We characterised DNA methylation and gene expression of four tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors DR4, DR5, DcR1 and DcR2 in three choriocarcinoma (JAR, JEG-3, BeWo) and two transformed (HTR-8/SVneo and HPT-8) cell lines. DR4 mRNA was detected in JAR, JEG-3, BeWo and HTR-8/SVneo cells, whereas DR5 was present in all detected cells. DcR1 transcripts were expressed only in JAR, JEG-3 and BeWo cells, whereas DcR2 transcripts were detected only in HTR-8/SVneo and HPT-8 cells. Hypermethylated DR4 promoter was observed in JAR, JEG-3, BeWo and HTR-8/SVneo cells, hypermethylated DcR1 promoter in HTR-8/SVneo and HPT-8 cells and hypermethylated DcR2 promoter in JAR, JEG-3 and BeWo cells. Restoration of DR4, DcR1 and DcR2 expression with decreased DNA methylation of these genes was induced by the DNA demethylation agent 5-aza-2'-deoxycytidine (5-aza-CdR) in trophoblast cells, whereas DR5 expression did not exhibit any change. Significant negative correlation between the expression and DNA methylation of these genes was also observed. In all tested cell lines, only HPT-8 demonstrated sensitivity to TRAIL-induced apoptosis. Combined treatment with 5-aza-CdR and TRAIL resulted in apoptosis in JAR, JEG-3, BeWo and HTR-8/SVneo cells but not in HPT-8 cells. The results indicate that DNA methylation is associated with TRAIL receptor expression and might be involved in trophoblast apoptosis.