Cryopreservation of human embryos by vitrification or slow freezing: a systematic review and meta-analysis

Purpose The objective of this study was to investigate the effects of vitrification on the preimplantation developmental competence of mouse 2-cell, 4-cell and 8-cell stage embryos. Methods Mouse 2-cell, 4-cell and 8-cell stage embryos were cryopreserved using the cryotop vitrification method and subsequently warmed on a later date. The embryos were then assessed by their morphology, blastocyst formation and hatching rates. Additionally, trophectoderm (TE) and inner cell mass (ICM) cell numbers were compared in hatched blastocysts from the control and experimental groups. Results Vitrified embryos at the 2-cell, 4-cell and 8-cell stages appeared morphologically normal after warming. The overall survival rate of vitrified embryos at various stages after warming was 96.7% and there were no significant differences among 2-cell stage (96.0%), 4-cell stage (96.8%) and 8-cell stage (97.1%) embryos (P>0.05). The blastocyst formation rate (69.4%) and hatching rate (52.6%) of vitrified 2-cell embryos were significantly lower than that from the control group and vitrified 8-cell embryos (P<0.05). In the vitrified 4-cell embryo group, the blastocyst formation rate (90.3%) was similar to the 8-cell group (91.2%), but the hatching rate (60.0%) was significantly lower than that of the non-vitrified control ( 84.1%) and vitrified 8-cell embryo (78.4%) groups (P<0.05). When further development to the fully hatched blastocyst stage was compared, hatched blastocysts derived from vitrified 2-cell, 4-cell and 8-cell embryos had significantly lower cell counts both in the ICM and TE, as compared to fresh blastocysts (P<0.05). Among the vitrified 2-cell, 4-cell and 8-cell embryo groups, there were no significant differences in the cell counts of ICM and TE (P>0.05). Conclusions Although cryotop vitrification was suitable for the cryopreservation of mouse embryos from the 2-cell stage, 4-cell stage and 8-cell stage without significant loss of survival, vitrification had an adverse effect on the development of 2-cell embryos. Mouse embryos at the 8-cell stage had the best tolerance for vitrification and would yield the highest level of post-vitrification developmental competence among early cleavage stage embryos. Nevertheless, it is unclear how these findings can be extrapolated to human embryos.

Section: Discussionmentioning

confidence: 99%

Vitrification of mouse embryos at 2-cell, 4-cell and 8-cell stages by cryotop method

Zhang

Cui

Ling

et al. 2009

“…This procedure is known as an open vitrification system. This idea was initially proposed for freezing Drosophila embryos [6], and drastic improvement in viability has since been shown in both animal studies [7][8][9][10] and clinical reports [11,12]. However, there are some potential drawbacks of the open vitrification system, such as the sterility of liquid nitrogen and the risk of cross-contamination during long-term storage [13,14].…”

Section: Introductionmentioning

confidence: 99%

Neonatal outcomes after the implantation of human embryos vitrified using a closed-system device

Iwahata

Hashimoto

Inoue³

et al. 2015

Purpose Closed vitrification poses a risk of adversely affecting embryo development, while it may minimize the risk of contamination. We assessed the effects of closed-system human embryo vitrification on fetal development after implantation, neonatal outcome, and clinical safety. Methods This was a retrospective cohort study conducted at a private fertility clinic. A total of 875 vitrified-warmed blastocysts that were single-transferred under hormone-replacement cycles between November 2011 and December 2013 were randomly divided into two groups (closed vitrification, n 313; open vitrification, n 562) after receiving the patients' consent forms. Developmental competence after implantation, including gestational age, birth weight, sex, Apgar score, and anomalies of newborns, after the transfer of blastocysts vitrified by closing vitrification was compared with that obtained in the case of open vitrification. Results There were no significant differences between the use of closed and open vitrification systems in embryo development after implantation, gestational age, birth weight, sex ratio, Apgar score, and congenital anomalies of newborns. Conclusion Human embryos can be vitrified using a closed vitrification system without impairment of neonatal development.

“…This idea was initially proposed for freezing Drosophila embryos (Mazur et al 1992). Animal data [11,15,22,31] and recent analyses of clinical reports [18,19] have revealed the benefits of vitrification such as low rates of cellular damage. However, the transition from freezing to vitrification is proceeding very slowly due to concerns regarding the sterility of liquid nitrogen and the risk of cross-contamination during long-term storage [2,3].…”

Section: Introductionmentioning

confidence: 99%

A closed system supports the developmental competence of human embryos after vitrification

Hashimoto

Amo

Hama

et al. 2013

Purpose Closed-system vitrification may enable the risk of contamination to be minimised. We performed three studies to compare the developmental competence of human embryos vitrified using either a closed vitrification system (CVS; Rapid-i®) or an open vitrification system (OVS; Cryo-top®). Methods The first study was performed in vitro using 66 zygotes previously vitrified at pronuclear stage. These were warmed and randomised 1:1 to revitrification using either the OVS or the CVS. After re-warming, embryo development and blastocyst cell number were assessed. For the second study, also performed in vitro, 60 vitrified-warmed blastocysts were randomised 1:1:1 into three groups (OVS or CVS revitrification, or no revitrification). The proportion of dead cells was assessed by staining. The third study was performed in vivo, using 263 high-grade blastocysts randomly assigned to vitrification using either the CVS (n=100) or the OVS (n=163). After warming, single blastocyst transfer was performed. Results There were no differences between the CVS and the OVS in survival rate (100 % vs. 97 %), blastulation rate (96 h: 50 % vs. 50 %; 120 h: 68 % vs. 56 %), proportion of good blastocysts (96 h: 32 % vs. 22 %, 120 h: 47 % vs. 41 %), or mean number of cells (137 vs. 138). The proportion of dead cells in blastocysts re-vitrified by CVS (31 %) was similar to that for OVS (38 %) and non-revitrification (32 %). In vivo, the implantation rate for blastocysts vitrified using the CVS (54 %) was similar to that with the OVS (53 %).Conclusion Our studies consistently indicate that human embryos may be vitrified using a CVS without impairment of developmental competence.