Effects of resveratrol supplementation to equine sperm during semen cryopreservation

During equine semen cryopreservation, most of the seminal plasma is removed, making sperm more susceptible to oxidative stress. This study evaluated the effect of resveratrol supplementation on the quality of frozen-thawed equine sperm. Semen from ten stallions was frozen using a control INRA 96 extender and extenders supplemented with 5-, 10-, 100-, and 150- µM resveratrol. Post-thaw evaluations included motility, kinematic parameters, morphology, plasma and acrosomal membrane integrity, mitochondrial potential, lipid peroxidation, nitrite, hydrogen peroxide, malondialdehyde levels, total reactive oxygen species (ROS), chromatin protamine deficiency, chromatin condensation, sperm binding to bovine oviduct explants, and gene expression of apoptosis-related genes such as B cell lymphoma 2 (BCL2) and BCL2-associated X (BAX), mitochondrial ROS modulator 1 associated with mitochondrial ROS production (ROMO1), sperm acrosome-associated 3 related with sperm binding capacity to the zona pellucida (SPACA3), and DNA damage repair gene 8-oxoguanine DNA glycosylase 1 (OGG1). Data were analyzed by ANOVA and Tukey test (P < 0.05). The 10 µM resveratrol treatment significantly increased sperm motility, mitochondrial activity, SPACA3 expression, and the number of sperm bound to oviduct explants compared to the control. Additionally, 10 µM resveratrol reduced total ROS, ROMO1, and BAX gene expression, indicating reduced oxidative stress and apoptosis. In conclusion, 10 µM resveratrol supplementation improved sperm metabolic activity, enhanced oviduct binding capacity, and demonstrated antioxidant and anti-apoptotic effects. Thus, incorporating resveratrol into freezing extenders could be a promising strategy to improve the fertilizing capacity of equine sperm in artificial insemination programs.