BACKGROUND:
Azoxymethane (AOM)-induced colorectal cancer (CRC) model has been extensively applied to the study of molecular mechanisms and the prevention/treatment of CRC. To date, genetic alterations in AOM model, such as K-ras mutation, are well documented, but the underlying epigenetic mechanisms are largely unknown. Aberrant activation of the WNT signalling pathway is critical for the initiation and progress of human CRC, which provokes the nuclear β-catenin translocation, the induction of WNT-downstream target genes and dysregulation of cell proliferation and differentiation. WNT-antagonist genes, SFRP1, 2, 5 are important in the regulation of the WNT pathway and are frequently silenced in human CRC. In this study, we investigated the role of SFRP5 methylation in the AOM model.
METHODS:
Rat colon tumors were obtained from AOM model (induced by two weekly AOM treatments and collected 30 weeks), and normal colon tissues were obtained from rats without AOM treatments. Promoter methylation of SFRP5 was analysed by bisulfite genomic sequencing.
Quantitative real-time RT-PCR was used to measure SFRP5, C-Myc and Cylin D1 mRNA; immunohistochemical staining was used to measure SFRP5, C-Myc and Cylin D1, β-catenin, DNMT1 expression and histone H3 acetylation.
RESULTS:
AOM-induced colon tumors displayed significant increased SFRP5 promoter hypermethylation. SFRP5 methylation was methylated in 80% of colon tumors (n=10), compared to 40% of normal colon (n=10). The methylation was associated with increased expression of DNMT1, β-catenin, C-Myc and Cylin D1 and decreased expression of SFRP5 and histone H3 acetylation.
CONCLUSIONS:
SFRP5 acts as a tumor suppressor gene in the AOM model, there is a link between increased DNMT1 expression and histone deacetylation, induction of promoter hypermethylation of SFRP5 gene and aberrant activation of Wnt/β-catenin signalling pathway. Further studies are warranted to determine the possible involvement of methylation of other WNT-antagonist genes and the potential effects of diet on restoration of WNT-antagonist gene expression.