Hyperbaric oxygen therapy ameliorates cyclophosphamide-induced spermatogenic impairment by suppressing the RhoA/ROCK...

Hyperbaric oxygen (HBO) therapy has shown potential to improve sperm quality and reproductive outcomes, but the mechanisms underlying its protective effects remain unclear. This study investigated whether HBO ameliorates cyclophosphamide (CYP)-induced spermatogenic impairmentand whether Rhoa/Rock signaling is involved. Male BALB/c mice were assigned to control, CYP, and CYP + HBO groups. CYP was administered intraperitoneally for five consecutive days, followed by HBO treatment at 2.5 ATA for 60 min/day, 5 days/week for 2 weeks. Sperm parameters, serum sex hormones, testicular histopathology, oxidative stress indices, and Rhoa/Rock-related proteins were assessed. Rhoa knockout (KO) mice were additionally used for mechanistic evaluation. Compared with CYP treatment alone, HBO treatment increased sperm count by 80.95% (P < 0.0001) and progressive motility by 160.96% (P < 0.0001), while decreasing the sperm abnormality rate by 38.88% (P < 0.0001). HBO also reduced testicular MDA levels by 40.90% (P < 0.0001) and increased SOD activity by 69.53% (P = 0.0005). These changes were accompanied by reduced Rhoa and Rock1 protein expression and increased Nrf2 and Ho-1 expression. In Rhoa KO mice, the beneficial effects of HBO on serum hormone levels, sperm parameters, testicular indices, oxidative stress markers, and Nrf2/Ho-1 expression were largely abolished. These findings indicate that HBO therapy alleviates CYP-induced testicular oxidative stress injury and spermatogenic impairment, with Rhoa/Rock signaling contributing to the protective response.