Optimizing thyroxine levels for enhanced buffalo sperm cryopreservation and fertility: a focus on quality, viability, and antioxidant protection

Introduction: This study investigated the effects of adding thyroxine (T4) to buffalo semen on sperm quality, oxidative markers, apoptosis-like changes, and fertility. Methods: Initially, we tested a wide range of T4 concentrations (0.1, 0.3, 0.9, 2.7, and 8.1 µg/dL) to evaluate their impact on motility and viability. Result and discussion: Lower concentrations (0.1–0.9 µg/dL) improved total and progressive motility and viability compared to higher concentrations (2.7 and 8.1 µg/dL). We assessed optimized doses (0.25, 0.5, 0.75, and 1 µg/dL) based on these findings. The 0.75 µg/dL group showed superior sperm velocity, viability, motion parameters, membrane, acrosome, and DNA integrity in equilibrated and frozen-thawed samples. Antioxidant markers (GPx, SOD, TAC) were enhanced, while MDA and apoptotic/necrotic cell levels were reduced, particularly in the 0.75 µg/dL group. Fertility trials revealed higher cryosurvival and conception rates in thyroxine-treated groups. In conclusion, T4 supplementation, especially at 0.75 µg/dL, enhances cryopreservation outcomes and fertility potential of buffalo bull semen.