A Method for Real-Time Assessment of Mitochondrial Respiration Using Murine Corneal Biopsy

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dc.creatorLiang, Wentao
dc.creatorHuang, Li
dc.creatorYuan, Tian
dc.creatorCheng, Rui
dc.creatorTakahashi, Yusuke
dc.creatorMoiseyev, Gennadiy P.
dc.creatorKaramichos, Dimitrios
dc.creatorMa, Jian-Xing
dc.creator.orcid0000-0002-8761-3824 (Karamichos, Dimitrios)
dc.date.accessioned2023-09-14T14:18:39Z
dc.date.available2023-09-14T14:18:39Z
dc.date.issued2023-08-29
dc.description.abstractPURPOSE: To develop and optimize a method to monitor real-time mitochondrial function by measuring the oxygen consumption rate (OCR) in murine corneal biopsy punches with a Seahorse extracellular flux analyzer. METHODS: Murine corneal biopsies were obtained using a biopsy punch immediately after euthanasia. The corneal metabolic profile was assessed using a Seahorse XFe96 pro analyzer, and mitochondrial respiration was analyzed with specific settings. RESULTS: Real-time adenosine triphosphate rate assay showed that mitochondrial oxidative phosphorylation is a major source of adenosine triphosphate production in ex vivo live murine corneal biopsies. Euthanasia methods (carbon dioxide asphyxiation vs. overdosing on anesthetic drugs) did not affect corneal OCR values. Mouse corneal biopsy punches in 1.5-mm diameter generated higher and more reproducible OCR values than those in 1.0-mm diameter. The biopsy punches from the central and off-central cornea did not show significant differences in OCR values. There was no difference in OCR reading by the tissue orientations (the epithelium side up vs. the endothelium side up). No significant differences were found in corneal OCR levels between sexes, strains (C57BL/6J vs. BALB/cJ), or ages (4, 8, and 32 weeks). Using this method, we showed that the wound healing process in the mouse cornea affected mitochondrial activity. CONCLUSIONS: The present study validated a new strategy to measure real-time mitochondrial function in fresh mouse corneal tissues. This procedure should be helpful for studies of the ex vivo live corneal metabolism in response to genetic manipulations, disease conditions, or pharmacological treatments in mouse models.
dc.description.sponsorshipFunded by National Institutes of Health/National Eye Institute grants (EY012231, EY028773, EY028949, EY032930, EY032931, EY033330, and EY033477).
dc.identifier.citationLiang, W., Huang, L., Yuan, T., Cheng, R., Takahashi, Y., Moiseyev, G. P., Karamichos, D., & Ma, J. X. (2023). A Method for Real-Time Assessment of Mitochondrial Respiration Using Murine Corneal Biopsy. Investigative ophthalmology & visual science, 64(11), 33. https://doi.org/10.1167/iovs.64.11.33
dc.identifier.issn1552-5783
dc.identifier.issue11
dc.identifier.urihttps://hdl.handle.net/20.500.12503/32411
dc.identifier.volume64
dc.publisherAssociation for Research in Vision and Ophthalmology
dc.relation.urihttps://doi.org/10.1167/iovs.64.11.33
dc.rights.holderCopyright 2023 The Authors
dc.rights.licenseAttribution 4.0 International (CC BY 4.0)
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/
dc.sourceInvestigative Ophthalmology & Visual Science
dc.subjectcornea
dc.subjectmitochondria
dc.subjectmetabolism
dc.subjectbiopsy
dc.subjectSeahorse
dc.subject.meshAnimals
dc.subject.meshMice
dc.subject.meshMice, Inbred C57BL
dc.subject.meshRespiration
dc.subject.meshBiopsy
dc.subject.meshCornea
dc.subject.meshAdenosine Triphosphate
dc.subject.meshMitochondria
dc.titleA Method for Real-Time Assessment of Mitochondrial Respiration Using Murine Corneal Biopsy
dc.typeArticle
dc.type.materialtext

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