LncRNA MCM3AP-AS1调节miR-424-5p/PSAT1轴对乳腺癌恶性进展的影响  

Effect of LncRNA MCM3AP-AS1 on Malignant Progression of Breast Cancer by Regulating miR-424-5p/PSAT1 Axis

作　　者：冯佳梅[1] 万华[1] 高晴倩[1] 瞿文超[1] 邵士珺[1] 孙佳晔 吴雪卿[1] FENG Jia-mei;WAN Hua;GAO Qing-qian;QU Wen-chao;SHAO Shi-jun;SUN Jia-ye;WU Xue-qing(artment of Breast,Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine,Shanghai,200011,)

机构地区：[1]上海中医药大学附属曙光医院乳腺科,上海200011

出　　处：《现代生物医学进展》2024年第14期2606-2612,2625,共8页Progress in Modern Biomedicine

基　　金：上海市卫生健康委员会科研课题计划项目(202040254)。

摘　　要：目的:探讨长链非编码核糖核酸(lncRNA)微小染色体维持蛋白3相关蛋白-反义链1(MCM3AP-AS1)调节微小核糖核酸(miR)-424-5p/磷酸丝氨酸氨基转移酶1(PSAT1)轴对乳腺癌恶性进展的影响。方法:采用实时荧光定量聚合酶链式反应(RT-qPCR)法检测人正常乳腺上皮细胞MCF-10A和乳腺癌细胞株MDA-MB-231、BT549、BT20中LncRNA MCM3AP-AS1、miR-424-5p和PSAT1信使核糖核酸(m RNA)的表达水平。构建LncRNA MCM3AP-AS1低表达模型、miR-424-5p敲低与过表达模型,并使用RT-qPCR方法验证转染模型构建的成功。分别采用四氮甲基唑蓝(MTT)法、Transwell实验检测乳腺癌细胞株的增殖、迁移、侵袭能力。免疫印迹法检测各组MDA-MB-231细胞中细胞周期蛋白D1(CyclinD1)、E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)和PSAT1蛋白的表达。经生物信息学分析后,采用双荧光素酶报告基因实验与RNA免疫共沉淀(RIP)实验分别验证LncRNA MCM3AP-AS1与miR-424-5p、miR-424-5p与PSAT1之间的靶向关系。结果:在乳腺癌细胞株MDA-MB-231、BT549、BT20中MCM3AP-AS1、PSAT1 m RNA的表达水平显著高于MCF-10A(P<0.05),miR-424-5p表达水平显著低于MCF-10A(P<0.05)。敲低MCM3AP-AS1或过表达miR-424-5p均可以降低MDA-MB-231细胞的吸光度(OD490)值、细胞迁移数目和细胞侵袭数目、CyclinD1、N-cadherin和PSAT1蛋白表达水平(P<0.05),提高细胞E-cadherin蛋白表达水平(P<0.05)。敲低miR-424-5p的表达逆转了下调MCM3AP-AS1对MDA-MB-231细胞增殖、迁移和侵袭以及相关蛋白表达的影响。双荧光素酶报告基因实验与RIP实验证实MCM3AP-AS1靶向负调控miR-424-5p表达,miR-424-5p靶向负调控PSAT1的表达。结论:LncRNA MCM3AP-AS1在乳腺癌中呈高表达,其低表达可通过靶向调节miR-424-5p/PSAT1轴抑制乳腺癌的恶性进展。Objective:To investigate the effect of long non-coding RNA(lncRNA)microchromosome maintenance protein 3-associated protein-antisense chain 1(MCM3AP-AS1)on malignant progression of breast cancer by regulating microribonucleic acid(miR)-424-5p/phosphoserine aminotransferase 1(PSAT1)axis.Methods:The expression levels of LncRNA MCM3AP-AS1,miR-424-5p and PSAT1 m RNA in human normal breast epithelial cells MCF-10A and breast cancer cell lines MDA-MB-231,BT549 and BT20 were detected by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR).LncRNA MCM3AP-AS1 low expression model,miR-424-5p knockdown and overexpression model were constructed,and the success of transfection model construction was verified by RT-q PCR method.The proliferation,migration and invasion of breast cancer cell lines were detected by methyl thiazolyl tetrazolium(MTT)assay and Transwell assay respectively.The expressions of cyclin d1(CyclinD1),E-cadherin(E-cadherin),N-cadherin(N-cadherin)and PSAT1 proteins in MDA-MB-231 cells were detected by Western blotting.After bioinformatics analysis,the targeting relationship between LncRNA MCM3 AP-AS1 and miR-424-5p,miR-424-5p and PSAT1 was verified by double luciferase reporter gene assay and RNA immunoprecipitation(RIP)assay respectively.Results:The expression levels of MCM3AP-AS1 and PSAT1 m RNA in breast cancer cell lines MDA-MB-231,BT549 and BT20 were significantly higher than those in MCF-10A(P<0.05),and the expression level of miR-424-5p was significantly lower than that in MCF-10A(P<0.05).Knockdown of MCM3AP-AS1 or overexpression of miR-424-5p could reduce the absorbance(OD490)value of MDA-MB-231 cells,the number of cell migration and cell invasion,the expression levels of CyclinD1,N-cadherin and PSAT1 proteins(P<0.05),and increase the expression level of E-cadherin protein(P<0.05).Knockdown of miR-424-5p reversed the effects of down-regulation of MCM3AP-AS1 on the proliferation,migration and invasion of MDA-MB-231 cells and the expression of related proteins.The dual luciferase report

关 键 词：MCM3AP-AS1 miR-424-5p PSAT1 乳腺癌 

分 类 号：R-33[医药卫生] R737.9