Angiopoietin chemotactic activities on neutrophils are regulated by PI-3K activation

Angiopoietins (Ang1 and Ang2) modulate blood vessel integrity during the angiogenic process through the activation of tyrosine kinase receptor (Tie2). We recently detected Tie2 expression on nentrophils and reported that angiopoietins induce acute proinflammatory events including nentrophil beta(2)-integrin activation and their adhesion onto endothelial cells. Herein, we investigated the effect of angiopoietins on neutrophil migration and their capacity to modulate CXCL8/ IL-8 chemotactic properties. Using a Boyden chamber assay, we observed that Ang1 and Ang2 (up to 10 nM; 60 min) increased the migration of nentrophils, and the maximal effect was achieved at 1 nM (72% and 114% increase, respectively) as compared with untreated cells. Angiopoietins induce a rapid and transient Akt phosphorylation, and pretreatment of neutrophils with PI-3K inhibitors, wortmannin (100 nM) and LY294002 (500 nM), reduced Ang1-mediated nentrophil migration by 100% and 78% and Ang2 chemotactic activity by 100% and 71%, respectively. Treatment of nentrophils with CXCL8/IL-8 (up to 50 nM; 60 min) increased basal neutrophil migration by 257% at its optimal concentration (10 nM), and pretreatment of nentrophils with corresponding PI-3K inhibitors reduced CXCL8/IL-8 (I nM) chemotactic effect. Pretreatment of nentrophils with Ang1 or Ang2 (10 nM; 15 min) potentiated nentrophil migration induced by CXCLB/IL-8 (I or 10 nM; 60 min) by 263% and 238% and by 1771% and 164%, respectively. Finally, both angiopoietins showed a synergistic effect on the induction of Akt phosphorylation mediated by CXCL8/IL-8. In summary, our data demonstrate that angiopoietins increase nentrophil migration through PI-3K activation and can enhance proinflammatory activities of other cytokines.