The combination of calcium ionophore A23187 and GM-CSF can safely salvage aged human unfertilized oocytes after ICSI.

Abstract

Artificial oocyte activation using calcium ionophores and enhancement of embryonic developmental potential by the granulocyte-macrophage colony-stimulating factor (GM-CSF) have already been reported. In this study, we evaluated the synergistic effect of these two methods on aged human unfertilized oocytes after intracytoplasmic sperm injection (ICSI). Then, we cultured the resulting embryos to the blastocyst stage and screened them for chromosomal abnormalities, to assess the safety of this protocol. Aged human oocytes deemed unfertilized after ICSI were activated, either by briefly applying the calcium ionophore A23187 alone (group A) or by briefly applying the ionophore and then supplementing the culture medium with recombinant human GM-CSF (rhGM-CSF) (group B). Next, the development was monitored in a time-lapse incubator system, and ploidy was analyzed by array comparative genomic hybridization (aCGH), after whole embryo biopsy and whole genome amplification. Differences between oocytes and resulting embryos in both groups were evaluated statistically. Oocytes unfertilized after ICSI can be activated with the calcium ionophore A23187 to show two pronuclei and two polar bodies. Addition of rhGM-CSF in the culture medium of A23187-activated oocytes enhances their cleaving and blastulation potential and results in more euploid blastocysts compared to the culture medium alone. This study shows that activating post-ICSI aged human unfertilized oocytes with a combination of a calcium ionophore and a cytokine can produce good-morphology euploid blastocysts.