Serpil SARIÖZKAN, Mustafa Numan BUCAK, Serhat BÜYÜKLEBLEBİCİ, Pürhan Barbaros TUNCER

Antioxidant supplementation ameliorates bull sperm parameters and fertilizing ability following the freeze-thaw process

This study’s goal was to reveal the effects of antioxidant supplementation on motility, motion characters, morphology, DNA integrity, and fertilizing potential of cryopreserved bovine sperm. The ejaculates were collected from three Holstein bulls. At least ten ejaculates were collected per bull. The ejaculates were immediately separated into five aliquots, diluted in the Cryobos (Magapor Co. Ltd., Zaragoza, Spain) commercial extender, including 2.5 mM taurine, 2.5 mM cysteine, 2.5 mM methionine, 2.5 mM glutamine, and no additives were used. Afterward, they were frozen in 0.25 mL French straws, and liquid nitrogen was used for the storage of semen. The supplementation of methionine resulted in a higher subjective motility rate in comparison with the other groups. Taurine led to the lowest post-thawed CASA motility rate (p < 0.05). The addition of antioxidants did not cause any improvements in sperm motion characteristics when compared to the controls (p > 0.05). Cysteine led to a higher protection of acrosome abnormality, in comparison with the other groups (p < 0.05). For the comet test, the minimum percentage of sperm with damaged DNA was obtained in the groups with cysteine and glutamine (p < 0.05). There was not any significant difference among the groups in terms of pregnancy rates (p > 0.05). In conclusion, cysteine supplementation to the semen extender prior to freezing ameliorated the post-thawed semen quality.

PDF

___

1. Watson P. Recent developments and concepts in the cryopreservation of spermatozoa and the assessment of their post-thawing function. Reproduction, Fertility and Development 1995; 7 (4): 871-891. doi: 10.1071/rd9950871
2. Medeiros CMO, Forell F, Oliveira ATD, Rodrigues JL. Current status of sperm cryopreservation: why isn’t it better? Theriogenology 2002; 57 (1): 327-344. doi:10.1016/S0093- 691X(01)00674-4
3. Agarwal A, Virk G, Ong C, du Plessis SS. Effect of oxidative stress on male reproduction. The World Journal of Men’s Health 2014; 32 (1): 1-17. doi: 10.5534/wjmh.2014.32.1.1
4. Flesch FM, Gadella BM. Dynamics of the mammalian sperm plasma membrane in the process of fertilization. Biochimica et Biophysica Acta (BBA)-Reviews on Biomembranes 2000; 1469 (3): 197-235. doi: 10.1016/S0304-4157(00)00018-6
5. Watson P. The causes of reduced fertility with cryopreserved semen. Animal Reproduction Science 2000; 60: 481-492. doi: 10.1016/s0378-4320(00)00099-3
6. Alvarez JG, Storey BT. Taurine, hypotaurine, epinephrine and albumin inhibit lipid peroxidation in rabbit spermatozoa and protect against loss of motility. Biology of Reproduction 1983; 29 (3): 548-555. doi: 10.1095/biolreprod29.3.548
7. Bansal AK, Bilaspuri GS. Impacts of oxidative stress and antioxidants on semen functions. Veterinary Medicine International 2010: 686137. doi: 10.4061/2011/686137
8. Lapointe J, Bilodeau JF. Antioxidant defenses are modulated in the cow oviduct during the estrous cycle. Biology of Reproduction 2003; 68 (4): 1157-1164. doi: 10.1095/ biolreprod.102.007476
9. Bucak MN, Tuncer PB, Sarıözkan S, Başpınar N, Taşpınar M et al. Effects of antioxidants on post-thawed bovine sperm and oxidative stress parameters: antioxidants protect DNA integrity against cryodamage. Cryobiology 2010; 61 (3): 248- 253. doi: 10.1016/j.cryobiol.2010.09.001
10. Sarıözkan S, Bucak MN, Tuncer PB, Ulutaş PA, Bilgen A. The influence of cysteine and taurine on microscopic–oxidative stress parameters and fertilizing ability of bull semen following cryopreservation. Cryobiology 2009; 58 (2): 134-138. doi: 10.1016/j.cryobiol.2008.11.006
11. Bucak MN, Sarıözkan S, Tuncer PB, Ulutaş PA, Akçadağ Hİ. Effect of antioxidants on microscopic semen parameters, lipid peroxidation and antioxidant activities in Angora goat semen following cryopreservation. Small Ruminant Research 2009; 81 (2): 90-95. doi: 10.1016/j.cryobiol.2010.05.005
12. Khlifaoui M, Battut I, Bruyas JF, Chatagnon G, Trimeche A, Tainturier D. Effects of glutamine on post-thaw motility of stallion spermatozoa: an approach of the mechanism of action at spermatozoa level. Theriogenology 2005; 63 (1): 138-149. doi: 10.1016/j.theriogenology.2004.04.012
13. Uysal O, Bucak MN. Effects of oxidized glutathione, bovine serum albumin, cysteine and lycopene on the quality of frozenthawed ram semen. Acta Veterinaria Brno 2007; 76 (3): 383- 390. doi: 10.2754/avb200776030383
14. Bucak M N, Uysal O. The role of antioxidants in freezing of Saanen goat semen. Indian Veterinary Journal 2008; 85 (2): 148-150.
15. Reed DJ, Orrenius S. The role of methionine in glutathione biosynthesis by isolated hepatocytes. Biochemical and Biophysical Research Communications 1977; 77 (4): 1257- 1264. doi: 10.1016/s0006-291x(77)80115-0
16. Bucak MN, Ateşşahin A, Varışlı Ö, Yüce A, Tekin N, Akçay A. The influence of trehalose, taurine, cysteamine and hyaluronan on ram semen: microscopic and oxidative stress parameters after freeze–thawing process. Theriogenology 2007; 67 (5): 1060-1067. doi: 10.1016/j.theriogenology.2006.12.004
17. Schäfer S, Holzmann A. The use of transmigration and Spermac™ stain to evaluate epididymal cat spermatozoa. Animal Reproduction Science 2000; 59 (3-4): 201-211. doi: 10.1016/s0378-4320(00)00073-7
18. Arabi M. Bull spermatozoa under mercury stress. Reproduction in Domestic Animals 2005; 40 (5): 454-459. doi: 10.1111/j.1439-0531.2005.00607.x
19. Henry MA, Noiles EE, Gao D, Mazur P, Critser JK. Cryopreservation of human spermatozoa. IV. The effects of cooling rate and warming rate on the maintenance of motility, plasma membrane integrity, and mitochondrial function. Fertility and Sterility 1993; 60 (5): 911-918. doi: 10.1016/ S0015-0282(16)56296-7
20. Kim SH, Yu DH, Kim YJ. Effects of cryopreservation on phosphatidylserine translocation, intracellular hydrogen peroxide, and DNA integrity in canine sperm. Theriogenology 2010; 73 (3): 282-292. doi: 10.1016/j.theriogenology. 2009.09.011
21. Kim SH, Yu DH, Kim YJ. Effects of cryopreservation on phosphatidylserine translocation, intracellular hydrogen peroxide, and DNA integrity in canine sperm. Theriogenology 2010; 73 (3): 282-292. doi: 10.1016/j.theriogenology.2009.09.011
22. Barroso G, Morshedi M, Oehninger S. Analysis of DNA fragmentation, plasma membrane translocation of phosphatidylserine and oxidative stress in human spermatozoa. Human Reproduction 2000; 15 (6): 1338-1344. doi: 10.1093/ humrep/15.6.1338
23. Bilodeau JF, Blanchette S, Gagnon C, Sirard MA. Thiols prevent H2 O2 -mediated loss of sperm motility in cryopreserved bull semen. Theriogenology 2001; 56 (2): 275-286. doi: 10.1016/ s0093-691x(01)00562-3
24. Barekat F, Tavalaee M, Deemeh MR, Bahreinian M, Azadi L et al. A preliminary study: N-Acetyl-L-Cysteine improves semen quality following varicocelectomy. International Journal of Fertility and Sterility 2016; 10 (1): 120-126. doi: 10.22074/ ijfs.2016.4777
25. Öğretmen F, İnanan BE, Kutluyer F, Kayım M. Effect of semen extender supplementation with cysteine on postthaw sperm quality, DNA damage, and fertilizing ability in the common carp (Cyprinus carpio). Theriogenology 2015; 83 (9): 1548- 1552. doi: 10.1016/j.theriogenology.2015.02.001
26. Ateşşahin A, Bucak MN, Tuncer PB, Kızıl M. Effects of antioxidant additives on microscopic and oxidative parameters of Angora goat semen following the freeze–thawing process. Small Ruminant Research 2008; 77 (1): 38-44. doi: 10.1016/j. smallrumres.2008.03.002
27. Amirat-Briand L, Bencharif D, Vera-Munoz O, Ali HBH, Destrumelle S, et al. Effect of glutamine on post-thaw motility of bull spermatozoa after association with LDL (low density lipoproteins) extender: preliminary results. Theriogenology 2009; 71 (8): 1209-1214. doi: 10.1016/j. theriogenology.2008.10.002
28. Sarıözkan S, Özdamar S, Türk G, Cantürk F, Yay A. In vitro effects of l-carnitine and glutamine on motility, acrosomal abnormality, and plasma membrane integrity of rabbit sperm during liquid-storage. Cryobiology 2014; 68 (3): 349-353. doi: 10.1016/j.cryobiol.2014.04.006
29. Petrunkina A, Waberski D, Günzel-Apel A, Töpfer-Petersen E. Determinants of sperm quality and fertility in domestic species. Reproduction 2007; 134 (1): 3-17. doi: 10.1530/REP07-0046
30. Agarwal A, Allamaneni S.S.R. The effect of sperm DNA damage on assisted reproduction outcomes. Minerva Ginecologica 2004; 56 (3): 235-245.
31. Henkel R, Hajimohammad M, Stalf T, Hoogendijk C, Mehnert C et al. Influence of deoxyribonucleic acid damage on fertilization and pregnancy. Fertility and Sterility 2004; 81 (4): 965-972. doi: 10.1016/j.fertstert.2003.09.044
32. Duru NK, Morshedi M, Schuffner A, Oehninger S. Cryopreservation-thawing of fractionated human spermatozoa and plasma membrane translocation of phosphatidylserine. Fertility and Sterility 2001; 75 (2): 263-268. doi: 10.1016/s0015- 0282(00)01694-0
33. Anzar M, He L, Buhr MM, Kroetsch TG, Pauls KP. Sperm apoptosis in fresh and cryopreserved bull semen detected by flow cytometry and its relationship with fertility. Biology of Reproduction 2002; 66 (2): 354-360. Doi: 10.1095/ biolreprod66.2.354
34. Peña FJ, Johannisson A, Wallgren M, Rodrıguez-Martınez H. Assessment of fresh and frozen–thawed boar semen using an Annexin-V assay: a new method of evaluating sperm membrane integrity. Theriogenology 2003; 60 (4): 677-689. doi: 10.1016/s0093-691x(03)00081-5
35. Peris SI, Morrier A, Dufour M, Bailey JL. Cryopreservation of ram semen facilitates sperm DNA damage: relationship between sperm andrological parameters and the sperm chromatin structure assay. Journal of Andrology 2004; 25 (2): 224-233. doi: 10.1002/j.1939-4640.2004.tb02782.x
36. Tuncer PB, Bucak MN, Büyükleblebici S, Sarıözkan S, Yeni D et al. The effect of cysteine and glutathione on sperm and oxidative stress parameters of post-thawed bull semen. Cryobiology 2010; 61 (3): 303-307. doi: 10.1016/j.cryobiol.2010.09.009