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Title: Probing Real Time Redox Events in Human Airway Epithelial Cells Exposed to an Environmental Peroxide
Air pollutants such as ozone, particulate matter, and secondary organic aerosols (SOA) induce intracellular oxidative stress via the generation of reactive oxygen species (ROS). While ROS play important roles in regulating signaling pathways, supra-physiological levels disrupt redox homeostasis and potentiate inappropriate oxidation of regulatory thiols. We examined the effect of isoprene hydroxy hydroperoxide (ISOPOOH), an environmentally derived peroxide that contributes to SOA, on the interplay between bioenergetics and intracellular redox status. We used live cell imaging of human airway epithelial cells (HAECs) expressing the genetically encoded ratiometric biosensors roGFP, iNAP1, and HyPER, to monitor changes in the glutathione redox potential (EGSH), NADPH and H2O2, respectively. Non-cytotoxic exposure to ISOPOOH induced transient increases in EGSH in HAECs that were markedly potentiated by glucose deprivation. ISOPOOH-induced changes in EGSH were not driven by intracellular H2O2. Following ISOPOOH exposure, the addition of 1 mM glucose rapidly restored baseline EGSH and reversed ISOPOOH-induced reductions in NADPH levels, while lower concentrations of glucose (30 uM) induced a bi-modal EGSH recovery. Alternatively, the addition of the glycolytic inhibitor 2-deoxyglucose (2-DG) did not block recovery of NADPH levels nor EGSH restoration. To impair the recovery of EGSH and NADPH levels, we employed a lentiviral vector system to knockdown glucose-6-phosphate dehydrogenase (G6PD), a key enzyme involved in NADPH synthesis. The resulting G6PD knockdown (~50%) did not block glucose-mediated recovery of EGSH, implicating that a partial knockdown of G6PD may not be sufficient to manipulate NADPH levels and thereby EGSH. These findings underscore early mechanisms involved in the cellular response to ISOPOOH while providing a unique live view of the dynamic regulation of redox homeostasis in the human lung during exposure to environmental oxidants. THIS ABSTRACT OF A PROPOSED PRESENTATION DOES NOT NECESSARILY REFLECT EPA POLICY.  more » « less
Award ID(s):
2001027
NSF-PAR ID:
10313555
Author(s) / Creator(s):
; ; ; ; ; ;
Date Published:
Journal Name:
28th Annual Meeting, Society for Redox Biology in Medicine
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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