Yu Lan, Jia Chanwei, Lan Yonglian, Song Rui, Zhou Liying, Li Ying, Liang Yu, Wang Shuyu
a Department of Reproductive Medicine , Beijing Obstetrics and Gynecology Hospital, Capital Medical University , Beijing , China.
Syst Biol Reprod Med. 2017 Oct;63(5):285-293. doi: 10.1080/19396368.2017.1362060. Epub 2017 Aug 10.
The aim of this study was to identify the parameters that are related to intactness and developmental potential of a day 3 embryo after warming to improve the selection criteria used to cryopreserve and transfer embryos. We also sought to compare slow freezing and vitrification methods of cryopreservation and to evaluate the viability of non-intact embryos. Embryos warmed following slow freezing (n=220) or vitrification (n=522) were divided into 3 groups according to the proportion of surviving blastomeres (I<50%; II=50-99%; and III=100%). The developmental potential of embryos, including the mitosis resumption rate, blastocyst formation rate, and formation rate of grade A blastocysts (i.e., fully expanded blastocysts with an inner cell mass and grade A or B trophectoderm) were retrospectively assessed in embryos. Cleavage-stage embryos with <50% blastomere survival were analyzed using next-generation sequencing (NGS). Logistic regression analysis showed that vitrification and grade 1 were independent predictive factors of embryo intactness and developmental potential (all p<0.05). On day 3, embryos with 4-6 cells or blastomere damage had lower developmental potential than those with 7-9 cells or intact blastomeres (all p<0.05). NGS results showed that the chromosomal status was completely normal in 8 embryos that developed into expanded blastocysts, whereas 4 out of 5 embryos in which development was arrested were abnormal. The results of this study suggest that vitrification is a better choice than slow freezing for embryo cryopreservation. Embryos showing poor quality (fragmentation >30% and/or a non-stage-specific cell size) and lower cell numbers (4-6 cells) on day 3 should be cultured to the blastocyst stage and then vitrified if they develop into good-quality blastocysts. The developmental potential of non-intact embryos is lower than that of intact embryos; however, after they are cultured to the fully expanded blastocyst stage, embryos with <50% blastomere survival appear to be better candidates for transfer. Abbreviations ART: assisted reproductive technology; grade A blastocyst: fully expanded blastocyst with an inner cell mass and grade A or B trophectoderm; NGS: next-generation sequencing; IVF: in vitro fertilization; ICSI: intracytoplasmic sperm injection; FET: frozen-thawed embryo transfer.
本研究的目的是确定与解冻后第3天胚胎的完整性和发育潜能相关的参数，以改进用于冷冻保存和移植胚胎的选择标准。我们还试图比较冷冻保存的慢速冷冻法和玻璃化法，并评估非完整胚胎的活力。将慢速冷冻（n = 220）或玻璃化（n = 522）后解冻的胚胎根据存活卵裂球的比例分为3组（I<50%；II = 50 - 99%；III = 100%）。回顾性评估胚胎的发育潜能，包括有丝分裂恢复率、囊胚形成率和A级囊胚（即完全扩张的囊胚，具有内细胞团和A级或B级滋养外胚层）的形成率。对卵裂球存活率<50%的卵裂期胚胎进行二代测序（NGS）分析。逻辑回归分析表明，玻璃化和1级是胚胎完整性和发育潜能的独立预测因素（所有p<0.05）。在第3天，4 - 6细胞或卵裂球受损的胚胎比7 - 9细胞或卵裂球完整的胚胎发育潜能更低（所有p<0.05）。NGS结果显示，8个发育为扩张囊胚的胚胎染色体状态完全正常，而5个发育停滞的胚胎中有4个异常。本研究结果表明，对于胚胎冷冻保存，玻璃化是比慢速冷冻更好的选择。第3天显示质量较差（碎片>30%和/或非特定阶段的细胞大小）且细胞数量较少（4 - 6细胞）的胚胎应培养至囊胚期，如果发育为优质囊胚则进行玻璃化冷冻。非完整胚胎的发育潜能低于完整胚胎；然而，在培养至完全扩张的囊胚期后，卵裂球存活率<50%的胚胎似乎是更好的移植候选对象。缩写词：ART：辅助生殖技术；A级囊胚：完全扩张的囊胚，具有内细胞团和A级或B级滋养外胚层；NGS：二代测序；IVF：体外受精；ICSI：卵胞浆内单精子注射；FET：冻融胚胎移植
