Endothelial cell activation by IL-1β in the presence of fibrinogen requires α_vβ₃

Objective - The purpose of this study was to investigate the receptor requirements for enhanced IL-1β-induced secretion of nitric oxide (NO) by endothelial cells (ECs) in the presence of fibrinogen. Methods and Results - ECs were exposed to IL-1β with or without fibrinogen and NO was measured as nitrite. NO production by EC exposed to fibrinogen (0.3plusmn;0.1 μmol/L) was comparable concentration to control (0.2±0.1 μmol/L), but IL-1β significantly increased NO production (0.8±0.1 μmol/L), and the combination of both fibrinogen and IL-1β resulted in a further increase to 2.2±0.2 μmol/L (P<0.002). 7E3 or LM609, antibodies to α_vβ₃, inhibited NO production stimulated by fibrinogen-bound IL-1β to 0.2±0.1 μmol/L (P<0.001) or 0.2±0.03 μmol/L (P<0.0001), respectively. These levels were comparable to control and significantly less than with IL-1β (P<0.002). EC or fibroblasts exposed to both fibrinogen and IL-1β, but not vitronectin and IL-1β, demonstrated positive Western blotting for α _vβ₃ after immunopurification with and- IL-1R, indicating specific association between α_vβ₃ and IL-1R. Dual immunofluorescence also revealed colocalization of α_vβ₃ and IL-1R only when the cells were exposed to both fibrinogen and IL-1β. Conclusion - The enhanced NO production by ECs in the presence of fibrinogen-bound IL-1β requires the coordinated effects of colocalized α_vβ₃ and IL-1R.