title: AMP-activated protein kinase (AMPK)-dependent and -independent pathways regulate hypoxic inhibition of transepithelial Na+ transport across human airway epithelial cells.
abstract: OBJECTIVE
  Pulmonary 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.
  
  
  METHODS
  H441 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.
  
  
  RESULTS
  AMPK was activated by exposure to 3% or 0.2% O(2) 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% O(2) 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 I(sc) (I(ouabain) ) and apical amiloride-sensitive Na(+) conductance (G(Na+) ). Modification of AMPK activity prevented the effect of hypoxia on I(ouabain) (Na(+) K(+) ATPase) but not apical G(Na+) . Scavenging of superoxide and inhibition of NADPH oxidase prevented the effect of hypoxia on apical G(Na+) (epithelial Na(+) channels).
  
  
  CONCLUSIONS
  Hypoxia 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.
authors.affiliation: Pharmacology and Cell Physiology Research Group, Division of Biomedical Sciences, St George's University of London, Cranmer Terrace, London, UK.
authors.forename: C D
authors.initials: CD
authors.lastname: Tan
authors.forename: R T
authors.initials: RT
authors.lastname: Smolenski
authors.forename: M I
authors.initials: MI
authors.lastname: Harhun
authors.forename: H K
authors.initials: HK
authors.lastname: Patel
authors.forename: S G
authors.initials: SG
authors.lastname: Ahmed
authors.forename: K
authors.initials: K
authors.lastname: Wanisch
authors.forename: R J
authors.initials: RJ
authors.lastname: Yáñez-Muñoz
authors.forename: D L
authors.initials: DL
authors.lastname: Baines
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport / Q000502:physiology*; D002460:Cell Line; D004847:Epithelial Cells / Q000502:physiology*; D005786:Gene Expression Regulation / Q000502:physiology; D005822:Genetic Vectors; D006801:Humans; D016086:Lentivirus; D010100:Oxygen / Q000378:metabolism / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000037:antagonists & inhibitors; D020545:Respiratory Mucosa / Q000166:cytology*; D012964:Sodium / Q000378:metabolism*; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics / Q000378:metabolism
chemical_list: D017382:Reactive Oxygen Species; D012964:Sodium; D055372:AMP-Activated Protein Kinases; D000254:Sodium-Potassium-Exchanging ATPase; D010100:Oxygen