INTRODUCTION

Alterations in DNA methylation profiles have been associated with cancer, and can be influenced by environmental factors such as smoking. A small but growing literature indicates there are reproducible and robust differences in methylation levels among smokers, never smokers, and ex-smokers. Here, we compared differences in salivary DNA methylation levels among current and ex-smokers (at least 2 years abstinent).

METHODS

Smokers (n=26) and ex-smokers (n=30) provided detailed smoking histories, completed the Paced Auditory Serial Addition Test (PASAT), and submitted a saliva sample. Whole-genome DNA methylation from saliva was performed, and ANCOVA models and a receiver operating characteristic (ROC) curve were used for the differences between groups and the performance of significant CpG sites.

RESULTS

After controlling for race, age, and gender, smokers had significantly lower methylation levels than ex-smokers in two CpG sites: cg05575921 (AHRR) and cg21566642 (ALPPL2). Based on the ROC analyses, both CpGs had strong classification potentials (cg05575921 AUC=0.97 and cg21566642 AUC=0.93) in differentiating smoking status. Across all subjects, the percent methylation of cg05575921 (AHRR) and cg21566642 (ALPPL2) positively correlated with the length of the last quit attempt (r=0.65 and 0.64, respectively, p<0.001) and PASAT accuracy (r=0.29 and 0.30, respectively, p<0.05).

CONCLUSIONS

In spite of the small sample size and preliminary research, our results replicate previously reported differences in AHRR hypomethylation among smokers. Furthermore, we show that the duration of smoking abstinence is associated with a recovery of methylation in ex-smokers, which may be linked to a reduced risk of smoking-associated diseases. The association with cognitive performance suggests that the hypomethylation of AHRR in saliva may reflect systemic exposure to cigarette-related toxicants that negatively affect cognitive performance, and should be validated in larger studies.