Short- and long-term testosterone treatment induce reversible subfertility in a mouse model of gender-affirming hormone therapy

Transgender and gender-diverse adults may utilize testosterone (T) gender-affirming hormone therapy (GAHT) to achieve embodiment goals. Some T-GAHT users choose to become gestational parents but T-GAHT alters the hypothalamic-pituitary-gonadal axis, which is critical to maintaining early pregnancies, along with the reproductive tract. Importantly, gestation requires cessation of T-GAHT as high T is detrimental to offspring sexual development. We examined Ts potential to impact the gestational capacity of mice and offspring reproductive health after ceasing short- or long-term T-GAHT treatment. Adult CD1 female (n=14-15/group) mice were implanted with a capsule containing 10 mg T or left blank (B). Implants were left for 6 or 12 weeks before removal. Following three days of recovery, T-GAHT and control mice were housed with a proven breeder CD1 male. Breeding pairs were allowed to produce two litters before dams were sacrificed and tissue collected. Offspring were weaned and their sexual development tracked before collecting tissue. Tissues from dams and female offspring were collected during diestrus. Students t-tests were used to compare between T and B treated animals of the same treatment duration. T-tests were also used for offspring measurements, which were averaged by dam and separated by dam treatment duration and litter. Short- and long-term T-GAHT increased the time between pairing and first parturition compared with controls (Short B 23.2±1.4, Short T 25.5±4, Long B 22.3±1.3, Long T 24.5±3.1). Long-term treatment decreased pup numbers in first litters (Short B=11.5±2, Short T=11.4±4, Long B=11.9±3, Long T=7.8±3 Pups). Both short- and long-term T-GAHT influenced pubertal onset, measured by vaginal opening and preputial sheath separation, which was earlier for first-litter male and female pups from T-GAHT mice. The detrimental effects of T-GAHT on dam fertility and offspring development disappeared when examining their second gestation and litters. T-GAHT treatment caused permanent changes to dam anatomy, including larger clitoris areas (Short B=6.3) and heavier uterine horns (Short B=115.3±39, Short T=157.1±57, Long B=128.8±33, Long T=162.8±45.1 mg), while the impact of T-GAHT on offspring tissues was minimal and restricted to first litters. Both short- and long-term T-GAHT are detrimental to gestation and disrupt offspring sexual development. However, these effects are not permanent, and the subfertility seen during the first gestational period is lost in the second. Concurrently, offspring measures from second litters are not significantly impacted by dam T-GAHT treatment. Dams previously given T-GAHT retain anatomical changes to their reproductive system, which do not appear to impact fertility.