miRNA-34a通过调控SOX4/RAS/MAPK信号通路对中枢神经系统淋巴瘤进展的影响  

Effect of miRNA-34a on the Progression of Central Nervous System Lymphomaby Regulating SOX4/RAS/MAPK Signaling Pathway

作　　者：周坚 潘晓冉 李小娟[1] ZHOU Jian;PAN Xiaoran;LI Xiaojuan(Hunan Provincial Brain Hospital/The Second People's Hospital of Hunan,Hunan Changsha 410000,China)

机构地区：[1]湖南省脑科医院/湖南省第二人民医院神经内科,湖南长沙410000

出　　处：《河北医学》2023年第2期189-194,共6页Hebei Medicine

基　　金：湖南创新型省份建设专项项目,(编号:2020JJ5293)。

摘　　要：目的:探讨微小RNA-34a(miR-34a)在原发性中枢神经系统淋巴瘤(PCNSL)中的作用及其潜在的分子机制。方法:通过实时荧光定量PCR(RT-qPCR)实验检测PCNSL组织中miR-34a的表达。体外培养Raji细胞,将miR-34a模拟物(miR-34a mimic)、模拟物对照(NC mimic)、miR-34a抑制剂(miR-34a inhibitor)、抑制剂对照(NC inhibitor)、miR-34a mimic+空白质粒(Vector)、miR-34a mimic+SOX4过表达质粒(pcDNA-SOX4)分别转染至细胞中。RT-qPCR实验检测细胞中miR-34a的表达;CCK-8实验和克隆形成实验检测细胞增殖能力;流式细胞术检测细胞凋亡率;Western blot检测细胞中SOX4蛋白和RAS/MAPK通路蛋白Ras、p-Raf-1、p-MEK的表达;生物信息学软件和双荧光素酶报告基因实验分别预测和验证miR-34a与SOX4的靶向关系。结果:PCNSL组织中miR-34a的表达显著降低(P<0.05)。与NC mimic组比较,miR-34a mimic组细胞中miR-34a的表达升高,细胞增殖能力显著降低,细胞凋亡率显著升高,Ras、p-Raf-1、p-MEK蛋白表达显著降低(P<0.05)。与NC inhibitor组比较,miR-34a inhibitor组细胞中miR-34a的表达降低,细胞增殖能力显著升高,细胞凋亡率显著降低,Ras、p-Raf-1、p-MEK蛋白表达显著升高(P<0.05)。与NC mimic组比较,共转染miR-34a mimic和WT-SOX4的细胞荧光素酶活性显著降低(P<0.05),而共转染miR-34a mimic和MUT-SOX4的细胞荧光素酶活性无显著性变化(P>0.05)。与miR-34a mimic+Vector组比较,miR-34a mimic+pcDNA-SOX4组Raji细胞增殖能力显著升高,细胞凋亡率显著降低,Ras、p-Raf-1、p-MEK蛋白表达显著升高(P<0.05)。结论:miR-34a通过靶向SOX4调控RAS/MAPK信号通路从而抑制PCNSL细胞增殖,促进细胞凋亡。Objective:To investigate the role of microRNA-34a(miR-34a)in primary central nervous system lymphoma(PCNSL)and its underlying molecular mechanisms.Methods:The expression of miR-34a in PCNSL tissues was detected by real-time quantitative PCR(RT-qPCR)experiments.Raji cell was cultured in vitro,and miR-34a mimic(miR-34a mimic),mimic control(NC mimic),miR-34a inhibitor(miR-34a inhibitor),inhibitor control(NC inhibitor),miR-34a mimic+Blank plasmid(Vector)and miR-34a mimic+SOX4 overexpression plasmid(pcDNA-SOX4)were transfected into cells respectively.The RT-qPCR assay was used to detect the expression of miR-34a in cells.Cell proliferation ability was detected by CCK-8 assay and clone formation assay.The apoptosis rate was detected by flow cytometry.Western blot was used to detect the expression of SOX4 protein and RAS/MAPK pathway proteins Ras,p-Raf-1,and p-MEK in cells.Bioinformatics software and dual luciferase reporter assay were used to predict and verify the targeting relationship between miR-34a and SOX4,respectively.Results:The expression of miR-34a in PCNSL tissues was significantly decreased(P<0.05).Compared with the NC mimic group,the expression of miR-34a in the miR-34a mimic group was increased,the cell proliferation ability was significantly decreased,the apoptosis rate was significantly increased,the protein expressions of Ras,p-Raf-1,and p-MEK were significantly decreased(P<0.05).Compared with the NC inhibitor group,the expression of miR-34a in the miR-34a inhibitor group was decreased,the cell proliferation ability was significantly increased,the apoptosis rate was significantly decreased,the protein expressions of Ras,p-Raf-1,and p-MEK were significantly increased(P<0.05).Compared with the NC mimic group,the luciferase activity of cells co-transfected with miR-34a mimic and WT-SOX4 was significantly decreased(P<0.05),while there was no significant change in luciferase activity of cells co-transfected with miR-34a mimic and MUT-SOX4(P>0.05).Compared with the miR-34a mimic+Vector group,the proliferatio

关 键 词：miRNA-34a 中枢神经系统淋巴瘤 SOX4 RAS/MAPK信号通路 增殖 

分 类 号：R733.1[医药卫生—肿瘤]