Diverse developmental velocities of blastocyst development emerge as early as a few hours after insemination: morphokinetic analysis of fertilisation in days 4-7 blastocysts

Purpose: The time at which embryo developmental velocity variations occur remains unknown. Therefore, we analysed the morphokinetic trajectory of human early fertilisation from the extrusion of the second polar body (PB2) to pronuclear breakdown (PNBD) and retrospectively compared its alterations during the fertilisation of day 4–7 blastocysts. Methods: Patients (n = 905) who underwent oocyte retrieval and blastocyst expansion between October 2019 and December 2020 were included. This study involved time-lapse technology monitoring of day 4 (n = 29), 5 (n = 1181), 6 (n = 244), and 7 (n = 12) blastocysts generated over 1009 intracytoplasmic sperm injection cycles. Morphokinetics during fertilisation were compared among the day 4–7 blastocyst groups. Results: The timing of fertilisation events in the cytoplasm and pronuclei (PN) differed among the day 4–7 blastocyst groups from the outset. PB2 extrusion occurred significantly earlier in day 4 blastocysts than in day 5 blastocysts. The duration of the cytoplasmic halo was shortened in day 4 blastocysts and prolonged in day 6 blastocysts compared with that in day 5 blastocysts. Paternal PN appeared earlier in day 4 blastocysts than in day 5 blastocysts, whereas their appearance was delayed in day 6 blastocysts. In paternal and maternal PN, the time interval from PN appearance to PNBD was shortened in day 4 blastocysts but was prolonged in day 6 and 7 blastocysts. Conclusions: Embryos that reached the blastocyst stage between days 4 and 7 displayed different morphokinetic speeds throughout fertilisation and as early as PB2 extrusion. Aberrant cytoplasmic granules, organelles, and PN rearrangement may contribute to these differences.