摘要
目的观察人结肠癌Caco-2细胞系P16基因启动子区甲基化状态,并探讨去甲基化制剂5-氮杂-2-脱氧胞苷(5-Aza-CdR)诱导高甲基化失活的P16基因重新表达的可能性及其对细胞生长的影响。方法用不同浓度的5-Aza-CdR处理Caco-2细胞系,MSP法检测用药前后P16基因的甲基化状态,RT-PCR方法检测P16基因mRNA表达。MTT法观察细胞生长速度,流式细胞仪检测细胞周期、细胞凋亡率。结果 P16基因在人结肠癌细胞系Caco-2中启动子区呈甲基化状态,经过5-Aza-CdR处理后,P16基因启动子区呈去甲基化状态,其mRNA重新表达。CpG岛去甲基化后能明显地抑制细胞的生长,诱导细胞凋亡,影响细胞周期分布,并具有良好的量效依赖关系。结论 5-Aza-CdR能够逆转P16基因甲基化状态,调控P16基因表达并有效地抑制肠癌细胞增殖。
Objective To investigate P16 gene methylation state in human colorectal cancer cell line Caco-2 and explore the possibility of re-expression of the hypermethylated and silenced P16 gene. Methods Cell line Caco-2 was treated with different concent rations of DNA methyltransferase inhibitor 5-Aza-CdR, MSP was used to detect promoter methylation state of P16 gene and RT-PCR were used to detect re-expression of mRNA before and after treatment with 5-Aza-CdR respectively. Cell growth speed was measured using MTT assay, cell cycle distribution and apoptosis rate were estimated using flow cytometry. Results Promoter hypermethylation of the P16 gene was detected in human colorectal cancer cell line Caco-2, After treatment with 5-Aza-CdR, the promoter region of the P16 gene exhibited a demethylation state, and 5-Aza-CdR induces P16 gene re-expression. 5-Aza-CdR effectively reversed the hypermethylation status of CpG island, induced colorectal cancer cell apoptosis and cell cycle arrest were observed in a dose-dependent manner. Conclusion P16 gene can be reversed by demethylation agent 5-Aza-CdR. demethylation agent can regulate the expression of the P16 gene, and effectively inhibits cell growth
出处
《基础医学与临床》
CSCD
北大核心
2011年第2期161-165,共5页
Basic and Clinical Medicine
基金
江西省教育厅青年科学基金(GJJ09453)
