miR-584-5p通过下调MMP-14抑制乳腺癌细胞生物学行为  

miR-584-5p inhibited the biological behavior of breast cancer cells by down-regulating MMP-14

作　　者：刘陈美琪 李海林 LIU Chenmeiqi;LI Hailin(Clinical Medical College of Jiamusi University,Jiamusi,Heilongjiang 154007,P.R.China;Department of Second General Surgery,Hongqi Hospital Affiliated to Mudanjiang Medical College,Mudanjiang,Heilongjiang 157011,P.R.China)

机构地区：[1]佳木斯大学临床医学院,黑龙江佳木斯154007 [2]牡丹江医学院附属红旗医院普外二科,黑龙江牡丹江157011

出　　处：《中国普外基础与临床杂志》2023年第3期302-308,共7页Chinese Journal of Bases and Clinics In General Surgery

基　　金：黑龙江省卫生健康委科研课题(项目编号:2020-171)。

摘　　要：目的探讨微小RNA-584-5p(miR-584-5p)对乳腺癌细胞生物学行为(增殖、迁移和侵袭)的影响及其作用机制。方法选择人正常乳腺上皮细胞MCF10A及乳腺癌细胞MDA-MB-231、SK-BR-3和MCF-7;取对数生长期的MCF-7细胞,应用LipofectamineTM 2000转染试剂盒对其进行转染,并分为7组:空白组(未转染的MCF-7细胞)、模拟物-阴性对照组(mimic-negative control,mimic-NC,mimic-NC组;转染mimic-NC)、miR-584-5p mimic组(转染miR-584-5p mimic)、pcDNA组[转染过表达基质金属蛋白酶-14(MMP-14)的pcDNA3.1质粒阴性对照(pcDNA3.1)]、MMP-14组[转染过表达MMP-14的pcDNA3.1质粒(pcDNA3.1-MMP-14)]、mimic-NC+MMP-14组(共转染mimic-NC和pcDNA3.1-MMP-14)及miR-584-5p mimic+MMP-14组(共转染miR-584-5p mimic和pcDNA3.1-MMP-14)。荧光定量PCR法检测MCF10A、MDA-MB-231、SK-BR-3和MCF-7细胞中的miR-584-5p表达水平以及各组MCF-7细胞中miR-584-5p及MMP-14 mRNA表达水平;蛋白印迹法检测各组MCF-7细胞中MMP-14蛋白表达;采用细胞计数试剂盒-8(CCK-8)、划痕实验和Transwell实验分别检测各组MCF-7细胞的增殖、迁移及侵袭情况;采用双荧光素酶报告基因实验检测miR-584-5p与MMP-14的靶向关系。结果与人正常乳腺上皮细胞MCF10A比较,乳腺癌MDA-MB-231、SK-BR-3和MCF-7细胞中miR-584-5p表达水平均降低(P<0.05),且MCF-7细胞中miR-584-5p表达水平最低。与空白组和mimic-NC组比较,miR-584-5p mimic组MCF-7细胞的miR-584-5p表达水平升高,MMP-14 mRNA和蛋白表达水平、增殖活力、划痕愈合率和侵袭细胞数降低或减少(P<0.05);与空白组和pcDNA组比较,MMP-14组MCF-7细胞的MMP-14 mRNA和蛋白表达水平、增殖活力、划痕愈合率及侵袭细胞数增高或增多(P<0.05);与MMP-14组及mimic-NC+MMP-14组比较,miR-584-5p mimic+MMP-14组MCF-7细胞的miR-584-5p表达水平升高,MMP-14 mRNA和蛋白表达水平、增殖活力、划痕愈合率和侵袭细胞数降低或减少(P<0.05);miR-584-5p mimic+MMP-14组MCF-7细胞的MMP-14 mRNA和�Objective To investigate the effect of microRNA-584-5p(miR-584-5p)on the biological behavior(proliferation,migration and invasion)of breast cancer cells and its mechanism.Methods Human normal breast epithelial cells MCF10A and breast cancer cells MDA-MB-231,SK-BR-3 and MCF-7 were selected;take MCF-7 cells in logarithmic growth phase,transfect them with LipofectamineTM 2000 transfection kit,and divide them into seven groups:blank group(untransfected MCF-7 cells),mimic-negative control(mimic-NC)group(transfected mimic-NC),miR-584-5p mimic group(transfected miR-584-5p mimic),pcDNA group[transfected with overexpression of matrix metalloproteinase-14(MMP-14)pcDNA3.1 plasmid negative control(pcDNA3.1)],MMP-14 group[transfected with overexpression of MMP-14 pcDNA3.1 plasmid(pcDNA3.1-MMP-14)],mimic-NC+MMP-14 group(co-transfected with mimic NC and pcDNA3.1-MMP-14),and miR-584-5p mimic+MMP-14 group(co-transfected with miR-584-5p mimic and pcDNA3.1-MMP-14).The mRNA expression levels of miR-584-5p in MCF10A,MDA-MB-231,SK-BR-3 and MCF-7 cells and the expression levels of miR-584-5p and MMP-14 mRNA of MCF-7 cell in each group were detected by fluorescence quantitative PCR.The protein expressions of MMP-14 of MCF-7 cell in each group were detected by Western blotting.The proliferation,migration and invasion of MCF-7 cell in each group were detected by cell counting kit-8(CCK-8),scratch test and Transwell test.The targeting relationship between miR-584-5p and MMP-14 was detected by double luciferase reporter gene assay.Results Compared with the human normal mammary epithelial cells MCF10A,the expression levels of miR-584-5p in breast cancer cells MDA-MB-231,SK-BR-3 and MCF-7 were decreased(P<0.05),and the expression level of miR-584-5p in MCF-7 cells was the lowest.Compared with the blank group and the mimic-NC group,the expression level of miR-584-5p of MCF-7 cells in the miR-584-5p mimic group was increased,and the expression levels of MMP-14 mRNA and protein,proliferation activity,scratch healing rate and invasive cell number

关 键 词：微小RNA-584-5p 基质金属蛋白酶-14 乳腺癌 生物学行为 

分 类 号：R737.9[医药卫生—肿瘤]