Effect of Cryopreservation Day on Pregnancy Outcomes in Frozen-thawed Blastocyst Transfer

동결 해동한 포배 이식에 있어서 동결시기가 임신결과에 미치는 영향

  • Published : 2010.03.31

Abstract

Objectives: Likewise fresh cycle, it is also important to select right blastocysts for transfer in purpose of improving the pregnancy and implantation rates in frozen-thawed embryo transfer (ET) cycles. To investigate the relationship between the developmental velocity at the time of cryopreservation and pregnancy rates, we compared pregnancy rates between the day 5 cryopreservation group and the day 6 cryopreservation group. Methods: Transfers of frozen-thawed blastocysts which had been cryopreserved by vitrification on day 5 or day 6 were performed between January 2006 and June 2007. Ethylene glycol, DMSO, and pull and cut straws were used for vitrification and artificial shrinkage was done in expanded blastocysts. Thawing was performed on the day before transfer and thawed blastocysts were cultured in for 15~18 hrs in Quinn's blastocyct media. Blastocyst survival was assessed before transfer and post-thaw survival was defined as >50% of cells remaining intact and blastocoele re-expansion by the time of transfer. Results: Transfers of thawed blastocyst had been cryopreserved on day 5 were 52 cycles and 41 transfer cycles were cryopreserved on day 6. Patient characteristics, the number of transferred embryos and the survival rate of thawed blastocysts were not different in each cryopreservation day. But the biochemical pregnancy, clinical pregnancy, ongoing pregnancy, and implantation rate were significantly high in transfer of frozen-thawed blastocyst which were cryopreserved on day 5. Conclusions: The clinical pregnancy and implantation rate of day-5 blastocyst showed significantly higher than those of day-6 blastocyst in frozen-ET cycles. This result indicated that developmental rate of blastocyst at cryopreservation time in frozen-thawed cycle is discriminative marker of pregnancy outcome as like in fresh cycle.

목 적: 배양 환경과 동결 기술이 발달함에 따라 동결 포배의 해동-이식의 빈도가 증가하고 있으며, 신선 주기와 마찬가지로 해동-이식 주기에서도 질 좋은 배아를 선별하는 것은 임신 성공 여부를 결정하는 아주 중요한 과제이다. 본 연구는 동결 당시 포배로의 발달 속도가 임신 결과에 미치는 영향을 알아보기 위하여, 수정 후 5일 및 6일째 동결한 포배의 해동-이식 후 임신율을 비교 분석하였다. 연구방법: 2006년 1월부터 12월까지 5일째 또는 6일째 동결한 포배를 해동하여 2007년 6월까지 융해 이식한 87명, 93주기를 대상으로 하였다. 동결법은 ethylene glycol과 DMSO를 이용한 유리화 동결법을 이용하였으며, 팽창 포배는 인위적인 수축을 시행 후 동결하였다. 해동 과정은 이식 전날 시행하여 15~18시간 배양액에서 배양 후 재팽창 여부를 확인하였다. 결 과: 5일째 동결한 포배를 해동-이식한 52주기와 6일째 동결한 포배를 해동-이식한 41주기에서 환자의 나이, 이식한 배아의 수, 해동 배아의 생존율 등 임신 결과에 영향을 미칠만한 요인들의 차이는 없었다. 그러나 생화학적 임신율, 임상적 임신율, 진행 임신율, 착상율 등은 5일째 동결한 포배를 해동-이식한 주기에서 높게 나타났다. 결 론: 5일째 동결한 포배를 해동-이식했을 때의 임신율은 6일째 동결한 포배를 해동-이식했을 때의 임신율보다 2배 이상 높았으며, 이는 신선 주기와 마찬가지로 해동-이식 주기에서도 동결 전 배아의 발달 속도의 차이를 임신 성공 예측의 중요한 지표로 사용할 수 있음을 시사한다.

Keywords

References

  1. Gardner DK, Vella P, Lane M, Wagley L, Schlenker T, Schoolcraft WB. Culture and transfer of human blastocysts increases implantation rates and reduces the need for multiple embryo transfers. Fertil Steril 1998; 69: 84-8. https://doi.org/10.1016/S0015-0282(97)00438-X
  2. Yokota Y, Sato S, Yokota M, Ishikawa Y, Makita M, Asada T, Araki Y. Successful pregnancy following blastocyst vitrification. Hum Reprod 2000; 15: 1802-3. https://doi.org/10.1093/humrep/15.8.1802
  3. Mukaida T, Nakamura S, Tomiyama T, Wada S, Kasai M, Takahashi K. Successful birth after transfer of vitrified human blastocyst with use of a cryo-loop containerless technique. Fertil Steril 2001; 76: 618-20. https://doi.org/10.1016/S0015-0282(01)01968-9
  4. Cho HJ, Son WY, Yoon SH, Lee SW, Lim JH. An improved protocol for dilution of cryoprotectant from vitrified human blastosyst. Hum Reprod 2002; 17: 2419-22. https://doi.org/10.1093/humrep/17.9.2419
  5. Son WY, Yoon SH, Lee SM, Lim JH. Pregnancy outcome following transfer of human blastocysts vitrified on electron microscopy grids after induced collapse of the blastocele. Hum Reprod 2003; 18: 137-9. https://doi.org/10.1093/humrep/deg029
  6. Alves da Motta EL, Alergretti JR, Baracat EC, Olive D, Serafini PC. High implantation and pregnancy rates with transfer of human blastocysts developed in preimplantation stage one and blastocyst media. Fetil Steril 1998; 70: 659-63. https://doi.org/10.1016/S0015-0282(98)00263-5
  7. Khorram O, Shapiro SS, Jones JM. Transfer of nonassisted hatched and hatching human blastocysts after in vitro fertilization. Fertil Steril 2000; 74: 163-5. https://doi.org/10.1016/S0015-0282(00)00567-7
  8. Shapiro BS, Richter KS, Harris DC, Daneshmand ST. A comparison of day 5 and day 6 blastocyst transfers. Fertili Steril 2001; 75: 1126-30. https://doi.org/10.1016/S0015-0282(01)01771-X
  9. Shapiro BS, Harris DC, Richter KS. Predictive value of 72-hour blastomere cell number on blastocyst development and success of subsequent transfer based on the degree of blastocyst development. Fertili Steril 2000; 73: 582-6. https://doi.org/10.1016/S0015-0282(99)00586-5
  10. Huisman GJ, Fauser BC, Eijkemans MJ, Pieters MH. Implantation rates after in vitro fertilization and transfer of a maximum of two embryos that have undergone three to five days of culture. Fertili Steril 2000; 73: 117-22. https://doi.org/10.1016/S0015-0282(99)00458-6
  11. Nagy ZP, Liu J, Joris G, Bocken G, Desmet B, Vankelecom A, et al. The influence of the site of sperm deposition and mode of oolemma breakage at intracytoplasmic sperm injection on fertilization and embryo development rates. Hum Reprod 1995; 10: 3171-777. https://doi.org/10.1093/oxfordjournals.humrep.a135881
  12. Balaban B, Urman B, Sertac A, Alatas C, Alsoy S, Mercan R. Blastocyst quality affects the success of blastocyst-stage embryo transfer. Fertili Steril 2000; 74: 282-7. https://doi.org/10.1016/S0015-0282(00)00645-2
  13. Scott L, Alvero R, Leondires M, Miller B. The morphology of human pronuclear embryos is positively related to blastocyst development and implantation. Hum Reprod 2000; 15: 2394 -403 https://doi.org/10.1093/humrep/15.11.2394
  14. Lane M, Gardner D. Differential regulation of mouse embryo development and viability by amino acids. J Reprod Fertil 1997; 109: 153-64. https://doi.org/10.1530/jrf.0.1090153
  15. Dokras A, Sargent IL, Barlow DH. Human blastocyst grading: an indicator of developmental potential? Hum Reprod 1993;8: 2119-27. https://doi.org/10.1093/oxfordjournals.humrep.a137993
  16. Gardner DK, Lane M, Stevens J, Schlenker T, Schoolcraft WB. Blastocyst score affects implantation and pregnancy outcome: towards a single blastocyst transfer. Fertil Steril 2000; 73: 1155-8. https://doi.org/10.1016/S0015-0282(00)00518-5
  17. Balaban B, Yakin K, Urman B. Randomized comparison of two different blastocyst grading systems. Fertil Steril 2006; 85: 559-63. https://doi.org/10.1016/j.fertnstert.2005.11.013
  18. Renard J, Philippon A, Menezo Y. In-vitro uptake of glucose by bovine blastocysts. J Reprod Fertil 1980; 58: 161-4. https://doi.org/10.1530/jrf.0.0580161
  19. Gardner DK, Leese HJ, Assessment of embryo viability prior to transfer by the noninvasive measurement of glucose uptake. J Exp Zool 1987; 242: 103-5. https://doi.org/10.1002/jez.1402420115
  20. Shu Y, Watt J, Gebhardt J, Dasig J, Appling J, Behr B. The value of fast blastocoele re-expansion in the selection of a viable thawed blastocyst for transfer. Fertil Steril 2009; 91: 401-6. https://doi.org/10.1016/j.fertnstert.2007.11.083
  21. Desai N, Goldfarb J. Examination of frozen cycles with replacement of a single thawed blastocyst. Reprod Biomed Online 2005; 11: 349-54. https://doi.org/10.1016/S1472-6483(10)60843-7