VASP‐mediated Rac 1 activation contributes to cAMP‐induced microvascular endothelial barrier stabilization

Abstract

Vasodilator‐stimulated phosphoprotein (VASP) is required for maintenance of endothelial barrier functions. To test our hypothesis that VASP is involved in regulation of Rac 1 activity, we studied cAMP‐dependent Rac 1 activation. Both, inhibition of Rac 1 activation by NSC‐23677 as well as of protein kinase A (PKA) by PKI completely blunted forskolin/rolipram (F/R)‐mediated cAMP increase to stabilize barrier functions measured as transendothelial resistence (TER). Because this indicates that PKA/Rac 1 activation is important for barrier stabilization, we tested this signalling pathway in VASP (‐/‐) cells. We found that F/R and isoproterenol reduced permeability measured as FITC‐dextran flux across VASP (‐/‐) monolayers, however, not below baseline levels of wild‐type cells (WT). Moreover, cAMP‐mediated Rac 1 activation was reduced to ~50% of WT levels and both PKA inhibition by PKI as well as PKA anchoring via A kinase anchoring peptides (AKAPs) by HT31 almost completely abolished Rac 1 activation in VASP (‐/‐) and WT endothelium. Accordingly, HT31 significantly reduced F/R‐mediated TER increase in WT cells and completely blocked the protective effect of cAMP on endothelial barrier properties. Our data underline the significant role of cAMP‐mediated Rac 1 activation for endothelial barrier stabilization and demonstrate that both AKAP‐mediated PKA anchoring and VASP are required for this process.