Abstract 40: Neutrophil Serine Protease PR3 Promotes Platelet Adhesion and Activation on von Willebrand Factor (vWF) Expressing Endothelial Cells
Introduction Neutrophils play important roles in inflammation, regulating vascular permeability and promoting thrombus formation. Upon activation, neutrophils release serine proteases that can activate coagulation factors and protease activated receptors (PARs). Proteinase 3 (PR3) is the most abundant serine protease in neutrophils; however, the role of PR3 in regulating vascular function and thrombosis is not clear. In this study we have used flow chamber assays and confocal microscopy to investigate PR3 activation of endothelial cells through PAR-2 and the recruitment and activation of platelets and leukocytes under shear conditions.
Results After endothelial cells were incubated with PR3 or a PAR-2 activating peptide, we observed significant release and surface presentation of von Willebrand (vWF) compared to untreated cells. In addition, we also found that PR3 increased endothelial cell adhesion molecule expression and ERK phosphorylation. Using flow chamber assays, we observed that PR3-treated endothelial cells expressing vWF were able to capture platelets from whole blood under venous shear conditions; however, blocking antibodies against either platelet GpIb or GpIIb/IIIa significantly reduced platelet adhesion. Using whole blood, we also found that significant numbers of leukocytes were recruited to PR3-treated endothelial cells. However, in platelet-depleted whole blood, or blood treated with antibodies against GpIb or GpIIb/IIIa, we observed a dramatic reduction in leukocyte adhesion. Therefore, leukocyte recruitment to PR3-treated endothelial cells required platelet activation and adhesion and this was mediated in part by vWF.
Conclusions Neutrophil PR3 can promote platelet adhesion to the vascular endothelium through a process requiring endothelial cell vWF, platelet GpIb and GpIIb/IIIa. PR3 also promotes leukocyte recruitment to the endothelium through a platelet-dependant process. This data demonstrates that PR3 can be a significant contributor to thrombus formation and leukocyte recruitment to the vascular endothelium and PR3 may be an important target for disrupting thrombus formation in vivo.
- © 2013 by American Heart Association, Inc.