Submitted on July 7, 2006
Accepted on February 7, 2007
The Antithrombotic Activity of Kininogen is Mediated by Inhibitory Effects of Domain 3 During Arterial Injury In Vivo
Sarmina Hassan1, Irma M. Sainz1, Mohammad M. Khan1, Harland N Bradford1, Irma Isordia-Salas1, Sakeem W Kashem1, R. Balfour Sartor2, and Robert W Colman3*
1 Thrombosis, Temple University, philadelphia, Pennsylvania, United States
2 Microbiology and Immunology, University of North Carolina, Chapel Hill, North Carolina, United States
3 Thrombosis Research Center, Temple University Sch Medicine, Philadelphia, Pennsylvania, United States
* To whom correspondence should be addressed. E-mail: colmanr{at}temple.edu.
High molecular weight kininogen (HK) and its domain 3 (D3) exhibit anticoagulant properties and inhibit platelet activation at low thrombin concentration in vitro. We hypothesized that the rapid occlusive thrombosis in HK-deficient (HKd) rats, following endothelial injury of the aorta, results from enhanced platelet aggregation by thrombin. The effects of D3 (G235-M357) or D3-derived peptides on thrombosis in vivo were tested. D3 and its exon 7C terminal peptide (E7CP, K270-Q292), expressed as glutathione-S-transferase fusion proteins (GST-D3, GST-E7CP) or GST alone, as well as cleaved HK (HKa) or synthetic peptide E7CP, were infused intravenously 10 minutes before endothelial injury. The blood flow was reduced down to 10% of baseline flow within 28 ± 5.2 minutes by a platelet-fibrin thrombus in GST-treated HKd rats compared with >240 minutes in GST-treated HK-normal rats (wild type). GST-D3, GST-E7CP, HKa or E7CP infusion prolonged the flow time to 233, >240, 223 and >240 minutes respectively in HKd rats. When GST-E7CP was infused 10 minutes after the injury, the blood flow was maintained for >240 minutes. Thrombin-antithrombin concentrations were elevated by injury in HKd rats receiving GST, from 35 to 55 µg/L and decreased with GST-E7CP, HKa or E7CP reconstitution to 40, 15 and 9 µg/L respectively. We conclude that HKd rats are prothrombotic and HKa, kininogen D3, and its fragment E7CP, modulate arterial thrombosis after endothelial injury.
