Diagnosing and preventing inherited disease: An analysis of multinucleated blastomere formation in human embryos

Human embryos were disaggregated into component blastomeres 42–72 h after insemination. The blastomeres were scored for the number of nuclei present and blastomeres of known nuclear morphology were returned to individual culture drops for 16–20 h, after which they were scored for cleavage and nuclear morphology. In all, 48% of mononucleated blastomeres cleaved during this period, but only 76% of these produced two mononucleated daughter blastomeres; in the remainder, one or more of the blastomeres was abnormally nucleated. During overnight culture, 30% of multinucleated blastomeres and 30% of anucleate blastomeres cleaved, the majority producing abnormally nucleated daughter blastomeres. The majority of blastomeres which showed no sign of cleavage after overnight culture retained the same nuclear morphology as when originally disaggregated. However, a small number of mononucleated blastomeres contained two nuclei after culture, indicating that karyokinesis may have taken place in the absence of cytokinesis. Overall, ∼30% of blastomeres with more than one nucleus seemed to arise by this mechanism, the remainder probably arising by errors of chromosome segregation and/or packaging at mitosis. In addition, 25/111 mononucleated daughter cells arose either after abnormal division of mononucleated parent cells or after division of multinucleated cells, suggesting that ∼23% of newly formed mononucleated cells might be chromosomally abnormal. The results of DNA quantitation indicated that very few (12/131, 9.2%) blastomeres (whether unior multinucleated) had a DNA content outside the 2–4C range. The embryos used for these studies had been cultured in one of three commonly used in-vitro fertilization (IVF) media: modified T6, Earle‘s balanced salts or Universal IVF medium (a commercial medium from Medi-Cult). A retrospective analysis was carried out of the number of embryos containing multinucleated blastomeres at disaggregation and of the total proportion of isolated blastomeres which were multinucleated in three groups of embryos, each of which had been cultured in one of the IVF media. Both these parameters were found to vary between cohorts of embryos cultured in the different media. The mechanism(s) by which culture medium composition might affect multinucleation of human blastomeres is discussed, as is the significance of these data for reliable preimplantation diagnosis of genetic status.