Efficacy of the anticaspase agent zVAD-fmk on post-thaw viability of canine spermatozoa.

Cryopreservation protocols for gametes are constantly improved with the aim of increasing the post-thaw viability of gametes. It is becoming clear that stress, resulting from cryopreservation, reduces cell numbers by apoptosis. Apoptosis, or programmed cell death, is a gene-activated event that occurs as a normal consequence of development and as a result of cellular stress. Apoptosis is mediated by the family of cysteine-dependent asparate-specific proteases (caspases). The present study was designed to test the hypothesis that addition of an anticaspase (zVAD-fmk) that has inhibitory properties against caspases and apoptosis to semen extenders and to the thaw medium would increase post-thaw viability of canine spermatozoa. Extenders were added in a two-step process. A dose of 100 microM caspase inhibitor was used. Four groups (n=6 for each) were composed based on the presence or absence of the caspase inhibitor: Group I (control), no caspase inhibitor in the extender or the thaw medium; Group II, caspase inhibitor in the thaw medium; Group III, caspase inhibitor in Extender II; and Group IV, caspase inhibitor in both Extender II and the thaw medium. Post-thaw motility, plasma membrane integrity, and acrosome status were investigated. The addition of caspase inhibitor to Extender II or to the thaw medium failed to improve the parameters that were studied. The results suggest that this caspase inhibitor may not be beneficial to the post-thaw motility of canine spermatozoa if used at this concentration.