ABSTRACT: Antioxidants have multiple protective roles in a variety of cells and thus can be used to protect sperm against cryo-damage during freezing, which affects fertility. The antioxidant resveratrol (3,5,4-trihydroxytrans-stilbene; RSV) has been reported to protect the animal sperm during cryopreservation, including human sperm. In this study, we assessed the protective effects of RSV supplementation on dog sperm cryopreservation. Semen was collected from four dogs and the effect of different concentrations of RSV (0, 100, 200, and 400 µM) on post-thaw sperm quality was examined. After thawing, sperm motility was assessed using computer-aided sperm analysis, and the structural integrity of the plasma membrane, acrosome, and chromatin was examined. In addition, their mitochondrial activity and gene expression were also assessed. Dog sperm cryopreserved with 200 µM RSV showed significant improvement in post-thaw sperm motility and viability compared with that of the control group (P<0.05). Moreover, RSV-supplemented samples showed significantly higher numbers of sperm with an intact plasma membrane, active mitochondria, and structural integrity of acrosomes and chromatin than that of control samples (P<0.05). Furthermore, gene expression showed that RSV supplemented samples showed lower expression of pro-apoptotic (BAX), reactive oxygen species (ROS) modulator oxidative stress-related (ROMO1) and 8-oxoguanine DNA glycosylase 1 (OGG1) whereas higher expression levels of anti-apoptotic (BCL2), protamine-2 (PRM2), protamine-3 (PRM3) and sperm acrosome-associated 3 (SPACA3) genes than control. Our results suggest that RSV, at its optimum concentration, can be efficiently used as an antioxidant in the cryopreservation of dog sperm.
