启动子区5′CpG岛去甲基化对人结肠癌细胞生物学表型的影响  

Effects of promoter region 5′CpG island demethylation on biological phenotype in human colorectal cancer cells

作　　者：方晓明[1] 郑树[1] 陈功星[1] 孙立峰[1] 吕庆华[1] 

机构地区：[1]浙江大学医学院附属第二医院

出　　处：《中国病理生理杂志》2005年第6期1111-1116,共6页Chinese Journal of Pathophysiology

基　　金：国家(973)重点基础研究发展归划项目(No.G1998051200);浙江省科技计划项目(No.011110541)

摘　　要：目的:探讨DNA启动子区5′CpG岛甲基化状态与人结肠癌RKO细胞增殖凋亡等生物学特征的关系。方法:应用特异性DNA甲基转移酶(DNMTs)抑制剂-5-氮-2′-脱氧胞苷(5-Aza-2′-deoxycytidine,5-Aza-CdR)处理肠癌RKO细胞72h,甲基化特异性PCR(methylation-specificPCR,MSP)及DNA测序法分析p16/CDKN2基因CpG岛甲基化状态;MTT、FCM、荧光染色及透射电镜检测启动子区去甲基化后细胞生长、形态和细胞周期凋亡的影响。结果:DNMTs抑制剂能较好地逆转启动子区胞嘧啶甲基化状态;CpG岛去甲基化后能明显地抑制肠癌细胞的生长,增加细胞群体倍增时间(P<0.01),诱导肠癌细胞凋亡,影响肠癌细胞周期分布,并具有良好的量效依赖关系。结论:通过逆转CpG岛高甲基化能有效地抑制肠癌细胞增殖,为临床治疗大肠癌提供新的作用靶点。AIM: To explore the relationship between methylation status of promoter region 5′CpG island and the biological phenotype in human colorectal cancer RKO cell lines. METHODS: RKO cells were treated with selective DNA methyltransferase (DNMTs) inhibitor, 5-Aza-2′-deoxycytidine (5-Aza-CdR), for 72 h. Methylation-specific PCR (MSP), T-A clone and DNA sequence analysis were used to detect 5′CpG island methylation status of p16/CDKN2 tumor suppresor gene. Cell growth, cell cycle arrest and apoptosis were analyzed by MTT, flow cytometry (FCM), fluorescent dye staining and transmission electron microscope. RESULTS: DNMTs inhibitor (5-Aza-CdR) effectively reversed the hypermethylation status of 5′CpG island. The effects of 5-Aza-CdR on cell growth inhibition (P<0.01), apoptosis and cell cycle arrest were observed in a dose-dependent manner. CONCLUSION: Selective DNMTs inhibitor inhibits cell growth by 5′CpG island demethylation, and this may be a potential new therapeutic target for colorectal cancer.

关 键 词：甲基化 结直肠肿瘤 细胞凋亡 细胞周期 脱氧胞苷 

分 类 号：R730.2[医药卫生—肿瘤] R730.5[医药卫生—临床医学]