Beneficial and harmful effects of cyclodextrin-vitamin E complex on cryopreserved ram sperm

Vitamin E is a potent molecule, especially when loaded in cyclodextrin, in modulating oxidative stress during the freeze-thawing process. The present study aimed to investigate the effect of different concentrations of cyclodextrin-vitamin E complex (CD-Vit E) on cryopreserved ram sperm. Ejaculates collected from five adult rams were pooled and divided into four aliquots. All aliquots were treated in Tris-extender (Tris-glucose-citric acid) containing 2 mg cholesterol-loaded methyl-β- cyclodextrin/120 × 10⁶ spermatozoa and either 0 (Control), 2, 4 or 6 mg CD-Vit E/120 × 10⁶ spermatozoa, corresponding to 0, 0.5, 1 or 1.5 of pure vitamin E, respectively. After incubation at 22 °C for 15 min and the addition of Tris-extender containing glycerol and egg yolk (v/v), all aliquots were frozen in liquid nitrogen. After thawing, motility (computer aided sperm analysis), viability (eosin staining), membrane integrity (HOST), acrosome integrity (Coomassie G-250 staining) and lipid peroxidation (Thiobarbituric acid assay) were evaluated. Compared to control, 2 mg CD-Vit E had a significant positive effect on total motility, progressive motility, movement linearity (LIN%), viability and lipid peroxidation. At 4 mg, however, CD-Vit E had a significant negative effect on total motility, progressive motility, membrane functionality and acrosome integrity. At a greater concentration (6 mg), the negative effects were greater as compared with inclusion of 4 mg in the cryoprotectant and the percentage of rapidly and moderately motile gametes and viability were also altered. In conclusion, the effect of CD-Vit E on cryopreserved ram sperm was concentration-dependent with the 2 mg amount having a beneficial effect while greater concentrations (4 and 6 mg) had a harmful effect on sperm motility and gamete integrity but without affecting oxidative stress status.