In vitro measurements of oxygen consumption rates in hTERT-RPE cells exposed to low levels of red light

Jeffrey C. Wigle; Cherry C. Castellanos

doi:10.1117/12.2210738

23 March 2016 In vitro measurements of oxygen consumption rates in hTERT-RPE cells exposed to low levels of red light

Jeffrey C. Wigle, Cherry C. Castellanos

Proceedings Volume 9695, Mechanisms of Photobiomodulation Therapy XI; 96950A (2016) https://doi.org/10.1117/12.2210738
Event: SPIE BiOS, 2016, San Francisco, California, United States

Abstract

Exposure to 2.88 J/cm² of red light induces an adaptive response against a lethal pulse of 2.0 μm laser radiation in hTERT-RPE cells in vitro, but not in a knockdown mutant for vascular endothelial growth factor c (VEGF-C). The generally accepted initiation sequence for photobiomodulation is that absorption of red light by cytochome c oxidase (CCOX) of the electron transport chain increases the binding affinity of CCOX for O₂ vs. nitric oxide (NO). This results in displacement of NO by O₂ in the active site of CCOX, thereby increasing cellular respiration and intracellular ATP. We've previously reported that red-light exposure induces a small, but consistently reproducible, increase in NO levels in these cells. But the relative importance of NO and oxidative phosphorylation is unclear because little is known about the relative contributions of NO and ATP to the response. However, if NO dissociation from CCOX actually increases oxidative phosphorylation, one should see a corresponding increase in oxygen consumption. A Seahorse Extracellular Flux Analyzer was used to measure oxygen consumption rates (OCR) in normal and mutant cells as a proxy for oxidative phosphorylation. Both basal respiration and maximum respiration rates in normal cells are significantly higher than in the mutant. The normal cells have a significant amount of “excess capacity,” whereas the VEGF-C(KD) have little or none. The OCR in exposed normal cells is lower than in unexposed cells when measured immediately after exposure. The exposures used for these experiments had no effect on the OCR in mutant cells.

Conference Presentation

Citation Download Citation

Jeffrey C. Wigle and Cherry C. Castellanos "In vitro measurements of oxygen consumption rates in hTERT-RPE cells exposed to low levels of red light", Proc. SPIE 9695, Mechanisms of Photobiomodulation Therapy XI, 96950A (23 March 2016); https://doi.org/10.1117/12.2210738

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