Antithrombotic activity of kininogen is mediated by inhibitory effects of domain 3 during arterial injury in vivo.
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 (GST) fusion proteins (GST-D3, GST-E7CP), or GST alone, as well as cleaved HK (HKa) or synthetic peptide E7CP, were infused intravenously 10 min before endothelial injury. Blood flow was reduced down to 10% of baseline flow within 28 +/- 5.2 min by a platelet-fibrin thrombus in GST-treated HKd rats compared with >240 min in GST-treated normal HK rats (wild type). GST-D3, GST-E7CP, HKa, or E7CP infusion prolonged the flow time to 233, >240, 223, and >240 min, respectively, in HKd rats. When GST-E7CP was infused 10 min after the injury, blood flow was maintained for >240 min. Thrombin-antithrombin concentrations were elevated by injury in HKd rats receiving GST from 35 to 55 microg/l and decreased with GST-E7CP, HKa, or E7CP reconstitution to 40, 15, and 9 microg/l, respectively. We conclude that HKd rats are prothrombotic and that HKa, kininogen D3, and its fragment E7CP modulate arterial thrombosis after endothelial injury.
Hassan
Sarmina
S
Sol Sherry Thrombosis Research Center, Temple Univeristy School of Medicine, 3400 N. Broad St., Rm. 418 OMS, Philadelphia, PA 19140, USA.
Sainz
Irma M
IM
Khan
Mohammad M
MM
Bradford
Harlan N
HN
Isordia-Salas
Irma
I
Kashem
Sakeen W
SW
Sartor
R Balfour
RB
Colman
Robert W
RW
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D000990:Antithrombin III; D001011:Aorta / Q000187:drug effects / Q000293:injuries / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D001783:Blood Flow Velocity; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000187:drug effects / Q000293:injuries / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D005337:Fibrin / Q000378:metabolism; D005343:Fibrinolytic Agents / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology / Q000627:therapeutic use; D005982:Glutathione Transferase / Q000235:genetics; D019679:Kininogen, High-Molecular-Weight / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology / Q000627:therapeutic use; D008297:Male; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D010447:Peptide Hydrolases / Q000097:blood; D010974:Platelet Aggregation; D017434:Protein Structure, Tertiary; D051381:Rats; D011917:Rats, Inbred Lew / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012039:Regional Blood Flow; D013917:Thrombin / Q000378:metabolism; D013927:Thrombosis / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology / Q000517:prevention & control
D005343:Fibrinolytic Agents; D019679:Kininogen, High-Molecular-Weight; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C046193:antithrombin III-protease complex; D000990:Antithrombin III; D005337:Fibrin; D005982:Glutathione Transferase; D010447:Peptide Hydrolases; D013917:Thrombin