Alterations of intraembryonic metabolites in preimplantation mouse embryos exposed to elevated concentrations of glucose: A metabolic explanation for the developmental retardation seen in preimplantation embryos from diabetic animals

Abstract

Preimplantation mouse embryos exposed to hyperglycemia, whether in vivo or in vitro, experience delayed development from the 2-cell to blastocyst stage. By comparing metabolites from embryos exposed to high vs. normal glucose conditions, a metabolic explanation for the delayed growth pattern was sought. Fertilized 1-cell embryos obtained from superovulated B6xCBA F1 mice were cultured for 96 h in medium containing 2.8 mM glucose (C) or in medium with added glucose to give 10 mM, 30 mM, or 52 mM glucose (HG). After incubation, each embryo was quick-frozen and freeze-dried. Metabolites were assayed by the ultramicrofluorometric technique and enzymatic cycling to obtain measurable levels in single embryos. Embryos cultured in HG exhibited 7-fold higher intracellular glucose levels than those cultured in C (C: 2.25 ± 0.6 vs. HG: 16.61 ± 2.4 mmol/kg wet weight; p < 0.001; C, n = 9; HG, n = 16). This accumulation of glucose was dose-related and stage-dependent. Citrate (C: 1.07 ± 0.14 vs. HG: 1.98 ± 0.12; p < 0.001), sorbitol (C: 0.41 ± 0.06 vs. HG: 0.57 ± 0.03; p < 0.01), malate (C: 0.81 ± 0.13 vs. HG: 1.72 ± 0.17; p < 0.001), and fructose (C: 2.1 ± 0.3 vs. HG: 5.3 ± 0.6; p < 0.001) were all significantly higher in HG. Also, these metabolites were highest in the most delayed embryos. Glycogen and 6-phosphogluconate levels were not significantly different. In conclusion, intraembryonic levels of glucose, and polyol pathway and Krebs cycle metabolites are elevated and correspond to the degree of developmental delay. These findings suggest that a metabolic abnormality may be responsible for retarded development experienced by embryos exposed to high glucose.