Browsing by Author "Varghese, Jacob"

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    A Comparison of the Developmental Competence of In Vitro- Versus In Vivo-Produced Mouse Embryos
    (2021-10-12) Varghese, Jacob; Thundathil, Jacob; Wong, Benjamin; Ungrin, Mark
    The prevalence of infertility in Canada has substantially increased over the last 30 years with one in six couples seeking medical intervention, many of which rely on assisted reproductive technologies (ARTs) including in vitro fertilization (IVF) and in vitro culture (IVC) of embryos. In clinical IVF programs, embryos undergo extended culture until the blastocyst stage (~6 days) under 5% or 2% O2 concentrations to facilitate success of single embryo transfer. Modest success rates of these culture systems warrant further research to refine culture conditions. The objectives of this study were to characterize stress-related responses of cultured embryos under 5% and 2% O2 in comparison to in vivo-derived blastocysts using a CD1 mouse model. We hypothesized 2% O2 during mouse in vitro embryo culture compromises developmental competence through altered transcriptional profiles of genes involved in the embryonic stress response and apoptosis relative to 5% O2-cultured and in vivo-derived blastocysts. The relative expressions of a cohort of stress-related genes in mouse blastocysts cultured under 5 or 2% O2 were quantified through RT-qPCR and compared to in vivo¬-derived embryos. Apoptotic responses were evaluated using an immunofluorescence assay specific for Caspase-3 as the executioner protein in apoptosis. The mean percentage of blastocysts developed in vivo or cultured under 5% O2 was significantly higher than those cultured under 2% O2. Furthermore, in vivo-derived embryos, or those cultured under 5% O2, had greater total cell numbers relative to those cultured under 2% O2. The expansion status of blastocysts was also greatest in embryos cultured under 5% O2. In general, stress response genes were significantly upregulated in embryos cultured under 2% O2, and the expression of antioxidant-related genes was significantly lower in cultured embryos versus those derived in vivo. Caspase-3 immunofluorescence was significantly higher in cultured embryos versus in vivo-derived embryos.Therefore, we inferred that 5% O2 systems may be optimal for culture of mouse embryos, warranting critical evaluation of culturing human embryos under continuous 2% O2 concentrations during clinical IVF programs.