摘要
目的观察去甲基化剂5-氮杂-2'-脱氧胞苷(5-Aza-CdR)对人低分化鼻咽癌细胞系CNE-2Z RASSF1A基因的去甲基化作用。方法用5-Aza-CdR处理CNE-2Z细胞后,应用甲基化特异性PCR(MSP)检测RASSF1A基因的甲基化情况,免疫细胞化学检测RASSF1A基因表达改变,细胞生长曲线和平板克隆形成试验分别检测细胞生长能力和克隆形成能力。结果 CNE-2Z细胞RASSF1A基因呈高甲基化和阴性表达。经5-Aza-CdR处理,第1～2天RASSF1A基因高甲基化状态不改变,第3天部分去甲基化,第4天后全去甲基化,5-Aza-CdR处理停止2天后仍保持全去甲基化。随着RASSF1A基因去甲基化,RASSF1A呈明显阳性表达、细胞生长和平板克隆形成能力明显下降。结论 RASSF1A基因在CNE-2Z细胞中因高甲基化而失表达。5-Aza-CdR可诱导CNE-2Z细胞RASSF1A基因完全去甲基化而增强其表达,明显抑制CNET-2Z细胞生长和克隆形成能力。
Objective To observe the effect of the demethylation agent 5-Aza-CdR on the demethylation of Rassf1a gene in poorly differentiated nasopharyngeal carcinoma cell line（CNE-2Z）.Methods The experimental cells were treated with 5-Aza-CdR,and the control cells were untreated.The methylation specific PCR（MSP） was used to test the methylation status of the cells.The immunocytochemistry was used to observe the expression of Rassf1a gene.The growth curve and the plate clone efficiency test were used to test the ability of the growth and clone formation of the cells.Results The Rassf1a gene in CNE-2Z cells was in high degree of methylation and negative expression.Treated with 5-Aza-CdR,Rasssf1a gene in the cells remained high methylation and negative expression on the first two days,half demethylation on the third day,and full demethylation on the fourth day.Two days after the 5-Aza-CdR was removed,the gene still remained full demethylation.As the methylation was deprived,the gene expression was recovered,the ability of growth and clone formation of the cells obviously decreased.Conclusion Rassf1a gene loses expression in CNE-2Z cells because of high methylation.5-Aza-CdR might fully induce demethylation of rassf1a gene and improve its expression in CNE-2Z cells,and obviously inhibit the ability of growth and clone formation of the cells
出处
《实用癌症杂志》
2012年第5期441-444,448,共5页
The Practical Journal of Cancer
基金
东莞市科技计划项目(201010815215)
