Associate Professor Oregon Health and Science University
Introduction: Cannabis is the most commonly used drug by reproductive age males. Our objective was to determine the effect of chronic delta-9-tetrahydrocannabinol (THC) use on the sperm epigenome and whether these changes are permanent or temporary with discontinuation of THC in a rhesus macaque (RM) model. Methods: We performed whole genome bisulfite sequencing (WGBS) of DNA extracted from sperm collected from 6 adult RM at baseline (pre-THC), at a high-THC dose (2.5mg/7kg/day), and post-THC discontinuation for 140 days (~2 spermatogenesis cycles). Bioinformatic analysis included QA/QC assessments, alignment to the rhesus reference genome, and identification of differentially methylated regions (DMRs). Results: Overlap of genes annotated to DMRs in our RM following THC with published gene lists of human cannabis users and non-users was more than expected by chance (p= 2.00e-82; Fig 1A). Pre-THC vs post-THC DMRs were similarly enriched for genes annotated to methylation differences between pre-cannabis exposure vs. post-cannabis discontinuation in humans (Fig 1B). Pre-THC vs high-THC DMRs annotated to 3,902 unique genes annotated and were significantly enriched for genes previously associated with autism spectrum disorder (ASD) (Fig 1C). We identified 2,759 DMRs between pre-THC and high-THC that intersected DMRs in high-THC vs post-THC sperm samples and demonstrated a reversal in methylation post-THC toward the level of methylation pre-THC (Fig 1D). A smaller number of DMRs between pre-THC and high-THC (n=1,898) did not return to baseline levels after THC abstinence (Fig 1E). Using Ingenuity Pathway Analysis, the top canonical pathway associated with THC and restored to baseline levels was the “synaptogenesis signaling pathway” and top biofunctions included “nervous system development and function” and “tissue development”. Similarly, the 1,898 non-restored DMRs were also enriched for genes in the “synaptogenesis signaling pathway” and “nervous system development and function”, as well as for genes in the “sperm motility” canonical pathway. Conclusions: Chronic THC use in RM alters genes related to nervous system development and function, and autism spectrum disorder that may impact longterm offspring outcomes. Discontinuation of THC resulted in restored methylation in only a subset of DMRs. SOURCE OF Funding: NICHD, NIDA, Silver Family
