Cryopreservation of epididymal cat spermatozoa: effects of in vitro antioxidative enzymes supplementation and lipid peroxidation induction

• Published in: Theriogenology 2010-05, Vol.73 (8), p.1076-1087
• Main Authors: Thuwanut, P., Chatdarong, K., Johannisson, A., Bergqvist, A.-S., Söderquist, L., Axnér, E.
• Format: Article
• Language: English
• Subjects: additives; Animal and Dairy Science; Animals; Antioxidant; antioxidant activity; antioxidants; Antioxidants - metabolism; Antioxidants - pharmacology; assisted reproductive technologies; catalase; Catalase - metabolism; Catalase - pharmacology; Cats; cryopreservation; Cryopreservation - methods; Cryopreservation - veterinary; Cryoprotective Agents - pharmacology; Enzymes - metabolism; Enzymes - pharmacology; epididymis; Feline; freeze-thaw cycles; germplasm conservation; glutathione peroxidase; Husdjursvetenskap; in vitro studies; iron; lipid peroxidation; Lipid Peroxidation - drug effects; Male; male fertility; Oxidative stress; protective effect; Reactive Oxygen Species - pharmacology; Semen Analysis - methods; Semen Analysis - veterinary; semen extenders; Semen Preservation - methods; Semen Preservation - veterinary; sperm motility; Sperm Retrieval - veterinary; Spermatozoa; superoxide dismutase; Superoxide Dismutase - metabolism; Superoxide Dismutase - pharmacology; Tromethamine - pharmacology; Up-Regulation - drug effects; Veterinary Science; Veterinärmedicin
• ISSN: 0093-691X; 1879-3231; 1879-3231
• DOI: 10.1016/j.theriogenology.2010.01.007

Reactive oxygen species and lipid peroxidation reaction, causes of sperm damage, can be diminished by action of antioxidative enzymes. This study aimed to investigate effects of (1) the antioxidative enzymes; catalase, glutathione peroxidase and superoxide dismutase, on epipididymal cat sperm quality and (2) the lipid peroxidation reaction induced by a transition metal (ferrous ion (II); Fe 2+) on sperm quality during the cryopreservation process. Epididymal spermatozoa harvested from 39 male cats were pooled and divided into 13 aliquots (n = 13). Each aliquot was resuspended with either a Tris egg yolk extender I (control; EE-I), or the Tris egg yolk extender I supplemented with 200 U/mL catalase (EE-CAT), or 10 U/mL glutathione peroxidase (EE-GPx), or 600 U/mL superoxide dismutase (EE-SOD), and then cryopreserved. After thawing, each sperm sample was subdivided into two groups; with and without lipid peroxidation induction (EE-I plus Fe 2+, EE-CAT plus Fe 2+, EE-GPx plus Fe 2+ and EE-SOD plus Fe 2+). Subjective sperm motility, membrane, and acrosome integrity were evaluated at the time of collection, after cooling, and at 0, 2, 4, and 6 h after thawing. Motility patterns assessed by computer-assisted sperm analysis (CASA), mitochondrial activity, and DNA integrity were evaluated during post-thaw incubation, whereas percentage of lipid peroxidation was detected at 0 and 6 h after thawing. The results demonstrate that catalase supplementation reduced linear motility and subjective motility immediately and 2 h after thawing (P < 0.05). Catalase supplementation, however, improved DNA integrity at 4 h (P < 0.05). Supplementation with glutathione peroxidase, compared to the control group, had a statistically significant positive effect on subjective motility at 0 and 6 h, linear motility at 6 h, mitochondrial activity at 6 h, membrane integrity at 2 and 6 h, and DNA integrity at 4 h after thawing. Although superoxide dismutase had a positive effect on sperm membrane integrity at 2 h after thawing (P < 0.05), it significantly reduced membrane integrity after cooling, linear motility at thawing, and acrosome integrity at 2 h after thawing. None of the three selected antioxidative enzymes significantly influenced acrosome integrity and none reduced the level of lipid peroxidation. Furthermore, induction of the lipid peroxidation reaction by Fe 2+ negatively affected most of the sperm quality parameters, i.e., motility and DNA integrity, during post-thaw sperm inc