Non-invasive imaging of mouse embryo metabolism in response to induced hypoxia

Abstract

Purpose

This study used noninvasive, fluorescence lifetime imaging microscopy (FLIM)-based imaging of NADH and FAD to characterize the metabolic response of mouse embryos to short-term oxygen deprivation. We investigated the response to hypoxia at various preimplantation stages.

Methods

Mouse oocytes and embryos were exposed to transient hypoxia by dropping the oxygen concentration in media from 5–0% over the course of ~1.5 h, then 5% O2 was restored. During this time, FLIM-based metabolic imaging measurements of oocyte/embryo cohorts were taken every 3 minutes. Experiments were performed in triplicate for oocytes and embryos at the 1- to 8-cell, morula, and blastocyst stages. Maximum hypoxia response for each of eight measured quantitative FLIM parameters was taken from the time points immediately before oxygen restoration.

Results

Metabolic profiles showed significant changes in response to hypoxia for all stages of embryo development. The response of the eight measured FLIM parameters to hypoxia was highly stage-dependent. Of the eight FLIM parameters measured, NADH and FAD intensity showed the most dramatic metabolic responses in early developmental stages. At later stages, however, other parameters, such as NADH fraction engaged and FAD lifetimes, showed greater changes. Metabolic parameter values generally returned to baseline with the restoration of 5% oxygen.

Conclusions

Quantitative FLIM-based metabolic imaging was highly sensitive to metabolic changes induced by hypoxia. Metabolic response profiles to oxygen deprivation were distinct at different stages, reflecting differences in metabolic plasticity as preimplantation embryos develop.

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Funding

This study received support from the following sources. A grant from Vivere Health (E.A.S.). Harvard Catalyst, The Harvard Clinical and Translational Science Center (National Institutes of Health Award UL1 TR001102); National Science Foundation (DMR-0820484 and PFI-TT-1827309); National Institutes of Health (R01HD092550-01); National Science Foundation Postdoctoral Research Fellowship in Biology (1308878 T.S.); Becker and Hickl GmbH sponsored research with the loaning of equipment for FLIM.

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EAS primarily contributed to study conception and design and data acquisition, analysis, and interpretation; drafted and critically revised the manuscript; and approved the final version for publication. TS contributed to study conception and design and data acquisition, analysis, and interpretation; drafted and critically revised the manuscript; and approved the final version for publication. DS contributed to study conception and design and data interpretation; critically revised the manuscript; and approved the final version for publication. MV contributed to design and data interpretation, and approved the final version for publication. DJN contributed to study conception, design and data interpretation, critically revised the manuscript, and approved the final version for publication.

Corresponding author

Correspondence to Emily A. Seidler.

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Conflicts of interest/Competing interests

TS and DJN co-hold patent US20150346100A1 pending for metabolic imaging methods for assessment of oocytes and embryos and patent US20170039415A1 issued for nonlinear imaging systems and methods for assisted reproductive technologies.

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Seidler, E.A., Sanchez, T., Venturas, M. et al. Non-invasive imaging of mouse embryo metabolism in response to induced hypoxia. J Assist Reprod Genet 37, 1797–1805 (2020). https://doi.org/10.1007/s10815-020-01872-w

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Keywords

  • Embryo metabolism
  • Hypoxia
  • Noninvasive assessment
  • Metabolic imaging
  • FLIM
  • Mitochondria