earticle

영어

We determined the optimal polyvinyl Alcohol(PVA) concentration in previous study. However, the sperm progressive motility was lower compared to viability. To improve the motility, we determined the effect of pH adjustment in PVA-extender and post-thaw incubation time on dog sperm viability, motility, reactive oxygen species(ROS) level and gene expression levels related to motility(SMCP) and apoptosis(Bcl-2). Spermatozoa collected from five dogs were resuspended (1×108 cell/mL) with Tris extender 1 containing 0.1, 0.2, 0.3 g PVA with non-adjustment of pH (6.45) and adjustment of pH (6.85), respectively. The extended spermatozoa were cooled to 4℃ for 1 h. The cooled spermatozoa were diluted (1:1) with Tris extender 2 containing 1 M glycerol and PVA concentrations used in the 1st dilution and then loaded in 0.5 mL straws. The straws were kept at 4℃ for additional 30 min and were frozen using LN2 vapor. Sperm viability was assessed after cooling and post-thaw incubation at 24°C for 20 min. Sperm progressive motility was assessed after cooling, post-cooling incubation at 24°C for 20 min, immediately after thawing and post-thaw incubation at 24°C for 20 min. In addition, ROS level and the relative abundance of Bcl-2 and SMCP mRNA(qRT-PCR) were measured following postthaw incubation. As results, adjustment of pH to 6.85 in PVA-Tris extender significantly (p<0.05) improved sperm progressive motility compared to non-adjustment groups with no significant effect on sperm viability, ROS and mRNA expression level of Bcl-2 and SMCP. Post-thaw incubation at 24°C for 20 min significantly improved the progressive motility in all pH adjusted groups. We conclude that adjustment of pH to 6.85 and post-thaw incubation in PVA-Tris extender could considerably improve motility in dog sperm cryopreservation.