Characterization of the first cell cycle in human zygotes: implications for cryopreservation

To study the temporal pattern of deoxyribonucleic acid (DNA) synthesis leading up to the first mitotic division in human one-cell stage zygotes.

In vitro fertilization program of a university hospital.

Couples donating spare embryos before the existence of an embryo freezing program.

Incorporation of 3H-thymidine was examined between 9 to 27 hours after insemination in 72 untransferred human zygotes containing two pronuclei. Microscopic observations on an additional 978 transferred zygotes extended the 3H-thymidine incorporation data.

The first thymidine incorporation was seen 9 to 10 hours after insemination, and the heaviest incorporation occurred between 11 and 13 hours. Subsequently, thymidine labeling declined, and chromosomal condensation and cell division first occurred approximately 19 to 20 hours. At 20 hours after insemination, 89% of the zygotes had two visible pronuclei (PN). In contrast, by 24 hours, 41% had no visible PN, whereas 5% had cleaved to the two-cell stage. By 27 hours, 38% had cleaved to the two-cell stage, and only 25% still had two visible PN.

These results suggest that the DNA-synthetic, S-phase of the human zygote is initiated by approximately 9 to 10 hours after insemination and is completed approximately 3 to 5 hours later. The duration of G2 phase and mitosis is in the range of 4 to 6 and 3 to 3.5 hours, respectively. Because zygotes may be particularly susceptible to damage during the S phase of the cell cycle, these findings suggest that the optimal time for freezing of human zygotes may be approximately 20 to 22 hours after insemination when the majority of zygotes should have entered the G2 phase, before pronuclear dissolution and chromosome condensation.