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(Arteriosclerosis, Thrombosis, and Vascular Biology. 2005;25:2222.)
© 2005 American Heart Association, Inc.

Thrombosis

Endothelial Cell Activation by IL-1ß in the Presence of Fibrinogen Requires _Vß₃

Abha Sahni; Sanjeev K. Sahni; Charles W. Francis

From Hematology/Oncology Unit, Department of Medicine, University of Rochester School of Medicine & Dentistry, Rochester, NY.

Correspondence to Abha Sahni, PhD, Department of Medicine, P.O. Box 610, University of Rochester Medical Center, 601 Elmwood Ave, Rochester, NY 14642. E-mail Abha_Sahni{at}urmc.rochester.edu

Objective— The purpose of this study was to investigate the receptor requirements for enhanced IL-1ß–induced secretion of nitric oxide (NO) by endothelial cells (ECs) in the presence of fibrinogen.

Methods and Results— ECs were exposed to IL-1ß with or without fibrinogen and NO was measured as nitrite. NO production by EC exposed to fibrinogen (0.3±0.1 µmol/L) was comparable concentration to control (0.2±0.1 µmol/L), but IL-1ß significantly increased NO production (0.8±0.1 µmol/L), and the combination of both fibrinogen and IL-1ß resulted in a further increase to 2.2±0.2 µmol/L (P<0.002). 7E3 or LM609, antibodies to _vß₃, inhibited NO production stimulated by fibrinogen-bound IL-1ß 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-1ß (P<0.002). EC or fibroblasts exposed to both fibrinogen and IL-1ß, but not vitronectin and IL-1ß, demonstrated positive Western blotting for _Vß₃ after immunopurification with anti- IL-1R, indicating specific association between _vß₃ and IL-1R. Dual immunofluorescence also revealed colocalization of _vß₃ and IL-1R only when the cells were exposed to both fibrinogen and IL-1ß.

Conclusion— The enhanced NO production by ECs in the presence of fibrinogen-bound IL-1ß requires the coordinated effects of colocalized _Vß₃ and IL-1R.

The ability of fibrinogen-bound IL-1ß to stimulate NO secretion by ECs is blocked by anti-_vß₃. Also, fibrinogen-bound IL-1ß promotes the specific association of _vß₃ with IL-1R, as shown by coimmunoprecipitation or immunofluorescence staining. Fibrinogen binding enhances IL-1ß–induced secretion of NO through the coordinated effects of colocalized _vß₃ and IL-1R.

Key Words: endothelial cells • fibrinogen • IL-1ß • nitric oxide