<b>Concentration-dependent effect of delta-9-tetrahydrocannabinol on epigenetic DNA modifiers in human peripheral blood mononuclear cells</b> - DMR/DhMR datasets from RRBS and RRoxBS

Cannabis is among the most used illicit substances in the world, and it is estimated that around 10% of regular cannabis users are susceptible to developing cannabis use disorder (CUD). We examined the effect of different concentrations of delta-9-tetrahydrocannabinol (THC) on epigenetic DNA modifiers - DNA methyltransferases (DNMTs) and ten-eleven translocation enzymes (TETs), cannabinoid <i>CB1</i> and <i>CB2</i> receptors, and cytokines <i>IL-1β, IL-6</i>, <i>IL-10</i>, and <i>TNF-α</i>. We used two <i>in vitro</i> 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 <i>DNMTs</i> and <i>TETs</i><i> </i>mRNA levels, enzymatic activity, or <i>CB1</i> and <i>CB2</i> mRNA levels at an average THC concentration (50 ng/ml, n=8 donors). However, repeated exposure to a high THC concentration (200 ng/ml, n=16 donors) statistically significantly downregulated <i>DNMTs</i> and upregulated <i>TETs</i>, <i>CB1</i> and <i>CB2</i> mRNA levels. Both concentrations of THC upregulated gene expression of <i>IL-1β, IL-6</i>, and <i>IL-10</i> and did not affect<i> </i><i>TNF-α</i> gene expression. On a genome-wide level, we found that after repeated THC incubations, a significant amount of differentially hydroxymethylated genes were hyperhydroxymethylated. An additional THC challenge shifted the hyperhydroxymethylated state to hypohydroxymethylation. Genes showing the strongest associations with THC exposure had functional significance in various signalling pathways. Our results indicate that repeated THC exposure induces a dose-dependent maladaptive response that may affect the expression of genes essential in the development of CUD.