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Role of Rac 1 and cAMP in endothelial barrier stabilization and thrombin‐induced barrier breakdown
Author(s) -
Spindler Volker,
Baumer Yvonne,
Werthmann Ruth,
Buenemann Moritz,
Waschke Jens
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.796.1
Subject(s) - forskolin , thrombin , intracellular , chemistry , rolipram , biophysics , small gtpase , microbiology and biotechnology , protein kinase a , kinase , signal transduction , biology , biochemistry , receptor , phosphodiesterase , enzyme , immunology , platelet
Barrier stabilizing effects of cAMP as well as of the small GTPase Rac 1 are well established. Moreover, it is generally believed that permeability‐increasing mediators such as thrombin disrupt endothelial barrier functions primarily via activation of Rho A. In this study, we provide evidence that decrease of both cAMP levels and of Rac 1 activity contribute to thrombin‐mediated barrier break‐down. Treatment of human dermal microvascular endothelial cells (HDMEC) with Rac 1‐inhibitor NSC‐23766 decreased transendothelial electrical resistance (TER) and caused intercellular gap formation. These effects were reversed by addition of forskolin/rolipram (F/R) to increase intracellular cAMP but not by the cAMP analogue O‐Me‐cAMP which primarily stimulates protein kinase A‐independent signaling via Epac/Rap 1. However, both F/R and O‐Me‐cAMP did not increase TER above control levels in the presence of NSC‐23766 in contrast to experiments without Rac 1 inhibition. Next, we tested the role of Rac 1 and cAMP in thrombin‐induced barrier break‐down. Thrombin‐induced drop of TER and intercellular gap formation were paralleled by a rapid decrease of cAMP as revealed by fluorescence resonance energy transfer (FRET). The efficacy of F/R or O‐Me‐cAMP to block barrier‐destabilizing effects of thrombin was comparable to Y27632‐induced inhibition of Rho kinase but was blunted when Rac 1 was inactivated by NSC‐23766.