Influence of the Addition of Zinc Oxide Nanoparticles to Cryopreservation Medium for Dog Epididymal Spermatozoa

The purpose of this study was to assess the effect of different concentrations of zinc oxide nanoparticles (ZnONPs) introduced to an extender for frozen-thawed epididymal dog spermatozoa. Epididymides from 22 castrated dogs were minced and cultured in a Tris buffer. The recovered spermatozoa were diluted in Tris-Citric acid-Fructose (TCF) extender with different concentrations of ZnONPs (100 and 200 µg/mL) and control (0.0 µg/mL). Diluted samples were equilibrated at 5 °C for 2 hours before being packed in 0.25 mL straws and stored in liquid nitrogen (-196 °C). After thawing at 37°C for 30 seconds, sperm motility, viability, membrane integrity, acrosome integrity, DNA integrity, and lipid peroxidation by malondialdehyde (MDA) concentration were all measured. The results were presented as mean ± SEM. Adding 100 and 200 µg/mL ZnONPs to the cryopreservation medium significantly (P < .05) improved motility and membrane integrity compared to the control. Viability and acrosome integrity were considerably (P < .05) better at 100 µg/mL ZnONPs than at 200 µg/mL ZnONPs and the control. MDA concentration was significantly (P < .05) decreased at 100 µg/mL ZnONPs compared to 200 µg/mL ZnONPs and the control. When 100 µg/mL ZnONPs were compared to 200 µg/mL ZnONPs and the control, the percentage of DNA damage was significantly (P < .05) reduced. Consequently, adding 100 µg/mL ZnONPs to TCF extender resulted in a significant increase in the proportion of motility, viability, membrane-intact, and acrosome-intact dog epididymal sperm, as well as the preservation of DNA integrity and the prevention of lipid peroxidation at the membrane level.