Effects of homocycteine on methylation status and mRNA expression of TNFSF4 gene in macrophages in vitro / 第三军医大学学报

ABSTRACT

Objective To investigate the effects of homocysteine (Hcy) on methylation status and mRNA expression of TNF superfamily member 4 (TNFSF4) gene in THP-1 macrophages. Methods Cultured THP-1 monocytes were induced to macrophages by 0.1 ?mol/L PMA treatment for 72 h, then the differentiated THP-1 macrophages were incubated with homocysteine (50-1 000 ?mol/L) for 24, 48 or 72 h. The status of methylation of TNFSF4 gene in THP-1 macrophages was analyzed by methylation specific polymerase chain reaction (MS-PCR).The expression level of TNFSF4 mRNA was determined by RT-PCR. Results The MS-PCR results showed that the unmethylated bands gradually became stronger, and the methylated bands gradually became weaker along with the prolongation of treatment time and the increased Hcy concentrations. The 72-hour treatment with Hcy induced a complete demethylation in the promotor region of TNFSF4 gene, where left the only unmethylated bands. Meanwhile, the TNFSF4 mRNA expression was also increased in a dose-dependent manner. Conclusion Hcy may contribute to atherogenesis by inducing demethylation in the promotion region of the TNFSF4 gene and increasing TNFSF4 expression in THP-1 macrophages.