Evaluation of the efficacy of icariin against heat stress-induced spermatogenic dysfunction in the testes of dogs.

INTRODUCTION: This study examines the detrimental effects of high-temperature environments on canine testicular function and reproductive health, and investigates the potential of Epimedii Folium, particularly its active component icariin, in alleviating these effects and improving testicular function. METHODS: A completely randomized single-factor design was employed, involving 24 adult male Beagle dogs (9.82 ± 0.73 kg) assigned randomly to four treatment groups, with six dogs in each group. The groups included a negative control group ("Control"), a positive control group exposed to testicular heat stress ("Model"), and two icariin-treated groups receiving daily doses of 0.5 g/kg (Icariin-L) and 1.0 g/kg (Icariin-H), respectively. All groups, except the negative control, underwent a testicular heat stress model to induce damage and assess the effects of icariin on sperm quality, testicular function, hormone levels, protein expression, and testicular histological changes. RESULTS: Icariin supplementation improved sperm quality under heat stress, as indicated by increased total sperm count and motility, along with a reduction in sperm malformation rate (p < 0.01). It also restored adenosine triphosphate (ATPase) activities (Na(+)-K(+)-ATPase, Mg(2+)-ATPase, and Ca(2+)-ATPase) and serum hormone levels (GnRH, LH, and E2) (p < 0.01). Western blot analysis revealed that icariin upregulated steroidogenic proteins (STAR, 17βHSD, and CYP450) and the tight junction protein ZO-1 (p < 0.01), while downregulating the BAX expression (a key regulator of mitochondrial apoptosis) and enhancing the BCL-2 expression (a major anti-apoptotic factor in the BCL-2 family) (p < 0.01). Histological assessments demonstrated that icariin mitigated heat-induced damage to seminiferous tubules, epithelial thinning, and spermatogonia degeneration. Furthermore, molecular docking analysis confirmed a strong binding affinity between icariin and 17βHSD, mediated by hydrogen bonding and hydrophobic interactions. CONCLUSION: These findings suggest that icariin may alleviate testicular heat stress by modulating testosterone synthesis, enhancing ATPase function, restoring blood-testis barrier integrity, and inhibiting apoptosis. The dose-dependent efficacy (1.0 g/kg > 0.5 g/kg) supports the potential of icariin as a possible therapeutic agent for improving reproductive health in dogs exposed to high-temperature environments.