AMP-activated protein kinase (AMPK)-dependent and -independent pathways regulate hypoxic inhibition of transepithelial Na(+) transport across human airway epithelial cells |
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Authors: | Tan Cd Smolenski Rt Harhun Mi Patel Hk Ahmed Sg Wanisch K Yáñez-Muñoz Rj Baines Dl |
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Institution: | Pharmacology and Cell Physiology Research Group, Division of Biomedical Sciences, St George's University of London, Cranmer Terrace, London, UK Department of Biochemistry, Medical University of Gdansk, Gdansk, Poland School of Biological Sciences, Royal Holloway, University of London, Egham, Surrey, UK. |
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Abstract: | BACKGROUND AND PURPOSEPulmonary transepithelial Na+ transport is reduced by hypoxia, but in the airway the regulatory mechanisms remain unclear. We investigated the role of AMPK and ROS in the hypoxic regulation of apical amiloride-sensitive Na+ channels and basolateral Na+K+ ATPase activity.EXPERIMENTAL APPROACHH441 human airway epithelial cells were used to examine the effects of hypoxia on Na+ transport, AMP : ATP ratio and AMPK activity. Lentiviral constructs were used to modify cellular AMPK abundance and activity; pharmacological agents were used to modify cellular ROS.KEY RESULTSAMPK was activated by exposure to 3% or 0.2% O2 for 60 min in cells grown in submerged culture or when fluid (0.1 mL·cm?2) was added to the apical surface of cells grown at the air–liquid interface. Only 0.2% O2 activated AMPK in cells grown at the air–liquid interface. AMPK activation was associated with elevation of cellular AMP : ATP ratio and activity of the upstream kinase LKB1. Hypoxia inhibited basolateral ouabain-sensitive Isc (Iouabain) and apical amiloride-sensitive Na+ conductance (GNa+). Modification of AMPK activity prevented the effect of hypoxia on Iouabain (Na+K+ ATPase) but not apical GNa+. Scavenging of superoxide and inhibition of NADPH oxidase prevented the effect of hypoxia on apical GNa+ (epithelial Na+ channels).CONCLUSIONS AND IMPLICATIONSHypoxia activates AMPK-dependent and -independent pathways in airway epithelial cells. Importantly, these pathways differentially regulate apical Na+ channels and basolateral Na+K+ ATPase activity to decrease transepithelial Na+ transport. Luminal fluid potentiated the effect of hypoxia and activated AMPK, which could have important consequences in lung disease conditions. |
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Keywords: | AMPK ROS airway epithelium Na+K+ ATPase ENaC |
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