharmMapper online websites were used to search related targets of Nuciferine.Enrichment analysis was conducted following the intersection.The STRING platform was used for protein interaction analysis,while Cytoscape was used to screen key targets,which were subsequently used for molecular docking.The proliferation level of DLBCL cells after Nuciferine treatment was detected by cell proliferation assay.The cycle arrest and apoptosis level of DLBCL cells after Nuciferine treatment was detected by flow cytometric analysis.The expression level of related proteins was validated by Western blot(WB)after Nuciferine treatment.Results:Through GeneCards,TCMSP database,SwissTargetPrediction and PharmMapper online websites,1422 related target genes of DLBCL and 247 related targets of Nuciferine have been screened.Combined with the above results,50 potential genes that might play key roles have been disclosed.Enrichment analysis has revealed that key targets are closely related to biological processes involved in tumor progression,such as cell proliferation,cell cycle and apoptosis;the“drug-target-pathway-disease”network is established.Molecular docking has shown that Nuciferine has good binding activity with key targets,and qRT-PCR assay is performed to verify the regularory role of Nuciferine.Time-and dose-dependently inhibitory effect of Nuciferine treatment on the proliferation was found in DLBCL cells by cell proliferation assay.Nuciferine treatment induced the elevation of G2/M phase cells in a dose-dependent manner and apoptosis of DLBCL cells in flow cytometric experiments.Furthermore,after Nuciferine treatment,the expression levels

关 键 词：弥漫大B细胞淋巴瘤 荷叶碱 网络药理学 分子对接 

分 类 号：R285.5[医药卫生—中药学]