5-氮杂-2′-脱氧胞苷逆转RASSF1A基因甲基化对人乳腺癌MCF-7细胞的影响  

Effect of the promoter methylation and the intervention by 5-aza-2'-deoxycytidine on RASSF1A gene of breast cancer cell line MCF-7

作　　者：周小娟[1] 王云梅[1] 刘娟[1] 李新红[1] 

机构地区：[1]西安交通大学医学院第一附属医院肿瘤内科,陕西西安710061

出　　处：《第四军医大学学报》2009年第21期2327-2331,共5页Journal of the Fourth Military Medical University

摘　　要：目的:探讨启动子甲基化对乳腺癌MCF-7细胞RASSF1A基因的作用及5-氮杂-2′-脱氧胞苷(5-Aza-CdR)干预对启动子甲基化的影响,为临床治疗乳腺癌奠定理论基础.方法:用1,3,5,7,9μmol/L不同浓度的5-Aza-CdR处理MCF-7乳腺癌细胞株,通过MTT检测5-Aza-CdR对MCF-7乳腺癌细胞株存活率的影响;建立5μmol/L,10μmol/L的浓度梯度流式细胞仪检测5-Aza-CdR对MCF-7乳腺癌细胞株生长周期及凋亡率的影响,MSP(甲基化PCR)检测5-Aza-CdR处理前后RASFF1A抑癌基因的甲基化状态,RT-PCR检测处理前后抑癌基因RASSF1A mRNA表达情况.结果:5-Aza-CdR抑制体外培养的人乳腺癌MCF-7细胞,诱导人乳腺癌MCF-7细胞凋亡和细胞周期停滞在G0/G1期,呈浓度依赖性.经5-Aza-CdR处理人乳腺癌MCF-7细胞后,MSP检测RASSF1A基因发生了甲基化,RT-PCR检测RASSF1A mRNA恢复了表达.结论:RASSF1A基因启动子甲基化是导致其失活的主要原因,5-Aza-CdR可以逆转乳腺癌细胞株MCF-7细胞RASSF1A基因启动子甲基化状态,使其重新表达.AIM：To approach the effect of promoter methylation to RASSF1A gene of breast cancer cell line MCF-7 and 5-Aza-CdR s impact on promoter methylation above. METHODS：Breast cancer cell lines MCF-7 were treated with 1,3,5,7 and 9 μmol/L 5-Aza-CdR respectively and MTT assay was used to detect the survival rate. Then cells were treated with 5 and 10 μmol/L 5-Aza-CdR,the cell cycle and apoptosis rate were analyzed by flow cytometry and the methylation was examined by PCR using methylation specific primer （MSP）,and the expression levels ofmRNA of RASSF1A were detected by reverse transcription-polymerase chain reaction （ RT-PCR ）. RESULTS ： After 5-Aza-CdR treatment, the proliferation of breast caner cell line MCF-7 is inhibited,the number of G0/G1 cell was increased, and 5-Aza- CdR blocked the cell cycle at G1/GO phase,the apoptotie rate was also increased significantly in a concentration dependent manner. After 5-Aza-CdR treatment , methylation specific primer （MSP） detected RASSF1A gene in breast cancer cell line generated meth- ylation, RT-PCR detected mRNA of RASSF1A gene in breast cancer cell line recovered express. CONCLUSION：The promot- er methylation of RASSF1A gene in breast cancer cell line MCF-7 is the main reason inducing itself deactivation, while 5-Aza-CdR can reverse that promoter methylation to make RASSF1A gene express again.

关 键 词：MCF-7细胞 RASSF1A基因 5-氮杂-2′-脱氧胞苷 

分 类 号：R737[医药卫生—肿瘤]