Optical imaging of cleavage stage bovine embryos using hyperspectral and confocal approaches reveals metabolic differences between on-time and fast-developing embryos
| Autor: | Ana Maria Reis Ferreira, Tiffany C. Y. Tan, Jeremy G. Thompson, Kylie R. Dunning, Clara Ana Santos Monteiro, Darren J X Chow, Gabriela Ramos Leal |
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| Rok vydání: | 2021 |
| Předmět: |
animal structures
DNA damage Confocal Embryonic Development Fertilization in Vitro Biology law.invention Andrology 03 medical and health sciences 0302 clinical medicine Human fertilization Food Animals Pregnancy Confocal microscopy law medicine Animals Blastocyst Small Animals Microscopy 030219 obstetrics & reproductive medicine Equine Optical Imaging 0402 animal and dairy science Embryo 04 agricultural and veterinary sciences Embryo Transfer Embryo Mammalian 040201 dairy & animal science Autofluorescence medicine.anatomical_structure embryonic structures Cattle Female Animal Science and Zoology Embryo quality |
| Zdroj: | Theriogenology. 159:60-68 |
| ISSN: | 0093-691X |
| Popis: | The assessment of embryo quality aims to enhance subsequent pregnancy and live birth outcomes. Metabolic analysis of embryos has immense potential in this regard. As a step towards this goal, here we assess the metabolism of bovine embryos using label-free optical imaging. We compared embryos defined as either on-time or fast-developing, as fast dividing embryos are more likely to develop to the blastocyst stage. Specifically, bovine embryos at 48 (Day 2) and 96 (Day 4) hours post fertilization were fixed and separated based on morphological assessment: on-time (Day 2: 2 cell; Day 4: 5–7 cell) or fast-developing (Day 2: 3-7 cell; Day 4: 8–16 cell). Embryos with different developmental rates on Day 2 and Day 4 were correlated with metabolic activity and DNA damage. Confocal microscopy was used to assess metabolic activity by quantification of cellular autofluorescence specific for the endogenous fluorophores NAD(P)H and FAD with a subsequent calculation of the optical redox ratio. Separately, hyperspectral microscopy was employed to assess a broader range of endogenous fluorophores. DNA damage was determined using γH2AX immunohistochemistry. Hyperspectral imaging showed significantly lower abundance of endogenous fluorophores in fast-developing compared to on-time embryos on Day 2, indicating a lower metabolic activity. On Day 4 of development there was no difference in the abundance of FAD between on-time and fast-developing embryos. There was, however, significantly higher levels of NAD(P)H in fast-developing embryos leading to a significantly lower optical redox ratio when compared to on-time embryos. Collectively, these results demonstrate that fast-developing embryos present a ‘quiet’ metabolic pattern on Day 2 and Day 4 of development, compared to on-time embryos. There was no difference in the level of DNA damage between on-time and fast-developing embryos on either day of development. To our knowledge, this is the first collective use of confocal and hyperspectral imaging in cleavage-stage bovine embryos in the absence of fluorescent tags. |
| Databáze: | OpenAIRE |
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