Abstract
Interactions between leukocytes and vascular endothelial cells are mediated by a complex set of membrane adhesion molecules which transduce bi-directional signals in both cell types. Endothelium of the cerebral blood vessels, which constitute the blood–brain barrier, strictly controls adhesion and trafficking of leukocytes into the brain. Investigating signaling pathways triggered by the engagement of adhesion molecules expressed on brain endothelial cells, we previously documented the role of ICAM-1 in activation of the tyrosine phosphorylation of several actin-binding proteins and subsequent rearrangements of the actin cytoskeleton. In the present study, we show that, whereas PECAM-1 is known to control positively the trans-endothelial migration of leukocytes via homophilic interactions between leukocytes and endothelial cells, PECAM-1 engagement on brain endothelial surface unexpectedly counteracts the ICAM-1-induced tyrosine phosphorylation of cortactin and rearrangements of the actin cytoskeleton. We present evidence that the PECAM-1-associated tyrosine phosphatase SHP-2 is required for ICAM-1 signaling, suggesting that its activity might crucially contribute to the regulation of ICAM-1 signaling by PECAM-1. Our findings reveal a novel activity for PECAM-1 which, by counteracting ICAM-1-induced activation, could directly contribute to limit activation and maintain integrity of brain vascular endothelium.
Highlights
These adhesion molecules have been well documented as signal transducers in leukocytes and endothelial cells, in as much as leukocyte adhesion to endothelial cells as well as antibody cross-linking were shown to activate multiple signaling pathways in both cell types
platelet/endothelial cell-adhesion molecule-1 (PECAM-1) cross-linking inhibits intercellular adhesion molecule-1 (ICAM-1)-induced actin cytoskeleton rearrangements In order to assess the existence of a putative cross-talk between the signaling pathways coupled to ICAM-1 and PECAM-1 in brain endothelial cells, we developed a procedure of sequential antibody cross-linking of ICAM-1 and PECAM-1 to mimic the supposed sequence of events supporting leukocyte extravasation
We show in the present study that PECAM-1 engagement by antibody cross-linking totally blocked ICAM-1-induced activation of RhoA and subsequent changes in actin cytoskeleton, as well as tyrosine phosphorylation of cortactin in rat brain endothelial cells
Summary
These adhesion molecules have been well documented as signal transducers in leukocytes and endothelial cells, in as much as leukocyte adhesion to endothelial cells as well as antibody cross-linking were shown to activate multiple signaling pathways in both cell types. No evidence to our knowledge has emerged on how the two activated signaling pathways coupled to ICAM-1 and PECAM-1 are integrated by endothelial cells and to what extent they might contribute in a sequential and coordinated manner to the endothelial response to leukocyte adhesion. We addressed the question of a putative cross-talk between these two signaling pathways by sequential antibody cross-linking of ICAM-1 and PECAM-1 at the surface of endothelial cells: this experimental approach has been shown by us and others to mimic leukocyte interaction with endothelial cells and to allow the biochemical analysis of endothelial response to leukocyte adhesion. The rat brain endothelial cell line RBE4 was used here as a robust model of brain microvascular endothelium (Schweitzer et al 1997; Hoffmann et al 2001); We report in the present study that PECAM-1 engagement unexpectedly down-regulated ICAM-1-induced tyrosine phosphorylation of cortactin and rearrangements of the actin cytoskeleton. The functional relevance of this finding is discussed in terms of regulation of BBB integrity in inflammatory situations
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