Survival and developmental potential of stored human early cleavage stage embryos.

Human early cleavage stage embryos which survive cryopreservation and thawing fully intact demonstrate similar developmental potential to equivalent non frozen embryos when returned to the in vivo environment, whereas blastomere loss is directly related to the loss of potential for subsequent implantation in thawed embryos. This suggests that blastomere lysis during freezing and thawing does not occur preferentially in non viable blastomeres. Prefreeze growth rate rather than prefreeze blastomere number per se correlates with the developmental potential of stored embryos. When blastomere loss occurs as a consequence of cryopreservation, development of thawed early cleavage stage embryos to the blastocyst stage in vitro is impaired and the resultant blastocysts have a reduced total cell content. Blastomere loss is more prevalent in embryos which have been biopsied for preimplantation genetic diagnosis but this increased sensitivity can be circumvented by modification of the standard cryopreservation protocol.

The role of insulin-like growth factor II and its receptor in mouse preimplantation development.

Insulin-like growth factor II (IGF-II) and its receptor, the IGF-II/mannose-6-phosphate (IGF-II/M6P) receptor, are first expressed from the zygotic genome at the two-cell stage of mouse development. However, their role is not clearly defined. Insulin-like growth factor II is believed to mediate growth through the heterologous type 1 IGF and insulin receptors, whereas the IGF-II/M6P receptor is believed to act as a negative regulator of somatic growth by limiting the availability of excess levels of IGF-II. These studies demonstrate that IGF-II does have a role in growth regulation in the early embryo through the IGF-II/M6P receptor. Insulin-like growth factor II stimulated cleavage rate in two-cell embryos in vitro. Moreover, this receptor is required for the glycaemic response of two-cell embryos to IGF-II and for normal progression of early embryos to the blastocyst stage. Improved development of embryos in crowded culture supports the concept of an endogenous embryonic paracrine activity that enhances cell proliferation. These responses indicate that the IGF-II/M6P receptor is functional and likely to participate in such a regulatory circuit. The functional role of IGF-II and its receptor is discussed with reference to regulation of early development.

The influence of prefreeze growth rate and blastomere number on cryosurvival and subsequent implantation of human embryos.

PURPOSE: To determine whether the relatively low implantation rate of cryopreserved Day 2 embryos with only 2 blastomeres can be increased as a consequence of increasing their blastomere content by extending the prefreeze culture time. METHODS: Of a total of 3480 Day 2 embryos studied, 1921 (55.2%) had reached the 4-cell stage by 40 h postinsemination (FAST) and were transferred or cryopreserved. The remaining embryos that underwent subsequent cell division by 46 h (INTERMEDIATE; 18.3% of total) or 66 h (SLOW; 20.3% of total) were also cryopreserved whereas the 6.2% that remained arrested at 66 h were discarded. Thawed embryos from each category were assessed for survival, post-thaw cleavage, and implantation. RESULTS: The proportion of thawed embryos that survived, the proportion of surviving embryos that underwent post-thaw cleavage, and the implantation rate of transferred embryos were all reduced in the slower growing cryopreserved embryos. CONCLUSIONS: The growth rate, and not the number of blastomeres per se, is a critical factor in predicting the developmental potential of cryopreserved embryos.

The developmental potential of cryopreserved human embryos.

Using rigorously matched non-frozen controls we have shown that cryopreservation does not alter the implantation potential of early cleavage stage (day 2) human embryos if no blastomere loss occurs. Thawed intact 4-cell embryos have a significantly higher implantation (fetal heart) rate (16.9%) than similar 2-cell embryos (7.2%). This difference is not due to blastomere number per se since increasing the cell number in frozen embryos by allowing an extended period in culture prior to freezing does not alter their intrinsic developmental potential. Blastomere loss, which occurred in almost half of all thawed embryos, is directly related to a reduction in developmental potential. We estimate that approximately 30% of the expected fresh embryo implantations are lost as a consequence of cryopreservation. Both preimplantation and peri-implantation losses may contribute to this outcome.