Ageing, chronic alcohol consumption and folate are determinants of genomic DNA methylation, p16 promoter methylation and the expression of p16 in the mouse colon

Older age, dietary folate and chronic alcohol consumption are important risk factors for the development of colon cancer. The present study examined the effects of ageing, folate and alcohol on genomic and p16-specific {DNA} methylation, and p16 expression in the murine colon. Old (aged 18 months; n 70) and young (aged 4 months; n 70) male {C57BL/6} mice were pair-fed either a {Lieber-DeCarli} liquid diet with alcohol (18 % of energy), a {Lieber-DeCarli} diet with alcohol (18 %) and reduced folate (0.25 mg folate/l) or an isoenergetic control diet (0.5 mg folate/l) for 5 or 10 weeks. Genomic {DNA} methylation, p16 promoter methylation and p16 gene expression were analysed by liquid {chromatography-MS,} methylation-specific {PCR} and real-time {RT-PCR,} respectively. Genomic {DNA} methylation was lower in the colon of old mice compared with young mice {(P} {\textless} 0.02) at 10 weeks. Alcohol consumption did not alter genomic {DNA} methylation in the old mouse colon, whereas it tended to decrease genomic {DNA} methylation in young mice {(P} = 0.08). p16 Promoter methylation and expression were higher in the old mouse colon compared with the corresponding young groups. There was a positive correlation between p16 promoter methylation and p16 expression in the old mouse colon {(P} {\textless} 0.02). In young mice the combination of alcohol and reduced dietary folate led to significantly decreased p16 expression compared with the control group {(P} {\textless} 0.02). In conclusion, ageing and chronic alcohol consumption alter genomic {DNA} methylation, p16 promoter methylation and p16 gene expression in the mouse colon, and dietary folate availability can further modify the relationship with alcohol in the young mouse.