Concentration-dependent effect of delta-9-tetrahydrocannabinol on epigenetic DNA modifiers in human peripheral blood mononuclear cells.

Cannabis is among the most used illicit substances in the world, and approximately 10% of regular cannabis users are estimated to be susceptible to developing cannabis use disorder (CUD). We examined the effect of different concentrations of delta-9-tetrahydrocannabinol (THC) on the epigenetic DNA modifiers DNA methyltransferases (DNMTs) and ten-eleven translocation enzymes (TETs); cannabinoid CB1 and CB2 receptors; and the cytokines IL-1β, IL-6, IL-10, and TNF-α. We used two in vitro study designs on human peripheral blood mononuclear cells (PBMCs) collected from healthy donors: (a) repeated THC incubations and (b) repeated THC incubations followed by an "abstinence" period and a THC challenge incubation. We observed no significant effects on DNMTs and TETs mRNA levels, enzymatic activity, or CB1 and CB2 mRNA levels at an average THC concentration (50 ng/ml, n = 8 donors). However, repeated incubations at a high THC concentration (200 ng/ml, n = 16 donors) significantly downregulated DNMTs and upregulated TETs, CB1, and CB2 mRNA levels. Both THC concentrations upregulated the gene expression of IL-1β, IL-6, and IL-10, but had no effect on TNF-α gene expression. At the genome-wide level, repeated THC incubations resulted in a significant number of differentially hydroxymethylated genes being hyperhydroxymethylated. An additional THC challenge shifted the hyperhydroxymethylated state to hypohydroxymethylation. The genes with the strongest associations with THC exposure were found to be functionally significant for various signaling pathways. These findings suggest that repeated incubations with high concentrations of THC may affect the expression of genes critical for the development of CUD through aberrant demethylation.