Petra L. Wale

Glucose consumption of single post-compaction human embryos is predictive of embryo sex and live birth outcome

BACKGROUND The aim of this study was to determine the relationship between nutrient utilization by the human embryo and its subsequent viability after transfer. METHODS The embryos of 50 patients having single blastocyst transfer were cultured individually from Day 3 in 10 µl drops of medium G2 under Ovoil in 5%O(2), 6%CO(2), 89%N(2). Patient inclusion in the study was maternal age ≤ 38. Embryos were moved to fresh drops of medium every 24 h. Spent media samples, including controls containing no embryo, were coded, frozen and subsequently analysed blind. Analysis of glucose was performed by microfluorimetry. The sex of children born was recorded. RESULTS Clinical pregnancy and live birth rates were 58 and 56%, respectively. Glucose consumption by embryos which resulted in a pregnancy was significantly higher on both Day 4 and Day 5 than that by embryos which failed to develop post-transfer (P < 0.01). Furthermore, on Day 4 female embryos consumed 28% more glucose compared with males (P < 0.05). Glucose uptake was independent of embryo grade. CONCLUSIONS The rapid screening of glucose metabolism by the human embryo on Day 4 and 5 may prove to be a useful metric in the development of algorithms for the selection of embryos for transfer in human IVF. Also, the observed sex-related metabolic difference provides preliminary data to support the hypothesis that male and female human embryos differ in their physiology due to the presence of two active X chromosomes and an altered proteome for a finite time during the preimplantation period.

Time-lapse analysis of mouse embryo development in oxygen gradients

Atmospheric oxygen (approximately 20%) in culture significantly impairs preimplantation embryo development. However, it is not known whether all stages of preimplantation embryo development are susceptible to oxygen toxicity. This study investigated the temporal responses of preimplantation embryos to oxygen conditions in vitro. Mouse embryos were cultured in atmospheric (approximately 20%) or lower (5%) oxygen concentrations for the first 48 h, followed by culture in the same or reciprocal oxygen concentrations for another 48 h: group 1 (control, 5 and 5%); group 2 (5 and 20%); group 3 (20 and 5%); and group 4 (20 and 20%). Time-lapse microscopy was performed with imaging of individual embryos at 15-min intervals. Compared with embryos cultured in 5% oxygen, embryos cultured in 20% oxygen were delayed at the 1st cleavage by 0.45 h (P<0.05), at the 2nd cleavage by 0.84 h (P<0.01) and at the 3rd cleavage by 3.19 h (P<0.001). Switching from 20% to 5% oxygen after 48 h did not completely alleviate earlier induced perturbations. Partial or complete culture in atmospheric oxygen resulted in significantly fewer blastocyst cell numbers compared with control (P<0.05). Oxygen can influence mouse embryo development at both the cleavage and post-compaction stages.

Oxygen Regulates Amino Acid Turnover and Carbohydrate Uptake During the Preimplantation Period of Mouse Embryo Development

ABSTRACT Oxygen is a powerful regulator of preimplantation embryo development, affecting gene expression, the proteome, and energy metabolism. Even a transient exposure to atmospheric oxygen can have a negative impact on embryo development, which is greatest prior to compaction, and subsequent postcompaction culture at low oxygen cannot alleviate this damage. In spite of this evidence, the majority of human in vitro fertilization is still performed at atmospheric oxygen. One of the physiological parameters shown to be affected by the relative oxygen concentration, carbohydrate metabolism, is linked to the ability of the mammalian embryo to develop in culture and remain viable after transfer. The aim of this study was, therefore, to determine the effect of oxygen concentration on the ability of mouse embryos to utilize both amino acids and carbohydrates both before and after compaction. Metabolomic and fluorometric analysis of embryo culture media revealed that when embryos were exposed to atmospheric oxygen during the cleavage stages, they exhibited significantly greater amino acid utilization and pyruvate uptake than when cultured under 5% oxygen. In contrast, postcompaction embryos cultured in atmospheric oxygen showed significantly lower mean amino acid utilization and glucose uptake. These metabolic changes correlated with developmental compromise because embryos grown in atmospheric oxygen at all stages showed significantly lower blastocyst formation and proliferation. These findings confirm the need to consider both embryo development and metabolism in establishing optimal human embryo growth conditions and prognostic markers of viability, and further highlight the impact of oxygen on such vital parameters.

Oxygen Affects the Ability of Mouse Blastocysts to Regulate Ammonium

ABSTRACT During embryo culture, ammonium is generated by amino acid metabolism and from the spontaneous deamination of amino acids at 37°C. Although ammonium has been shown to be embryo toxic, few studies have investigated the mechanism(s) by which the early embryo can regulate ammonium. Whilst 20% oxygen represents a source of stress to the developing embryo, it is not known how oxygen affects the physiology of the embryo in the presence of other sources of stress. The aim of this study was, therefore, to investigate possible pathways involved in ammonium sequestration in the preimplantation embryo and the effect of oxygen on the regulation of these pathways. Glutamine and alanine were investigated as possible ammonium sequestration pathways. Amino acid utilization by blastocysts was determined after culture from the postcompaction stage with 0, 150, or 300 μM ammonium (in either 5% or 20% oxygen) and with or without 500 μM L-methionine sulfoximine (MSO), an inhibitor of glutamine synthetase. In the presence of MSO, ammonium production was significantly increased and glutamate was no longer consumed. Glutamine synthetase inhibition with MSO significantly decreased glutamine formation. Ammonium and oxygen independently altered overall amino acid turnover. Together, 5% oxygen and ammonium promoted glutamine production, whereas in the presence of 20% oxygen and ammonium, glutamine was consumed. Data reveal that both oxygen and ammonium affect amino acid utilization by the developing embryo, however, 20% oxygen appears to have the greater impact. Mouse blastocysts can alleviate ammonium stress by its transamination to both glutamine and alanine, but only under physiological conditions.