Role of protein kinase G in barrier-protective effects of cGMP in human pulmonary artery endothelial cells

Aigul Moldobaeva, Laura E. Welsh-Servinsky, Larissa A. Shimoda, R. Scott Stephens, Alexander D. Verin, Rubin M. Tuder, David B. Pearse

Research output: Contribution to journalArticle

34 Scopus citations

Abstract

Increases in endothelial cGMP prevent oxidant-mediated endothelial barrier dysfunction, but the downstream mechanisms remain unclear. To determine the role of cGMP-dependent protein kinase (PKG)I, human pulmonary artery endothelial cells (HPAEC) lacking PKGI expression were infected with a recombinant adenovirus encoding PKG (Ad.PKG) and compared with uninfected and control-infected (Ad.βgal) HPAEC. Transendothelial electrical resistance (TER), an index of permeability, was measured after H 2O2 (250 μM) exposure with or without pretreatment with 8-(4-chlorophenylthio)guanosine 3′,5′-cyclic monophosphate (CPT-cGMP). HPAEC infected with Ad.PKG, but not Ad.βgal, expressed PKG I protein and demonstrated Ser239 and Ser157 phosphorylation of vasodilator-stimulated phosphoprotein after treatment with CPT-cGMP. Adenoviral infection decreased basal permeability equally in Ad.PKG- and Ad.βgal-infected HPAEC compared with uninfected cells. Treatment with CPT-cGMP (100 μM) caused a PKGI-independent decrease in permeability (8.2 ± 0.6%). In all three groups, H2O 2 (250 μM) caused a similar ∼35% increase in permeability associated with increased actin stress fiber formation, intercellular gaps, loss of membrane VE-cadherin, and increased intracellular Ca2+ concentration ([Ca2+]i). In uninfected and Ad.βgal-infected HPAEC, pretreatment with CPT-cGMP (100 μM) partially blocked the increased permeability induced by H2O2. In Ad.PKG-infected HPAEC, CPT-cGMP (50 μM) prevented the H2O 2-induced TER decrease, cytoskeletal rearrangement, and loss of junctional VE-cadherin. CPT-cGMP attenuated the peak [Ca2+] i caused by H2O2 similarly (23%) in Ad.βgal- and Ad.PKG-infected HPAEC, indicating a PKGI- independent effect. These data suggest that cGMP decreased HPAEC basal permeability by a PKGI-independent process, whereas the ability of cGMP to prevent H2O2-induced barrier dysfunction was predominantly mediated by PKGI through a Ca2+-independent mechanism.

Original languageEnglish (US)
Pages (from-to)L919-L930
JournalAmerican Journal of Physiology - Lung Cellular and Molecular Physiology
Volume290
Issue number5
DOIs
StatePublished - May 1 2006
Externally publishedYes

Keywords

  • Acute lung injury
  • Hydrogen peroxide
  • Pulmonary edema
  • cAMP

ASJC Scopus subject areas

  • Physiology
  • Pulmonary and Respiratory Medicine
  • Physiology (medical)
  • Cell Biology

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