Tissue factor-induced coagulation triggers platelet thrombus formation as efficiently as fibrillar collagen at arterial blood flow conditions.

Abstract

The relative importance of vessel wall tissue factor (TF) in initiating thrombogenesis is not well defined. In contrast , vessel wall collagens have been well documented as potent inducers of thrombus formation. We compared the potency of a human TF/phospholipid surface with that of a surface consisting of human type III collagen fibrils in triggering thrombus formation in native human blood at venous and arterial blood flow conditions. A commercial preparation, Thromborel S, was used as a source of human TF. Biochemical characterization of this preparation revealed small amounts of FVII, FIX, and FX proteins. Coagulant activity of these proteins was associated with the FVII protein only, although it was a very low activity. Studies with anti-TF antibodies in a one-stage clotting assay showed that the procoagulant activity of Thromborel was mainly a result of TF. The molar ratio of TF to phospholipid was 1:2 x 10 7. Thrombus formation in flowing nonanticoagulated human blood drawn directly from an antecubital vein was triggered by either Thromborel S or collagen fibrils coated on Thermanox coverslips in a parallel-plate perfusion chamber device. A 1:50 Thromborel S dilution gave maximal fibrin T issue factor (TF) initiates coagulation through its interaction with the plasma coagulation protein FVTI/FVIIa. 12 Activated FVII (FVIIa) bound to TF activates the coagulation proteins FX and FIX, 34 which ultimately results in thrombin generation, fibrin formation, and platelet activation and aggregation. The wall shear rate has a profound impact on TF/ FVIIa-dependent FX activation, 5-7 as well as on fibrin deposition and platelet thrombus formation. 815 The significance of TF for fibrin deposition and thrombus formation on subendothelium, 7-13 ' 16 stimulated endothe-lial cells, 1718 and the extracellular matrix of stimulated endothelium 15 has been investigated in nonanticoagu-lated human blood by various perfusion chamber devices. These experiments have revealed a significant decrease in fibrin deposition with increasing shear and a concomitant enhancement of platelet thrombus formation. Immobilized purified TF reconstituted in phospho-lipid vesicles and TF expressed on fibroblasts have also been exposed to purified FVII and FX in flowing deposition (90% surface coverage) at a wall shear rate of 100 s"'. However, pretreatment of the TF surface with a monoclonal anti-TF antibody reduced this fibrin deposition by 93% (P<.001). Thus, TF was essential for the procoagulant activity of the Thromborel S surface in this flow system also. At higher wall shear rates (650 and 2600 s" 1), less fibrin was deposited, but the platelet thrombus formation on the fibrin mesh increased dramatically. Platelet thrombus formation was elicited on top of the fibrin mesh; platelets adhered exclusively to the fibrin strands formed on the Thromborel S surface. However, while much less fibrin was deposited on the collagen (/ > <.001), the platelet-collagen adhesion was substantially higher than the platelet-fibrin adhesion on the Thromborel S ...

Redistribution and Hemostatic Action of Recombinant Activated Factor VII Associated with Platelets

“…22 Notably, one of the TF sources used was known to contain small amounts of FVII. 23 The present data indicate that rFVIIa is internalized by platelets in a dose-dependent manner both in healthy subjects and in patients with severe FVII deficiency. To our knowledge, there have not been previous reports of FVIIa inside platelets.…”

“…22 Of note, one of the TF preparations used in these studies was known to contain traces of FVII. 23 It was therefore hypothesized that platelets may be able to incorporate FVIIa or even TF-FVIIa complexes. Redistribution of rFVIIa into platelets could protect this factor from physiological clearance mechanisms and thus explain the prolonged hemostatic action of rFVIIa under some clinical conditions.…”

Antithrombotic Efficacy of the Vitamin K Antagonist Fluindione in a Human Ex Vivo Model of Arterial Thrombosis

“…14,15,17,26 Equine collagen (Collagen Reagent Horm, Nycomed) was spraycoated onto plastic coverslips to a final density of 0.5 g/cm 2 . TF, purified from human placenta (Thromborel, Behring), was diluted 1:133 in coating buffer (0.1 mol/L sodium carbonate, pH 9.5), and coverslips were incubated in 2 mL of the Thromborel dilution for 24 hours at 4°C.…”

“…Perfusion experiments were performed with a parallelplate perfusion chamber device at 37°C. 14,15,17,26 After blood samples were collected (see below), native blood was drawn directly from an antecubital vein of the volunteers through a 19-gauge infusion set (Ohmeda) over a collagen-coated coverslip positioned in the parallelplate perfusion chamber. The blood flow rate was maintained at 10 mL/min by a peristaltic roller pump (Multiperpex LKB, Pharmacia) placed distal to the chamber.…”

“…[13][14][15][16][17] In this model, native blood is drawn from volunteers through a parallel-plate chamber device, where it interacts with a thrombogenic surface in well-established blood flow conditions, mimicking wall shear rates encountered in moderately stenosed arteries (2600 s Ϫ1 ). Two relevant thrombogenic molecules, collagen and tissue factor (TF), which are present in atherosclerotic plaques and primarily responsible for thrombus formation in vivo, 18,19 are exposed to blood.…”