Reproductive and Developmental Biology

  • 제35권2호
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  • Pages.137-141
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  • 2011
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  • 1738-2432(pISSN)
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  • 2288-0151(eISSN)
한국동물번식학회 (The Korean Society of Animal Reproduction)
권호 표지

Study on Development of Canine Oocytes Treated by In Vitro Fertilization and ICSI

  • Park, Ji-Hoon (College of Veterinary Medicine, Chungnam National University) ;
  • Chung, Young-Ho (Dept. of Companion Animal and Animal Resourses Science, Joongbu University) ;
  • Kim, Sang-Keun (College of Veterinary Medicine, Chungnam National University)
  • 투고 : 2011.05.07
  • 심사 : 2011.05.30
  • 발행 : 2011.06.30
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초록

These study was to investigate the in vitro fertilization and viability of fresh and vitrified oocytes. Also, the developmental capacity of IVF and intracytoplasmic sperm injection (ICSI) oocytes were investigated. Then vitrification was performed with the use of 20% ethylene glycol + 20% DMSO + 0.5 M sucrose + 10% FCS + TCM-199 medium. Vitrification immature oocytes are cultured in vitrification solution for 10 min afterwards transferred to expose at room temperature for 5 min. and transferred to the ice water for 5 min. The oocytes were sealed in a 1.0 mm straw and placed in a $LN_2$ container. Frozen oocytes were rapidly thawed in a water bath at $30{\sim}35^{\circ}C$, and then placed in TCM-199 medium containing 0.5 M sucrose for 5 min each, respectively, at $38^{\circ}C$. After being washed for 2~3 times, using fresh medium the oocytes were cultured in TCM-l99 medium supplemented with 5% FCS at $38^{\circ}C$ in 5% $CO_2$ and air. The normal morphology of fresh and vitrified-thawed oocytes were $87.1{\pm}2.1%$ and $54.8{\pm}2.5%$, respectively. The viability rates of fresh and vitrified-thawed oocytes were $70.0{\pm}2.2%$ and $41.9{\pm}2.6%$, respectively. Viability rates of vitrified-thawed oocytes were lower than that of fresh follicular oocytes (p<0.05). The in vitro maturation rates of fresh and vitrified oocytes were $45.1{\pm}3.6%$ and $28.9{\pm}4.4%$, respectively. The IVF rates of fresh follicular and vitrified-thawed oocytes were 34.00.2% and $20.2{\pm}2.6%$, respectively. The in vitro maturation and fertilization rates of vitrified-thawed oocytes were lower than those of the fresh follicular oocytes (p<0.05). A total of 350 oocytes were fixed and stained after co-incubation with spermatozoa, of which 88 had identifiable nuclear material. After IVF for 20 hrs, $25.1{\pm}3.4%$ of the oocytes found to have been penetrated by spermatozoas. Oocytes were fixed and stained after ICSI, and 105 oocytes contained identifiable nuclear material. After IVF and ICSI for 20 hrs, $34.3{\pm}3.4%$ and $59.0{\pm}2.0%$ of the oocytes were found to have been penetrated by spermatozoas. The developmental rates upon ICSI were significantly higher than those of the IVF method (p<0.05).

키워드

참고문헌

  1. Abe Y, Lee DS, Kim SK, Suzuki H (2008): Vitrification of canine oocytes. J Mamm Ova Res 25:32-36. https://doi.org/10.1274/jmor.25.32
  2. Bedford SJ, Kurokawa M, Hinrichs K, Fissore RA (2003): Intracellular calcium oscillations and activation in horse oocytes injected with stallion sperm extracts or spermatozoa. Reproduction 126:489-499. https://doi.org/10.1530/rep.0.1260489
  3. Candy CJ, Wood M, Whittingham DG, Merriman JA, Choudhury (1994): Cryopreservation of immature mouse oocytes. Hum Reprod 9:1738-1742.
  4. Cuello C, Sntonia MG, Parrilla I, TomeI J, Vazquez JM, Roca J, Berthelot F, Martinat-Botte F, Martinez EA (2004): Vitrification of porcine embryos at various development stages using different ultra-rapid cooling procedures. Theriogenology 62:353-361. https://doi.org/10.1016/j.theriogenology.2003.10.007
  5. Fujihira T, Kishida R, Fukui Y (2004): Developmental capacity of vitrified immature porcine oocytes following ICSI: Effects of cytochalasin B and cryoprotectants. Cryobiology 49:286-290. https://doi.org/10.1016/j.cryobiol.2004.08.004
  6. Hewitt DA, England GCW (1999): Synthetic oviductal fluid and oviductal cell coculture for canine oo-cytes maturation in vitro. Anim Reprod Sci 55(1): 63-75. https://doi.org/10.1016/S0378-4320(98)00162-6
  7. Isachenko V, Soler C, lsachenko E, Perez-Sanchez F, Grishchenko V (1998): Vitrification of immature porcine oocytes: Effects of lipid droplets, temperature, cytoskeleton, and addition and removal of cryoprotectant. Cryobiology 36:250-253. https://doi.org/10.1006/cryo.1998.2079
  8. Kasai M, Komi JH, Takakamo A, Tssnoda H, Sakurai T, Machida T (1990): A simple method for mouse embryo cryopreservation in a low toxicity vitrification solution, without appreciable loss of viability. J Reprod Fertil 89:91-97. https://doi.org/10.1530/jrf.0.0890091
  9. Leibo SP, Oda K (1993): High survival of mouse zygote and embryos cooled rapidly or slowly in ethylene glycol plus polyvinyl pyrrolidone. Cryo-Letters 14:133-144.
  10. Otoi T, Shin T, Kraemer D, Westhusin ME (2004): Influence of maturation culture period on the development of canine oocytes after in vitro maturation and fertilization. Reprod Nutr Dev 44:631-637. https://doi.org/10.1051/rnd:2004065
  11. Otoi T, Shimizu R, Naoi H, Wongsrikeao P, Agung B, Taniguchi M (2005): Meiotic competence of canine oocytes embedded in collagen gel. Reprod DO-mest Anim 41:17-21.
  12. Rall WF, Fahy GM (1985): Ice-free cryopreservation of mouse embryos at $-196{^{\circ}C}$ by vitrification. Nature 313:573-575. https://doi.org/10.1038/313573a0
  13. Reyes M De, Carrion R, Barros C (2006): In vitro fertilization of in vitro matured canine oocytes using frozen-othawed dog semen. Theriogenology 66:1682-1684. https://doi.org/10.1016/j.theriogenology.2006.02.002
  14. Renard JP, Nguyen BX, Garnier V (1984): Two-step freezing of two-cell rabbit embryos after partial dehydration at room temperature. J Reprod Fert 71:573-580. https://doi.org/10.1530/jrf.0.0710573
  15. Robinski B, Arav A, Devires AL (1991): Cryopreservation of oocytes using directional cooling and antifreeze glycoproteins. Cryo-Lettters 12:93-106.
  16. Rodrigues BA, Carboneiro dos Santos L, Rodrigues JL (2004): Embryonic development of in vitro matured and in vitro fertilized dog oocytes. Mol Reprod Dev 67:215-223. https://doi.org/10.1002/mrd.10394
  17. Schmidt M, Hyttle P, Grece T, Avery B (1993: Ultrastructure of frozen thawed bovine in vitro matured oocytes. Theriogenology 39:304. https://doi.org/10.1016/0093-691X(93)90159-3
  18. Songsasen N, Yu I, Leibo SP (2002): Nuclear maturation of canine oocytes cultured in protein-free media. Mol Reprod Dev 62:407-415. https://doi.org/10.1002/mrd.10130
  19. Songsasen N, Wildt DE (2005): Size of the donor follicle, but not stage of reproductive cycle or seasonality, influences meiotic competency of selected domestic dog oocytes. Mol Reprod Dev 72:113-119. https://doi.org/10.1002/mrd.20330
  20. Suzuki T, Nishikata Y (1992): Fertilization and cleavage of frozen thawed bovine oocytes by one step dilution methos in vitro. Theriogenology 37:306. https://doi.org/10.1016/0093-691X(92)90375-2
  21. Toth TL, Jones HW, Baka SC, Muasher S, Veeck LL, Lanzendorf SE (1994): Fertilization and in vivo development of cryopreserved human prophase I oo-cytes. Fertil Steril 61:891-894.
  22. Vajta G, Holm P, Kuwayama M, Booth PJ, Jacobsen A, Greve T, Callesen H (1998): Open pulled straw vitrification: A new way to reduce cryoinjuries of bovine ova and embryos. Mol Reprod Dev 51:53-58. https://doi.org/10.1002/(SICI)1098-2795(199809)51:1<53::AID-MRD6>3.0.CO;2-V
  23. van Blerkom J (1989): Maturation at high frequency of germinal vesicle-stage mouse oocyte after cryopreservation: Alterations in cytoplasmic, nuclear, nucleolar and chromosomal structure and organization associated with vitrification. Human Reprod 4:883-898.
  24. Yamada S, Shimazu Y, Kawaji H, Nakazawa M, Naito K, Toyoda Y (1992): Maturation, fertilization, and development of dog oocytes in vitro. BioI Reprod 46:853-858. https://doi.org/10.1095/biolreprod46.5.853