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Increased number of cells and metabolic activity in male human preimplantation embryos following in vitro fertilization

The number of cells and metabolic activity of male and female human preimplantation embryos were examined to determine whether male embryos are more advanced than female embryos following in vitro fertilization (IVF). The metabolic activity of embryos fertilized normally was assessed daily by non-invasive measurement of pyruvate and glucose uptake and lactate production between days 2 and 6 after insemination. On day 6, the numbers of nuclei from the trophectoderm and inner cell mass of blastocysts were counted by differential labelling and fluorescence microscopy. Nuclei were then recovered and the sex of the embryos identified using nested primers to amplify the amelogenin gene and pseudogene sequences on the X and Y chromosomes, respectively. Development of male and female embryos were then compared retrospectively. From 69 of 178 (39%) embryos that developed to the blastocyst stage, the sex of 57 was determined; 21 (37%) were male and 36 (63%) female. The number of cells in male embryos was significantly greater on day 2 (P < 0.005), and this difference was maintained up to the blastocyst stage (in both the trophectoderm and the inner cell mass), although differences were not always significant. Pyruvate uptake was significantly higher by male embryos between days 2 and 5 (P < 0.05). Glucose uptake and lactate production were significantly higher in male embryos on days 4–5 (P <0.05); this difference was not significant on days 5–6. Extrapolation from differences in the number of cells indicates that female embryos are approximately 4.5 h delayed in their development from day 2 onwards compared with male embryos. As there was no evidence for a differential cleavage rate beyond this stage, it is suggested that differences between males and females occur early, either at fertilization, or during the first or second cleavage.