The final study consisted of a mix of methadone maintained former heroin addicts, healthy controls, opioid-treated chronic pain patients, and non-opioid treated chronic pain patients. DNA was extracted from whole blood and both global methylation at LINE-1 repeats and local methylation patterns at 22 CpG loci in the OPRM1 promoter region were analyzed with respect to the above groups. Increased methylation at CpG +126 and global LINE-1 were seen in the methadone patients as compared with controls, and additional analyses including smoking as a covariate confirmed hypermethylation at LINE-1 but not CpG +126. Similarly, in the opioid-treated pain patients versus non-opioid-treated pain patients, global methylation at LINE-1 was significantly higher, as was local methylation at CpG +126. LINE-1 methylation but not OPRM1 methylation at any CpG was also higher in the methadone patients as compared to opioid-prescribed pain patients. Interestingly, LINE-1 methylation but not OPRM1 methylation in the chronic pain patients was significantly correlated with pain intensity, but not in the methadone patients.
