Role of Rac 1 and cAMP in Endothelial Barrier Stabilization and Thrombin-induced Barrier Breakdown

被引:87
作者
Baumer, Y. [1 ]
Spindler, V. [1 ]
Werthmann, R. C. [2 ]
Buenemann, M. [2 ]
Waschke, J. [1 ]
机构
[1] Univ Wurzburg, Inst Anat & Cell Biol, D-97070 Wurzburg, Germany
[2] Univ Wurzburg, Inst Pharmacol & Toxicol, D-97070 Wurzburg, Germany
关键词
RHO GTPASES; SPHINGOSINE; 1-PHOSPHATE; OXIDIZED PHOSPHOLIPIDS; EXCHANGE FACTOR; IN-VIVO; ACTIVATION; PERMEABILITY; INTEGRITY; CELLS; EPAC1;
D O I
10.1002/jcp.21819
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
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 RhoA. In this study,we provide evidence that decrease of both cAMP levels and of Rac I activity contribute to thrombin-mediated barrier breakdown. 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 8-pCPT-2'-O-Methyl-cAMP (O-Me-cAMP) which primarily stimulates protein kinase A (PKA)-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 I inhibition. Because Rac I was required for maintenance of barrier functions as well as for cAMP-mediated barrier stabilization, we tested the role of Rac I and cAMP in thrombin-induced barrier breakdown. 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 I was inactivated by NSC-23766. Taken together, these data indicate that decrease of cAMP and Rac I activity may be an important step in inflammatory barrier disruption. J. Cell. Physiol. 220: 716-726, 2009. (C) 2009 Wiley-Liss, Inc.
引用
收藏
页码:716 / 726
页数:11
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