Association of promoter methylation statuses of congenital heart defect candidate genes with Tetralogy of Fallot

Background: Although a lower methylation level of whole genome has been demonstrated in Tetralogy of Fallot (TOF) patients, little is known regarding changes in specific gene DNA methylation profiles and the possible associations with TOF. In current study, the promoter methylation statuses of congenital heart defect (CHD) candidate genes were measured in order to further understand epigenetic mechanisms that may play a role in the development of TOF. Methods: The methylation levels of CHD candidate genes were measured using the Sequenom MassARRAY platform. QRT-PCR was used to analyze the mRNA levels of CHD candidate genes in the right ventricular myocardium of TOF cases and normal controls. Results: Methylation status analysis was performed on the promoter regions of 71 CHD candidate genes (113 amplicons). We found significant differences in methylation status, between TOF cases and controls, in 26 amplicons (26 genes) (p < 0.05). Of the 26 amplicons, 17 were up regulated and 9 were down regulated. Additionally, 14 of them were located in the CpG islands, 7 were located in the CpG island shores, and 5 were covering the regions near the transcription start site (TSS). The methylation status was subsequently confirmed and mRNA levels were measured for 7 represented candidate genes, including EGFR, EVC2, NFATC2, NR2F2, TBX5, CFC1B and GJA5. The methylation values of EGFR, EVC2, TBX5 and CFC1B were significantly correlated with their mRNA levels (p < 0.05). Conclusions: Aberrant promoter methylation statuses of CHD candidate genes presented in TOF cases may contribute to the TOF development and have potential prognostic and therapeutic significance for TOF disease.