AN ANALYSIS OF MULTINUCLEATED BLASTOMERE FORMATION IN HUMAN EMBRYOS

Human embryos were disaggregated into component blastomeres 42-72 h af ter 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 blastom eres cleaved during this period, but only 76% of these produced two mo nonucleated daughter blastomeres; in the remainder, one or more of the blastomeres was abnormally nucleated. During overnight culture, 30% o f 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 overni ght culture retained the same nuclear morphology as when originally di saggregated. However, a small number of mononucleated blastomeres cont ained two nuclei after culture, indicating that karyokinesis may have taken place in the absence of cytokinesis. Overall, similar to 30% of blastomeres with more than one nucleus seemed to arise by this mechani sm, the remainder probably arising by errors of chromosome segregation and/or packaging at mitosis. In addition, 25/111 mononucleated daught er cells arose either after abnormal division of mononucleated parent cells or after division of multinucleated cells, suggesting that simil ar to 23% of newly formed mononucleated cells might be chromosomally a bnormal. The results of DNA quantitation indicated that very few (12/1 31, 9.2%) blastomeres (whether uni-or multinucleated) had a DNA conten t outside the 2-4C range. The embryos used for these studies had been cultured in one of three commonly used in-vitro fertilization (IVF) me dia: modified T6, Earle's balanced salts or Universal IVF medium (a co mmercial 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 whi ch were multinucleated in three groups of embryos, each of which had b een cultured in one of the IVF media. Both these parameters were found to vary between cohorts of embryos cultured in the different media. T he mechanism(s) by which culture medium composition might affect multi nucleation of human blastomeres is discussed, as is the significance o f these data for reliable preimplantation diagnosis of genetic status.