Recent research suggests activation of protease activated receptors (PAR) on the surface of endothelial and epithelial cells may play a role in general mechanisms of inflammation. We hypothesize that mast cell tryptase activation of endothelial cell PAR-2 is coupled to increased calcium-independent phospholipase A₂ (iPLA₂) activity and increased platelet activating factor (PAF) production that may play a role in inflammatory cell recruitment at sites of vascular injury. Stimulation of human coronary artery endothelial cells (HCAEC) with 20 ng/ml tryptase increased iPLA₂ activity, arachidonic acid release and PAF production. These tryptase-stimulated responses were inhibited by pretreatment with the iPLA₂-selective inhibitor bromoenol lactone (BEL, 5 µM, 10 min). Similar patterns of increased iPLA₂ activity and PAF production were also seen when HCAEC were treated with SLIGKV, which represents the tethered ligand sequence for the human PAR-2 once the receptor is cleaved by tryptase. Tryptase stimulation also increased cell surface expression of P-selectin, decreased electrical resistance and increased neutrophil adherence to the endothelial cell monolayer. The tryptase stimulated increase in both cell surface P-selectin expression and neutrophil adhesion were also inhibited with BEL pretreatment. We conclude that tryptase stimulation of HCAEC contributes importantly to early inflammatory events following vascular injury by activation of iPLA₂, leading to arachidonic acid release, PAF production, cell surface P-selectin expression and increased neutrophil adherence.