Effect of additives on short-term storage of ram semen

Year 2021, , 22 - 28, 30.04.2021

Caner Ozturk , Şükrü Dursun , Gaye Bulut , Tahir Karaşahin

https://doi.org/10.31797/vetbio.833211

Abstract

The aim of this study was to investigate the effects of short-term stored ram semen (24-hour intervals) on spermatological parameters up to 72 hours. In the study, % motility, viability and acrosome integrity were evaluated.
In thıs study four ejaculates from each ram (four merino rams) were used. The ejaculates were divided into seven equal-sized pieces after pooling. The study groups were formed as resveratrol (2 and 4 mM), trolox (2 and 4 mM), BSA (3 and 6 mg/ml) and control added to tris-based diluent. Diluted samples were stored at 4 ° C until 72 hours. Spermatozoa motility, viability and acrosome integrity were evaluated at 0, 24, 48 and 72 hours. Motility examination was evaluated with phase contrast microscope at 400X magnification, vitality and acrosome integrity were evaluated with fluorescent microscope.
At the 72nd hour of the study, the BSA (3 and 6mM) groups were assessed for motility (53.75 ± 2.50%; 55.00 ± 4.08%) and acrosome integrity (53.70 ± 3.39%; 57.10 ± 4.68%) in the control group (43.75 ± 2.50%; 46.53 ± 3.58%) and statistical difference (p <0.05). In the Trolox (4mM) group (67.22 ± 3.71%), the highest viability was achieved at 72 hours compared to all groups, and a statistical difference (p <0.05) was determined with the control group (46.53 ± 3.53).
As a result of the study, it was determined that BSA and trolox added to ram semen extender were beneficial in the short-term storage of ram semen and had a protective effect on spermatological parameters.

Keywords

BSA, short term storage, trehalose, trolox

Koç spermasının kısa süreli saklanmasında katkı maddelerinin etkisi

Abstract

Bu çalışmanın amacı, kısa süreli saklanan koç spermasının (24 saatlik aralıklarla) 72 saate kadar spermatolojik parametreler üzerine etkilerini araştırmaktı. Çalışmada % motilite, canlılık ve akrozom bütünlüğü verileri değerlendirildi.
Çalışmada her koçtan (4 baş Merinos koçu) alınan dört ejakülat kullanıldı. Ejakülatlar pooling yapıldıktan sonra eşit hacimli yedi parçaya bölündü. Çalışma grupları, tris bazlı sulandırıcıya eklenen resveratrol (2 ve 4 mM), troloks (2 ve 4 mM), BSA (3 ve 6 mg/ml) ve kontrol olmak üzere oluşturuldu. Sulandırılan örnekler 72. saate kadar 4°C de muhafaza edildi. Spermatozoa motilitesi, canlılık ve akrozom bütünlüğü 0, 24, 48 ve 72. saatte değerlendirildi. Motilite muayenesi faz kontrast ataçmanlı mikroskopta 400X lük büyütmede, canlılık ve akrozom bütünlüğü ise floresan mikroskop ile değerlendirildi.
Çalışmanın 72. saatinde, BSA (3 ve 6mM) grupları motilite (53.75±2.50%; 55.00±4.08%) ve akrozom bütünlüğü (53.70±3.39%; 57.10±4.68%) değerlendirmelerinde kontrol grubu (43.75±2.50%; 46.53±3.58) ile istatistiksel farklılık (p<0.05) gösterdi. Troloks (4mM) grubunda (67.22±3.71%), tüm gruplara kıyasla 72. saatte en yüksek canlılık sonucuna ulaşılmış ayrıca kontrol grubu (46.53±3.53) ile istatistiksel farklılık (p<0.05) belirlenmiştir.
Çalışmanın sonucunda koç sperma sulandırıcısına eklenen BSA ve troloksun koç spermasının kısa süreli saklamasında faydalı olduğu ve spermatolojik parametreler üzerine koruyucu etki gösterdiği belirlenmiştir.

Keywords

BSA, kısa süreli saklama, trehaloz, troloks

References

• Ball, B. A., Medina, V., Gravance, C. G., & Baumber, J. (2001). Effect of antioxidants on preservation of motility, viability and acrosomal integrity of equine spermatozoa during storage at 5 C. Theriogenology, 56(4), 577-589.
• Baumber, J., Sabeur, K., Vo, A., & Ball, B. A. (2003). Reactive oxygen species promote tyrosine phosphorylation and capacitation in equine spermatozoa. Theriogenology, 60(7), 1239-1247.
• Bilodeau, J. F., Blanchette, S., Gagnon, C., & Sirard, M. A. (2001). Thiols prevent H2O2-mediated loss of sperm motility in cryopreserved bull semen. Theriogenology, 56(2), 275-286.
• Branco, C. S., Garcez, M. E., Pasqualotto, F. F., Erdtman, B., & Salvador, M. (2010). Resveratrol and ascorbic acid prevent DNA damage induced by cryopreservation in human semen. Cryobiology, 60(2), 235-237.
• Brzezińska-Ślebodzińska, E., Ślebodziński, A. B., Pietras, B., & Wieczorek, G. (1995). Antioxidant effect of vitamin E and glutathione on lipid peroxidation in boar semen plasma. Biological trace element research, 47(1-3), 69-74.
• Bucak, M. N., & Tekin, N. (2007). Protective effect of taurine, glutathione and trehalose on the liquid storage of ram semen. Small Ruminant Research, 73(1-3), 103-108.
• da Silva Maia, M., Bicudo, S. D., Sicherle, C. C., Rodello, L., & Gallego, I. C. S. (2010). Lipid peroxidation and generation of hydrogen peroxide in frozen-thawed ram semen cryopreserved in extenders with antioxidants. Animal Reproduction Science, 122(1-2), 118-123.
• De La Lastra, C. A., & Villegas, I. (2007). Resveratrol as an antioxidant and pro-oxidant agent: mechanisms and clinical implications. Biochemical Society Transactions, 35 (5), 1156–1160.
• Diaz-Jimenez, M., Dorado, J., Ortiz, I., Consuegra, C., Pereira, B., Gonzalez-De Cara, C. A., Aguilera R., Mari G., Mislei B., Love C.C., & Hidalgo, M. (2018). Cryopreservation of donkey sperm using non-permeable cryoprotectants. Animal reproduction science, 189, 103-109.
• Domínguez-Rebolledo, Á. E., Fernández-Santos, M. R., García-Álvarez, O., Maroto-Morales, A., Garde, J. J., & Martínez-Pastor, F. (2009). Washing increases the susceptibility to exogenous oxidative stress in red deer spermatozoa. Theriogenology, 72(8), 1073-1084.
• El-Kon, I. (2011). Testing usability of bovine serum albumin (BSA) for preservation of Egyptian buffalo semen. American-Eurasian Journal of Agricultural & Environmental Sciences, 11(4), 495-502.
• Garcez, M. E., dos Santos Branco, C., Lara, L. V., Pasqualotto, F. F., & Salvador, M. (2010). Effects of resveratrol supplementation on cryopreservation medium of human semen. Fertility and sterility, 94(6), 2118-2121.
• Giaretta, E., Bucci, D., Mari, G., Galeati, G., Love, C. C., Tamanini, C., & Spinaci, M. (2014). Is resveratrol effective in protecting stallion cooled semen? Journal of Equine Veterinary Science, 34(11-12), 1307-1312.
• Giulivi, C., & Cadenas, E. (1993). Inhibition of protein radical reactions of ferrylmyoglobin by the water-soluble analog of vitamin E, Trolox C. Archives of biochemistry and biophysics, 303(1), 152-158.
• Gungor, S., Ozturk, C., & Omur, A. D. (2017). Positive effects of trehalose and cysteine on ram sperm parameters. Veterinární medicína, 62(5), 245-252.
• Gürler, H., Malama, E., Heppelmann, M., Calisici, O., Leiding, C., Kastelic, J. P., & Bollwein, H. (2016). Effects of cryopreservation on sperm viability, synthesis of reactive oxygen species, and DNA damage of bovine sperm. Theriogenology, 86(2), 562-571.
• Jang, M., Cai, L., Udeani, G. O., Slowing, K. V., Thomas, C. F., Beecher, C. W., Fong, H. H., Farnsworth, N. R., Kinghorn, A. D., Mehta, R. G., Moon, R. C., & Pezzuto J. M. (1997). Cancer chemopreventive activity of resveratrol, a natural product derived from grapes. Science, 275(5297), 218-220.
• Ledesma, A., Fernández-Alegre, E., Cano, A., Hozbor, F., Martínez-Pastor, F., & Cesari, A. (2016). Seminal plasma proteins interacting with sperm surface revert capacitation indicators in frozen-thawed ram sperm. Animal reproduction science, 173, 35-41.
• Mata-Campuzano, M., Álvarez-Rodríguez, M., Tamayo-Canul, J., López-Urueña, E., de Paz, P., Anel, L., Martínez-Pastor F., & Álvarez, M. (2014). Refrigerated storage of ram sperm in presence of Trolox and GSH antioxidants: Effect of temperature, extender and storage time. Animal reproduction science, 151(3-4), 137-147.
• Matsuoka, T., Imai, H., Kohno, H., & Fukui, Y. (2006). Effects of bovine serum albumin and trehalose in semen diluents for improvement of frozen-thawed ram spermatozoa. Journal of Reproduction and Development, 52(5), 675-683.
• Minaei, M. B., Barbarestani, M., Nekoonam, S., Abdolvahabi, M. A., Takzare, N., Asadi, M. H., Hedayatpour A., & Amidi, F. (2012). Effect of Trolox addition to cryopreservation media on human sperm motility. Iranian journal of reproductive medicine, 10(2), 99.
• O’Hara, L., Hanrahan, J. P., Richardson, L., Donovan, A., Fair, S., Evans, A. C. O., & Lonergan, P. (2010). Effect of storage duration, storage temperature, and diluent on the viability and fertility of fresh ram sperm. Theriogenology, 73(4), 541-549.
• Ourique, G. M., Finamor, I. A., Saccol, E. M., Riffel, A. P., Pês, T. S., Gutierrez, K., Gonçalves, P.B.D., Baldisserotto, B., Pavanato M. A., & Barreto, K. P. (2013). Resveratrol improves sperm motility, prevents lipid peroxidation and enhances antioxidant defences in the testes of hyperthyroid rats. Reproductive Toxicology, 37, 31-39.
• Peña, F. J., Johannisson, A., Wallgren, M., & Martinez, H. R. (2003). Antioxidant supplementation in vitro improves boar sperm motility and mitochondrial membrane potential after cryopreservation of different fractions of the ejaculate. Animal reproduction science, 78(1-2), 85-98.
• Said, T. M., Gaglani, A., & Agarwal, A. (2010). Implication of apoptosis in sperm cryoinjury. Reproductive biomedicine online, 21(4), 456-462.
• Sarıözkan, S., Bucak, M. N., Tuncer, P. B., Ulutaş, P. A., & Bilgen, A. (2009). The influence of cysteine and taurine on microscopic–oxidative stress parameters and fertilizing ability of bull semen following cryopreservation. Cryobiology, 58(2), 134-138.
• Silva, S. V., Soares, A. T., Batista, A. M., Almeida, F. C., Nunes, J. F., Peixoto, C. A., & Guerra, M.M.P. (2013). Vitamin E (Trolox) addition to Tris-egg yolk extender preserves ram spermatozoon structure and kinematics after cryopreservation. Animal reproduction science, 137(1-2), 37-44.
• Taylor, K., Roberts, P., Sanders, K., & Burton, P. (2009). Effect of antioxidant supplementation of cryopreservation medium on post-thaw integrity of human spermatozoa. Reproductive Biomedicine Online, 18(2), 184-189.
• Tremellen, K. (2008). Oxidative stress and male infertility—a clinical perspective. Human reproduction update, 14(3), 243-258.
• Uğuralp, S., Usta, U., & Mizrak, B. (2005). Resveratrol may reduce apoptosis of rat testicular germ cells after experimental testicular torsion. European Journal of Pediatric Surgery, 15(5), 333-336.
• Uysal, O., & Bucak, M. N. (2007). Effects of oxidized glutathione, bovine serum albumin, cysteine and lycopene on the quality of frozen-thawed ram semen. Acta Veterinaria Brno, 76(3), 383-390.
• Valko, M., Leibfritz, D., Moncol, J., Cronin, M. T., Mazur, M., & Telser, J. (2007). Free radicals and antioxidants in normal physiological functions and human disease. The international journal of biochemistry & cell biology, 39(1), 44-84.
• Zhang, X., Li, X., & Xia, L. (2015). Expression of a thermo-alkaline lipase gene from Talaromyces thermophilus in recombinant Pichia pastoris. Biochemical Engineering Journal, 103, 263-269.
• Zhu, Z., Li, R., Fan, X., Lv, Y., Zheng, Y., Hoque, S. A., Wu D., & Zeng, W. (2019). Resveratrol improves boar sperm quality via 50AMP-activated protein kinase activation during cryopreservation. Oxidative Medicine and Cellular Longevity, 1-15.