Hyperspectral microscopy can detect metabolic heterogeneity within bovine post-compaction embryos incubated under two oxygen concentrations (7% versus 20%)

• Published in: Human reproduction (Oxford) 2017-10, Vol.32 (10), p.2016-2025
• Main Authors: Sutton-McDowall, Melanie L., Gosnell, Martin, Anwer, Ayad G., White, Melissa, Purdey, Malcolm, Abell, Andrew D., Goldys, Ewa M., Thompson, Jeremy G.
• Format: Article
• Language: English
• ISSN: 0268-1161; 1460-2350
• DOI: 10.1093/humrep/dex261

Abstract STUDY QUESTION Can we separate embryos cultured under either 7% or 20% oxygen atmospheres by measuring their metabolic heterogeneity? SUMMARY ANSWER Metabolic heterogeneity and changes in metabolic profiles in morula exposed to two different oxygen concentrations were not detectable using traditional fluorophore and two-channel autofluorescence but were detectable using hyperspectral microscopy. WHAT IS KNOWN ALREADY Increased genetic and morphological blastomere heterogeneity is associated with compromised developmental competence of embryos and currently forms the basis for embryo scoring within the clinic. However, there remains uncertainty over the accuracy of current techniques, such as PGS and time-lapse microscopy, to predict subsequent pregnancy establishment. STUDY DESIGN, SIZE, DURATION The impact of two oxygen concentrations (7% = optimal and 20% = stressed) during post-fertilisation embryo culture was assessed. Cattle embryos were exposed to the different oxygen concentrations for 8 days (D8; embryo developmental competence) or 5 days (D5; metabolism measurements). Between 3 and 4 experimental replicates were performed, with 40–50 embryos per replicate used for the developmental competency experiment, 10–20 embryos per replicate for the fluorophore and two-channel autofluorescence experiments and a total of 21–22 embryos used for the hyperspectral microscopy study. PARTICIPANTS/MATERIALS, SETTING, METHODS In-vitro produced (IVP) cattle embryos were utilised for this study. Post-fertilisation, embryos were exposed to 7% or 20% oxygen. To determine impact of oxygen concentrations on embryo viability, blastocyst development was assessed on D8. On D5, metabolic heterogeneity was assessed in morula (on-time) embryos using fluorophores probes (active mitochondria, hydrogen peroxide and reduced glutathione), two-channel autofluorescence (FAD and NAD(P)H) and 18-channel hyperspectral microscopy. MAIN RESULTS AND THE ROLE OF CHANCE Exposure to 20% oxygen following fertilisation significantly reduced total blastocyst, expanded and hatched blastocyst rates by 1.4-, 1.9- and 2.8-fold, respectively, compared to 7% oxygen (P < 0.05), demonstrating that atmospheric oxygen was a viable model for studying mild metabolic stress. The metabolic profiles of D5 embryos was determined and although metabolic heterogeneity was evident within the cleavage stage (i.e. arrested) embryos exposed to fluorophores, there were no detectable difference in fluorescence