Abstract
A novel strategy, focused on the induction of sub-lethal oxidative stress to optimize sperm cryosurvival, has been used before cryopreservation. The present study compared the effect of preconditioning with various concentrations of nitric oxide-donor (sodium nitroprusside, SNP) and peroxynitrite-generator (3-morpholinosydnonimine, SIN-1) on in vitro sperm functions and lipid peroxidation status (LPO) of cryopreserved Karan-Fries (KF) crossbred bull semen. To optimize the concentration of additives, spermatozoa obtained from 36 ejaculates were supplemented with different concentrations of SNP (0.01, 0.05, 0.1μM) and SIN-1 (80, 160, 200μM) versus control in the extender. The post-freezing sperm motility and viability were greater (p < 0.05) in 0.1μM SNP and 80μM SIN-1 in comparison to other concentrations used. Furthermore, the spermatozoa obtained from 48 ejaculates were supplemented with 0.1μM SNP and 80μM SIN-1 in the extender. A significant increase (p < 0.05) was observed in progressive motility, viability and membrane integrity in SNP and SIN-1 treated extender at 24h, 15days, and 2-month post-cryopreservation (PC) periods. There was no significant difference in sperm abnormality in the extended groups and the control group. The seminal plasma of SNP-treated extender had less (p < 0.05) lipid peroxidation as compared to SIN-1 treated and control groups. In post-thaw semen, both SNP and SIN-1 showed a higher (p < 0.05) proportion of acrosome intact (FITC-PNA) sperm with a greater decrease (p < 0.05) in membrane scrambling and lipid peroxidation. SNP and SIN-1 improved (p < 0.05) the proportion of sperm with higher mitochondrial membrane potential (Δψm) as compared to the control. In conclusion, it seems that the preconditioning of SNP and SIN-1 at lower doses may have beneficial effects on post-thawed crossbred bull sperm quality.
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