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Dog Sperm Cryopreservation Using Glucose in Glycerol-free TRIS: Glucose Concentration, Exposure Time  

Yu, Il-Jeoung (Department of Theriogenology and Reproductive Biotechnologies, College of Veterinary Medicine and Bio-safety Research Institute, Chonbuk National University)
Publication Information
Journal of Veterinary Clinics / v.30, no.6, 2013 , pp. 442-448 More about this Journal
Abstract
The aim of the present study was to develop glycerol-free TRIS extender using glucose for dog sperm cryopreservation. We determined the appropriate concentration of glucose in glycerol-free TRIS and the exposure time in glycerol-free TRIS containing 0.3 M glucose at $4^{\circ}C$. Ejaculates of six dog sperm were cooled in glycerol-free TRIS through $4^{\circ}C$ for 100 min, cooled at $4^{\circ}C$ in TRIS with different glucose concentrations 0 M, 0.04 M, 0.1 M, 0.2 M and 0.3 M, respectively for 30 min followed by cryopreservation. After thawing at $37^{\circ}C$ for 25 sec, membrane and acrosome integrities of dog sperm were evaluated. In addition, the effect of exposure time (10, 30, 50 and 70 min) of sperm to glycerol-free TRIS containing 0.3 M glucose at $4^{\circ}C$ on progressive motility, viability, and DNA integrity following sperm cryopreservation was studied. Membrane integrity and acrosome integrity were assessed by 6-carboxyfluoresceindiacetate (6-CFDA)/propidium iodide (PI) fluorescent staining and Pisum sativum agglutinin conjugated to fluorescein isothiocyanate, respectively. DNA integrity was assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling, using flow cytometry. Sperm frozen in glycerol-free TRIS supplemented with 0.2 M or 0.3 M glucose have an intact plasma membrane (CFDA+/PI-) after cryopreservation than sperm frozen in the extenders with lower glucose concentrations (p<0.05). Acrosome integrity was significantly higher in the 0.3 M group than less than 0.1 M groups (p<0.05). The sperm DNA fragmentation index did not differ according to exposure time, although progressive motility was significantly higher in the 50 min exposure group than the other groups (p<0.05). These results indicate that cryopreservation of dog sperm is feasible and yields more motile sperm following freezing and thawing in glycerol-free TRIS containing 0.3 M glucose with the exposure time for 50 min at $4^{\circ}C$.
Keywords
Dog; sperm; cryopreservation; Glycerol-free TRIS; glucose;
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1 Rota B, Ström B, Linde-Forsberg C, Rodriguez-Martinez H, Effects of equex STM paste on viability of frozen-thawed dog spermatozoa during in vitro incubation at $38{^{\circ}C}$. Theriogenology 1997; 47: 1093-1101.   DOI   ScienceOn
2 Sanchez R, Risopatron J, Schulz M, Villegas J, Isanchenko V, Kreinberg R, Isachenko E. Canine sperm vitrification with sucrose: effect on sperm function. Angrologia 2011; 43: 233-241.
3 Sariozkan S, Bucak MN, Canturk F, Ozdamar S, Yay A, Tuncer PB, Ozcan S, Dorgucu N, Caner Y. The effect of different sugars on motility, morphology and DNA damage during the liquid storage of rat epididymal sperm at 4oC. Cryobiology 2012; 65: 93-97.   DOI
4 Shamsi MB, Imam SN, Dada R. Sperm DNA integrity assays: diagnostic and prognostic challenges and implications in management of infertility. J Assist Reprod Genet 2011; 28:1073-1085.   DOI
5 Strauss G, Shcurtenberger P, Hauser H. The interaction of saccharides with lipid layer vesicles: stabilization during freeze-thawing and freezing-drying. Biochim Biophys Acta 1986; 858: 169-180.   DOI   ScienceOn
6 Zribi N, Chakroun NF, Abdallah FB, Elleuch H, Sellami A, Gargouri J, Rebai T, Fahfakh F, Keskes LA. Effect of freezing-thawing process and quercetin on human sperm survival and DNA integrity. Cryobiology 2012; 65: 326-331.   DOI   ScienceOn
7 Yu I, Leibo SP. Recovery of motile, membrane-intact spermatozoa from canine epididymides stored for 8 days at $4{^{\circ}C}$. Theriogenology 2002; 57: 1179-1190.   DOI   ScienceOn
8 Watson PF. The causes of reduced fertility with cryopreserved semen. Anim Reprod Sci 2000; 60-61: 481-492.   DOI   ScienceOn
9 Holt WV. Basic aspects of frozen storage of semen. Anim Reprod Sci 2000; 62: 3-22.   DOI   ScienceOn
10 Koshimoto C, Gamliel E, Mazur P. Effect of osmolality and oxygen tension on the survival of mouse sperm frozen to various temperatures in various concentrations of glycerol and raffinose. Cryobiology 2000; 41: 204-231.   DOI   ScienceOn
11 Merino O, Risopatrón J, Sánchez R, Isachenko E, Figueroa E, Isachenko I, Fish (Oncorhynchus mykiss) spermatozoa cryoprotectant-free vitrification: stability of mitochondrion as criterion of effectiveness. Anim Reprod Sci 2011; 124: 125-131.   DOI   ScienceOn
12 Koshimoto C, Mazur P. The effect of the osmolality of sugar-containing media, the type of sugar, and the mass and molar concentration of sugar on the survival of frozenthawed mouse sperm. Cryobiology 2002; 45: 80-90.   DOI   ScienceOn
13 Malo C, Gil L, Gonzalez N, Cano R, de Blas L, Espinosa E. Comparing sugar type supplementation for cryopreservation of boar semen in egg yolk based extender. Cryobiology 2010; 21: 17-21.
14 Marcias Garcia B, Ortega Ferrusola C, Aparicio IM, Miro- Moran A, Morillo Rodriguez A, Gallardo Bolanos JM, Gonzalez Fernandez L, Balao da Silva CM, Rodriguez Martinez H, Tapia JA, Pena FJ. Toxicity of glycerol for the stallion spermatozoa: effects on membrane integrity and cytoskeleton, lipid peroxidation and mitochondria membrane potential. Theriogenology 2012; 77: 1280-1289.   DOI   ScienceOn
15 Naing SW, Wahid H, Mohd Azam K, Rosnina Y, Zuki AB, Kazhal S, Bukar MM, Thein M, Kyaw T, San MM. Effect of sugars on characteristics of Boer goat semen after cryopreservation. Anim Reprod Sci 2010; 122: 23-28.   DOI   ScienceOn
16 Paasch U, Sharma RK, Gupta AK, Grunewald S, Mascha EJ, Thomas AJ, Glander HJ, Agarwal A. Cryopreservation and thawing is associated with varying extent of activation of apoptotic machinery in subsets of ejaculated human spermatozoa. Biol Reprod 2004; 71: 1828-1837.   DOI   ScienceOn
17 Peris SL, Bilodeau JF, Duflour M, Bailey JL. Impact of cryopreservation and reactive oxygen species on DNA integrity, lipid peroxidation, and functional parameters in ram sperm. Mol Reprod Dev 2007; 74: 878-892.   DOI   ScienceOn
18 Rota A, Milani C, Cabianca G, Martini M. Comparison between glycerol and ethylene glycol for dog semen cryopreservation. Theriogenology 2006; 65: 1848-1858.   DOI   ScienceOn
19 Arenas Núñez MA, Juárez-Mosqueda MD, Gutiérrez-Pérez O, Anzaldúa Arce SR, Izquierdo AC, Rodríguez RM, Trujillo Ortega ME. Glycerol decreases the integrity of the perinuclear theca in boar sperm. Zygote 2013; 21: 172-177.   DOI   ScienceOn
20 Asien EG, Medina VH, Venturino A. Cryopreservation and post-thawed fertility of ram semen frozen in different trehalose concentrations. Theriogenology 2002; 57: 1801-1808.   DOI   ScienceOn
21 Bailey JL, Lessard C, Jacques J, Breque C, Dobrinski I, Zeng W, Galantino-Homer HL. Cryopreservation of boar semen and its future importance to the industry. Theriogenology 2008; 70: 1251-1259.   DOI   ScienceOn
22 Gomez-Fernandez J, Gomez-Izquierdo E, Tomas C, Moce E, de Mercado E. Effect of different monosaccharides and disaccharides on boar sperm quality after cryopreservation. Anim Reprod Sci 2012; 33: 109-116.
23 Fuller BJ. Cryoprotectants: the essential antifreezes to protect life in the frozen state. CryoLetters 2004; 25: 375-388.
24 Aboagla EM, Terada T. Trehalose-enhanced fluidity of the goat sperm membranes and its protection during freezing. Biol Reprod 2003; 69: 1245-1250.   DOI   ScienceOn
25 Hermansson U, Linde Forsberg G. Freezing of stored, chilled dog spermatozoa, Theriogenology 2006; 65: 584-593.   DOI   ScienceOn
26 Ricci C, Perticarari S, Fragonas E, Giolo E, Canova S, Pozzobon C, Guaschino S, Presani G. Apoptosis in human sperm: its correlation with semen quality and the presence of leukocytes. Hum Reprod 2002; 17: 2665-2672.   DOI   ScienceOn