Effects of 5-Aza-2-deoxycytidine on DNA Methylation and Expression of p16 and MGMT Gene in Lung Cancer Cell Line SPC-A-1
|School||Anhui Medical University,|
|Keywords||5-Aza-2’-deoxycytidine p16 gene MGMT gene DNA methylation lung cancer|
Objective To investigate the effects of 5-Aza-2 -deoxycytidine ( 5-Aza-dC) on DNA methylation and expression of p16 and MGMT gene in the human lung cancer cell line SPC-A-1 ,and to explore the mechanism of p16 and MGMT gene silencing in human lung cancer cells and the regulating effect of the demethylating agent on p16 and MGMT gene expression.Methods SPC-A-1 cells were cultured in RPMI-1640 medium and were treated with 5μmol/L of DNA methyltransferase inhibitor 5-Aza-dC. Methylation-specific polymerase chain reaction( MSP) was used to detect the promoter methylation state of the p16 and MGMT gene. RT-PCR was used to detect the mRNA expression of p16 and MGMT before and after treatment with 5-Aza-dC, respectively.Results Before treatment with 5-Aza-dC,p16 and MGMT expressions were absent,and promoter hypermethylation of the p16 and MGMT gene were detected in SPC-A-1 cells, After treatment with 5-Aza-dC, the promoter region of the p16 and MGMT gene exhibited a demethylation state, and their mRNA expressions were increased.Conclusion Promoter hypermethylation is a major mechanism of p16 and MGMT gene silencing in human lung cancer cells, and can be reversed by the demethylating agent 5-Aza-dC, which can regulate the expressions of the p16 and MGMT gene.