IVF clinics increasingly culture embryos individually to facilitate morphometric and genetic analyses. However, culturing embryos individually rather than in groups deprives them of paracrine factors produced by neighbouring embryos and this reduces blastocyst cell numbers. Further examination of the differences between embryos cultured individually or in groups, including their response to other stresses in vitro and the role of paracrine factors, is therefore warranted.
Zygotes from CBA x C57BL/6 mice were cultured individually in 2 µL G1/G2 medium or in groups of 10 in 20 µL at 5% oxygen. Time-lapse microscopy revealed that individually cultured embryos were delayed reaching the 8-cell stage (P<0.01), and the resulting blastocysts had fewer cells and a reduced proportion of inner cell mass cells compared to embryos cultured in groups (P<0.05). Increasing the drop size from 2 µL to 20 µL further reduced cell numbers of individually cultured embryos (P<0.05), presumably due to dilution of embryo-secreted factors. In support of this, the addition of embryo-conditioned media to single embryos increased cell numbers compared to controls (P<0.001).
To determine the effect of an additional in vitro stress, embryos were cultured in atmospheric oxygen. The effect of combined single culture and atmospheric oxygen on cell numbers was more detrimental than each condition alone (P<0.001), indicating that there is a cumulative effect of these stresses.
Despite the differences observed during the preimplantation stages, embryos cultured individually or in groups performed equally in outgrowth assays and no differences were observed in fetal or placental weights on day 15 of pregnancy.
When considering the relevance of these findings to clinical IVF, individual culture may reduce the numbers of transferrable blastocysts available, even if no fetal consequences are apparent. Furthermore, many clinics routinely culture embryos both individually and in atmospheric oxygen, which have cumulative detrimental effects on blastocyst development.