Cessation of chronic delta-9-tetrahydrocannabinol use partially reverses impacts on male fertility and the sperm epigenome in rhesus macaques.

OBJECTIVE: To determine whether discontinuation of delta-9-tetrahydrocannabinol (THC) use mitigates THC-associated changes in male reproductive health using a rhesus macaque model of daily THC edible consumption. DESIGN: Research animal study. SETTING: Research institute environment. PATIENT(S): Adult male rhesus macaques (age, 8-10 years; n = 6). INTERVENTION(S): Chronic daily THC edible administration at medically and recreationally relevant contemporary doses followed by cessation of THC use. MAIN OUTCOME MEASURE(S): Testicular volume, serum male hormones, semen parameters, sperm deoxyribonucleic acid (DNA) fragmentation, seminal fluid proteomics, and whole genome bisulfite sequencing of sperm DNA. RESULT(S): Chronic THC use resulted in significant testicular atrophy, increased gonadotropin levels, decreased serum sex steroid levels, changes in seminal fluid proteome, and increased DNA fragmentation with partial recovery after discontinuation of THC use. For every increase of 1 mg/7 kg/day in THC dosing, there was a significant decrease in the total testicular volume bilaterally by 12.6 cm3 (95% confidence interval [CI], 10.6-14.5), resulting in a 59% decrease in volume. With THC abstinence, the total testicular volume increased to 73% of its original volume. Similarly, with THC exposure, there were significant decreases in the mean total testosterone and estradiol levels and a significant increase in the follicle-stimulating hormone level. With increasing THC dose, there was a significant decrease in the liquid semen ejaculate volume and weight of coagulum; however, no other significant changes in the other semen parameters were noted. After discontinuing THC use, there was a significant increase in the total serum testosterone level by 1.3 ng/mL (95% CI, 0.1-2.4) and estradiol level by 2.9 pg/mL (95% CI, 0.4-5.4), and the follicle-stimulating hormone level significantly decreased by 0.06 ng/mL (95% CI, 0.01-0.11). Seminal fluid proteome analysis revealed differential expression of proteins enriched for processes related to cellular secretion, immune response, and fibrinolysis. Whole genome bisulfite sequencing identified 23,558 CpGs differentially methylated in heavy-THC vs. pre-THC sperm, with partial restoration of methylation after discontinuation of THC use. Genes associated with altered differentially methylated regions were enriched for those involved in the development and function of the nervous system. CONCLUSION(S): This is the first study demonstrating that discontinuation of chronic THC use in rhesus macaques partially restores adverse impacts to male reproductive health, THC-associated sperm differentially methylated regions in genes important for development, and expression of proteins important for male fertility.

Chronic prenatal delta-9-tetrahydrocannabinol exposure adversely impacts placental function and development in a rhesus macaque model.

Cannabis use in pregnancy is associated with adverse perinatal outcomes, which are likely mediated by the placenta. However, the underlying mechanisms and specific vasoactive effects of cannabis on the placenta are unknown. Our objective was to determine the impact of chronic prenatal delta-tetrahydrocannabinol (THC, main psychoactive component of cannabis) exposure on placental function and development in a rhesus macaque model using advanced imaging. Animals were divided into two groups, control (CON, n = 5) and THC-exposed (THC, n = 5). THC-exposed animals received a THC edible daily pre-conception and throughout pregnancy. Animals underwent serial ultrasound and MRI at gestational days 85 (G85), G110, G135 and G155 (full term is ~ G168). Animals underwent cesarean delivery and placental collection at G155 for histologic and RNA-Seq analysis. THC-exposed pregnancies had significantly decreased amniotic fluid volume (p < 0.001), placental perfusion (p < 0.05), and fetal oxygen availability (p < 0.05), all indicators of placental insufficiency. Placental histological analysis demonstrated evidence of ischemic injury with microinfarctions present in THC-exposed animals only. Bulk RNA-seq demonstrated that THC alters the placental transcriptome and pathway analysis suggests dysregulated vasculature development and angiogenesis pathways. The longer-term consequences of these adverse placental findings are unknown, but they suggest that use of THC during pregnancy may deleteriously impact offspring development.

Chronic exposure to delta-9-tetrahydrocannabinol impacts testicular volume and male reproductive health in rhesus macaques.

OBJECTIVE: To determine the dose-dependent effect of delta-9-tetrahydrocannabinol (THC) exposure on male testes and reproductive health in a nonhuman primate model. DESIGN: Research animal study. SETTING: Research institute. ANIMAL(S): Adult male rhesus macaques 8-10 years of age (n = 6). INTERVENTION(S): Daily edible THC at medically and recreationally relevant doses. MAIN OUTCOME MEASURE(S): Testicular volume and epididymal head width, serum levels of inhibin B, albumin, total testosterone, prolactin, follicle-stimulating hormone, estradiol, and luteinizing hormone; semen volume; and sperm motility, morphology, and concentration. RESULT(S): For each 1 mg/7 kg/day increase in THC dosing, there was a marked loss in total bilateral testicular volume of 11.8 cm3 (95% confidence interval [CI]: 8.3-15.4). In total, average bilateral testicular volume decreased by 58%. Significant dose-response decreases in mean total testosterone level by 1.49 ng/mL (95% CI: 0.83-2.15) and in estradiol level by 3.8 pg/mL (95% CI: 2.2-5.4) were observed, but significant increases in the levels of follicle-stimulating hormone by 0.06 ng/mL (95% CI: 0.02-0.10), luteinizing hormone by 0.16 ng/mL (95% CI: 0.08-0.25), and prolactin by 7.4 ng/mL (95% CI: 3.4-11.3) were observed. There were no statistically significant changes in semen parameters. CONCLUSION(S): In rhesus macaques, chronic exposure to THC resulted in significant dose-response testicular atrophy, increased serum gonadotropin levels, and decreased serum sex steroids, suggestive of primary testicular failure. Further studies are needed to determine if reversal of these observed adverse effects would occur if THC was discontinued and for validation of thefindings in a human cohort.

The effects of delta-9-tetrahydrocannabinol exposure on female menstrual cyclicity and reproductive health in rhesus macaques.

OBJECTIVE: To determine the dose-dependent effect of contemporary marijuana exposure on female menstrual cyclicity and reproductive endocrine physiology in a nonhuman primate model. DESIGN: Research animal study. SETTING: Research institute environment. ANIMALS: Adult female rhesus macaques (6-12 years of age; n = 8). INTERVENTIONS: Daily delta-9-tetrahydrocannabinol (THC) edible at medically and recreationally relevant contemporary doses. MAIN OUTCOME MEASURES: Menstrual cycle length (MCL), anti-Müllerian hormone, prolactin, basal follicle-stimulating hormone (FSH), estradiol (E2) and progesterone, luteinizing hormone (LH), and thyroid-stimulating hormone. RESULTS: The average before THC weight was 6.9 kg (standard deviation, 0.8), and at the highest THC dosing, the average weight was 7.2 kg (standard deviation, 0.8). With increasing THC dosing, MCL and FSH concentrations increased, while basal E2 concentration was stable. The average MCL concentration increased 4.0 days for each mg/7 kg/day of THC (95% CI, 1.4-6.6 days). Follicle-stimulating hormone concentration increased significantly with increasing THC dose, 0.34 ng/mL for each mg/7 kg/day of THC (95% CI, 0.14-0.57 ng/mL). No significant trends were observed between THC dosing and average basal progesterone, anti-Müllerian hormone, prolactin, LH, or thyroid-stimulating hormone concentrations. CONCLUSIONS: In rhesus macaques, a dose response toward increased MCL and basal FSH concentrations but plateau of basal E2 and LH concentrations was observed with increasing THC dosing, suggesting ovulatory dysfunction. Further studies are needed to determine the effects of a longer duration of exposure and whether the significant increase in MCL and FSH concentrations results in reduced fecundity.