Metabolism of murine and human embryos: Search for a growth indicator in vitro culture medium

Abstract

Ham's F-10 medium was analyzed biochemically before and after growth of murine and human embryos. Ham's F-10 medium (280 mosm/kg, pH 7.4) alone, by means of reverse-phase high-performance liquid chromatography, demonstrated one major hydrophilic peak, which eluted at 4 to 8 minutes in a 10% to 48% acetonitrile gradient. This peak showed a single peptide of 50 kilodaltons in one-dimensional polyacrylamide gel electrophoresis and size exclusion high-performance liquid chromatography. After the growth of two-cell murine embryos to eight-cell embryos or blastocysts, the major hydrophilic peak was greatly reduced or absent in the culture medium, and in turn a major hydrophobic peak appeared that eluted at 29 minutes. The major hydrophobic peak could not be focused in one-dimensional polyacrylamide gel electrophoresis, but a high content of polar and nonpolar amino acids was revealed in N-terminal sequencing and mass spectrometric analysis. This shift in the peaks was not detected when embryos were cultured in the presence of 0.02% sodium azide. In vitro culture of human zygotes from the pronuclear stage to two to eight cells caused a similar disappearance of the major hydrophilic peak concomitant with the appearance of one to three major hydrophobic peaks in the culture medium. We conclude that the change in profile of culture medium from hydrophilic to hydrophobic peaks on high-performance liquid chromatography is indicative of the metabolic pattern of murine and human embryos. These data also indicate that murine and human embryos do not secrete any major peptide during their development in vitro.