MIP-2 causes differential activation of RhoA in mouse aortic versus pulmonary artery endothelial cells.

Previously, we have shown that endothelial cell chemotaxis to the proangiogenic chemokine MIP-2 (macrophage inflammatory protein-2) is much greater in mouse aortic endothelial cells (EC) than pulmonary arterial endothelial cells (PA EC). This was true despite the observation that both cell types display comparable levels of the ligand receptor, CXCR(2) (8). Since the systemic arterial circulation is proangiogenic in the adult lung and the pulmonary circulation is relatively resistant to neovascularization, we questioned whether the observed functional heterogeneity is related to inherent differences in cell signaling cascades of the two EC subtypes. Specifically, we measured activation of Rac1 and RhoA, both thought to be involved in EC cell migration. Rac1 showed inconsistent and minimal changes in both cell types after MIP-2 treatment (p>0.05). However, activated RhoA was increased upon exposure to MIP-2 only in aortic EC (61% increase; p<0.05). Decreased RhoA activation after treatment of aortic EC with specific siRNA for RhoA resulted in a functional decrease in EC chemotaxis to MIP-2 (17% increase; p<0.05). Additionally, increased RhoA activation in PA EC with adenoviral infection of RhoA caused an increase in PA EC chemotaxis to MIP-2 (46% increase; p<0.05). Inhibition of RhoA activity with the Rho kinase inhibitor, Y27632, blocked aortic EC chemotaxis and stress fiber formation. Thus, RhoA activation is increased after MIP-2 treatment in mouse aortic endothelial cells but not in pulmonary artery endothelial cells. We conclude that RhoA is part of a signaling pathway essential for aortic cell migration after CXCR(2) ligation. This result provides one explanation for the difference in chemotaxis observed in these two endothelial subtypes that express similar levels of CXCR(2).