The Use of κ-Carrageenan in Egg Yolk Free Extender Improves the Efficiency of Canine Semen Cryopreservation

SIMPLE SUMMARY: Semen cryopreservation generates sperm damage and reduces fertilization capacity as a consequence of reactive oxygen species formation, a known trigger of the apoptosis cascade. The present study aimed to examine the effects of κ-carrageenan supplementation to the freezing medium on canine sperm cryo-survival. To this end, the sperm was frozen in egg-yolk-free diluent containing different κ-carrageenan concentrations (0.0%, 0.1%, 0.2%, 0.3%, and 0.5%). The addition of κ-carrageenan to the extender at a 0.2% concentration resulted in a significant increase in the total motility and rapid progressive motility. Increasing levels of κ-carrageenan increased the spermatozoa acrosome integrity (p < 0.05). Apoptosis levels were significantly lower in the 0.1% and 0.2% κ-carrageenan treatment. Moreover, the κ-carrageenan supplemented group exhibited an upregulation of the relative expression of anti-apoptotic and down-regulation of oxidative state-related genes (p < 0.05). Overall, the introduction of κ-carrageenan as a component of egg-yolk-free freezing diluent could be useful for defining the cryopreservation media and improving the overall efficiency of cryopreserved canine semen. ABSTRACT: κ-Carrageenan is a plant polysaccharide derived from red seaweeds reported to possess potential medicinal and antioxidants activities. The present study aimed to identify the cryoprotective effects of κ-carrageenan on the quality of frozen-thawed canine semen. Twenty-eight ejaculates were collected and diluted in a Tris egg-yolk-free extender supplemented with various concentrations of κ-carrageenan (0.0%, 0.1%, 0.2%, 0.3%, and 0.5%). The addition of κ-carrageenan to the extender at a 0.2% concentration induced a significant increase in the total motility (TM) and the rapid progressive motility (RPM) of canine sperm. Among the experimental groups, the highest percentage of sperms with intact acrosomes was found in the 0.5% κ-carrageenan group (p < 0.05). Apoptosis levels were significantly lower in the 0.1% and 0.2% κ-carrageenan treatment. Moreover, sperm in the κ-carrageenan supplemented group showed a significantly higher expression of antiapoptotic (Bcl-2) and lower expression of NADPH oxidase (NOX5), spermine synthase (SMS), and spermine oxidase (SMOX) genes than those in the control group. In conclusion, the addition of κ-carrageenan to the freezing extender improved the overall efficiency of frozen-thawed dog spermatozoa.