title: Enhanced depolarization-induced pulmonary vasoconstriction following chronic hypoxia requires EGFR-dependent activation of NAD(P)H oxidase 2.
abstract: OBJECTIVE
  Chronic hypoxia (CH) enhances depolarization-induced myofilament Ca(2+) sensitization and resultant pulmonary arterial constriction through superoxide (O(2)(-))-dependent stimulation of RhoA. Because NAD(P)H oxidase (NOX) has been implicated in the development of pulmonary hypertension, we hypothesized that vascular smooth muscle (VSM) depolarization increases NOX-derived O(2)(-) production leading to myofilament Ca(2+) sensitization and augmented vasoconstrictor reactivity following CH. As epidermal growth factor receptor (EGFR) mediates Rac1-dependent NOX activation in renal mesangial cells, we further sought to examine the role EGFR plays in this response.
  
  
  RESULTS
  Vasoconstrictor responses to depolarizing concentrations of KCl were greater in lungs isolated from CH (4 wk, 0.5 atm) rats compared to normoxic controls, and this effect of CH was abolished by the general NOX inhibitor, apocynin. CH similarly augmented KCl-induced vasoconstriction and O(2)(-) generation (assessed using the fluorescent indicator, dihydroethidium) in Ca(2+)-permeabilized, pressurized small pulmonary arteries. These latter responses to CH were prevented by general inhibition of NOX isoforms (apocynin, diphenylene iodonium), and by selective inhibition of NOX 2 (gp91ds-tat), Rac1 (NSC 23766), and EGFR (AG 1478). Consistent with these observations, CH increased KCl-induced EGFR phosphorylation, and augmented depolarization-induced Rac1 activation in an EGFR-dependent manner.
  
  
  METHODS
  This study establishes a novel signaling axis in VSM linking membrane depolarization to contraction that is independent of Ca(2+) influx, and which mediates myofilament Ca(2+) sensitization in the hypertensive pulmonary circulation.
  
  
  CONCLUSIONS
  CH augments membrane depolarization-induced pulmonary VSM Ca(2+) sensitization and vasoconstriction through EGFR-dependent stimulation of Rac1 and NOX 2.
authors.affiliation: Vascular Physiology Group, Department of Cell Biology and Physiology, University of New Mexico Health Sciences Center, Albuquerque, NM 87131, USA.
authors.forename: Charles E
authors.initials: CE
authors.lastname: Norton
authors.forename: Brad R S
authors.initials: BR
authors.lastname: Broughton
authors.forename: Nikki L
authors.initials: NL
authors.lastname: Jernigan
authors.forename: Benjimen R
authors.initials: BR
authors.lastname: Walker
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Resta
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism; D000860:Hypoxia; D008297:Male; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011189:Potassium Chloride / Q000494:pharmacology; D011651:Pulmonary Artery / Q000187:drug effects* / Q000378:metabolism*; D051381:Rats; D014661:Vasoconstriction / Q000187:drug effects*; D014662:Vasoconstrictor Agents / Q000494:pharmacology*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism
chemical_list: D020746:Calcium Channels, L-Type; D008562:Membrane Glycoproteins; D014662:Vasoconstrictor Agents; D011189:Potassium Chloride; D006861:Hydrogen Peroxide; C500593:Cybb protein, rat; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D066246:ErbB Receptors; D020830:rac1 GTP-Binding Protein; D002118:Calcium