Embryo morphokinetics derived from fresh and vitrified bovine oocytes predict blastocyst development and nuclear abnormalities

Abstract

Embryo development is a dynamic process and critical stages may go unnoticed with the use of traditional morphologic assessments, especially the timing of embryonic divisions and aberrant zygotic cleavage patterns. Bovine embryo development is impaired after oocyte vitrification, but little is known about the underlying morphokinetic behavior. Here, bovine zygotes from fresh (n = 708) and vitrified oocytes (n = 182) were monitored by time-lapse imaging and the timing and nature of early blastomere divisions were modeled to find associations with blastocyst development at day 8. The predictive potential of morphokinetic parameters was analyzed by logistic regression and receiver operating characteristic curve analysis to determine optimal cut-off values. Lag-phase was highly correlated with embryo development. Remarkably, 100% of zygotes that reached the blastocyst stage showed a lag-phase. Fast first cleavage increased the chance of blastocyst development to 30% with a cut-off of 32 h and 22 min. Aberrant zygotic cleavage events, including multipolar division, unequal blastomere sizes, and membrane ruffling resulted in decreased blastocyst development. Multipolar division leads to uneven blastomeres, which was associated with anuclear and multinuclear blastomeres, indicating genome segregation errors. Moreover, we described for the first time morphokinetics of embryos derived from vitrified bovine oocytes. Vitrification severely affected blastocyst development, although lower cryoprotectant concentration in equilibration solutions seems to be less detrimental for embryo yield. Impaired development was linked to slow cleavages, lower lag-phase incidence, and increased early embryonic arrest. Typically, less than 15% of the embryos produced from vitrified oocytes reached more than eight cells. Interestingly, the rate of abnormal first cleavage events was not affected by oocyte vitrification. In conclusion, time to first cleavage, the presence of a lag-phase, and the absence of aberrant zygotic cleavage were the best predictors of bovine blastocyst development for both fresh and vitrified oocytes.