Thrombotic responses of endothelial outgrowth cells to protein-coated surfaces.

There is significant clinical need for viable small-diameter vascular grafts. While there are many graft biomaterials in development, few have been clinically successful. Evaluation of grafts with a clinically relevant model is needed to drive development.

Immobilization of actively thromboresistant assemblies on sterile blood-contacting surfaces.

Rapid one-step modification of thrombomodulin with alkylamine derivatives such as azide, biotin, and PEG is achieved using an evolved sortase (eSrtA) mutant. The feasibility of a point-of-care scheme is demonstrated herein to site-specifically immobilize azido-thrombomodulin on sterilized commercial ePTFE vascular grafts, which exhibit superior thromboresistance compared with commercial heparin-coated grafts in a primate model of acute graft thrombosis.

Reversal of rivaroxaban anticoagulation by haemostatic agents in rats and primates.

Rivaroxaban is an oral, direct factor Xa inhibitor for the management of thromboembolic disorders. Despite its short half-life, the ability to reverse rivaroxaban anticoagulation could be beneficial in life-threatening emergencies. The potential of prothrombin complex concentrate (PCC; Beriplex®), activated PCC (aPCC; FEIBA®) or recombinant activated factor VII (rFVIIa; NovoSeven®) to reverse rivaroxaban in rats and baboons was investigated.

Antithrombotic effect of antisense factor XI oligonucleotide treatment in primates.

Objective: During coagulation, factor IX (FIX) is activated by 2 distinct mechanisms mediated by the active proteases of either FVIIa or FXIa. Both coagulation factors may contribute to thrombosis; FXI, however, plays only a limited role in the arrest of bleeding. Therefore, therapeutic targeting of FXI may produce an antithrombotic effect with relatively low hemostatic risk.

Antithrombotic activity of protein S infused without activated protein C in a baboon thrombosis model.

Protein S (ProS) is an essential plasma protein that enhances the anticoagulant activity of activated protein C (APC). In vitro , purified native human Zn2+-containing ProS also exerts direct anticoagulant activity by inhibiting prothrombinase and extrinsic FXase activities independently of APC. We investigated antithrombotic effects of ProS infused without APC in a baboon shunt model of thrombogenesis that employs a device consisting of arterial and venous shear flow segments.

A biologically active surface enzyme assembly that attenuates thrombus formation.

Activation of hemostatic pathways by blood-contacting materials remains a major hurdle in the development of clinically durable artificial organs and implantable devices. We postulate that surface-induced thrombosis may be attenuated by the reconstitution onto blood contacting surfaces of bioactive enzymes that regulate the production of thrombin, a central mediator of both clotting and platelet activation cascades. Thrombomodulin (TM), a transmembrane protein expressed by endothelial cells, is an established negative regulator of thrombin generation in the circulatory system.

Safety and antithrombotic efficacy of moderate platelet count reduction by thrombopoietin inhibition in primates.

Most heart attacks and strokes are caused by blood clots (thrombi) that block the vasculature. Because disease-causing arterial thrombosis depends on blood platelets, platelet inhibitors such as aspirin and clopidogrel effectively decrease the risk of thrombosis; however, they also impair platelet-dependent hemostasis that staunches bleeding from wounds and can therefore produce excessive bleeding. Experimental studies show that a reduction in the number of platelets also inhibits thrombosis, but these treatments also interfere with platelet function.

Transport-reaction model of mural thrombogenesis: comparisons of mathematical model predictions and results from baboon models.

Thrombogenesis depends on biochemical reactions affected by blood flow dynamics. While mathematical models of mural thrombogenesis provide a means of understanding how blood flow affects thrombus growth, comparisons to experimental data are needed to validate the models and enable prediction of thrombus growth under diverse conditions. In this paper, we present mathematical models of mural thrombogenesis under flow and validation of the models with experimental data collected from a thrombogenic vascular graft segment.

Inhibition of acute vascular thrombosis in chimpanzees by an anti-human tissue factor antibody targeting the factor X binding site.

Tissue factor (TF) antagonists targeting the factor VII (FVII) binding domain have been shown to interrupt acute vascular thrombus formation without impairing haemostasis in non-human primates. In this study, we evaluate whether a human/mouse chimeric monoclonal antibody (ALT-836, formerly known as Sunol-cH36) blocking the factor X/factor IX (FX/FIX) binding site of tissue factor could achieve similar clinical benefits in an arterial thrombosis model induced by surgical endarterectomy in chimpanzees. In this model, sequential surgical endarterectomies on right and left superficial femoral arteries were performed 30 days apart in five chimpanzees.

Prevention of vascular graft occlusion and thrombus-associated thrombin generation by inhibition of factor XI.

The protease thrombin is required for normal hemostasis and pathologic thrombogenesis. Since the mechanism of coagulation factor XI (FXI)-dependent thrombus growth remains unclear, we investigated the contribution of FXI to thrombus formation in a primate thrombosis model. Pretreatment of baboons with a novel anti-human FXI monoclonal antibody (aXIMab; 2 mg/kg) inhibited plasma FXI by at least 99% for 10 days, and suppressed thrombin-antithrombin (TAT) complex and beta-thromboglobulin (betaTG) formation measured immediately downstream from thrombi forming within collagen-coated vascular grafts.

Capture of flowing endothelial cells using surface-immobilized anti-kinase insert domain receptor antibody.

In humans, self-endothelialization of synthetic grafts is severely limited, but a recent interesting idea is to attract endothelial progenitor cells (EPCs) from peripheral blood onto grafts via antibodies directed at proposed EPC markers. Results with anti-CD34 antibodies have shown some promise, but it is unclear whether CD34 is the best marker for cells with re-endothelializing potential. Much evidence points to kinase insert domain receptor (KDR) as an important indicator of endothelial potential if not a definitive marker.

Procoagulant properties of flow fields in stenotic and expansive orifices.

In the United States, over 125,000 mechanical heart valves (MHVs) are implanted each year. Flow through the MHV hinge can cause thromboemboli formation. The purpose of this study was to examine various orifice geometries representing the MHV hinge region and how these geometries may contribute to platelet activation and thrombin generation.

Fibrinogen gamma' chain carboxy terminal peptide selectively inhibits the intrinsic coagulation pathway.

The minor gammaA/gamma' isoform of fibrinogen contains a high affinity binding site for thrombin exosite II that is lacking in the major fibrinogen isoform, gammaA/gammaA fibrinogen. The biological consequences of gamma' chain binding to thrombin were therefore investigated. Coagulation assays, thrombin activity assays, and a primate thrombosis model were used to characterize the biological effects of the gamma' 410-427 peptide.

Thrombin formation in vitro in response to shear-induced activation of platelets.

Introduction: Thromboembolic events caused by implanted vascular devices present serious medical challenges. In particular bileaflet mechanical heart valves (MHVs) are prone to thrombus formation in the hinge region due to a combination of high shear stress and stagnation regions. Most studies of shear-induced platelet activation and aggregation have been performed using viscometers, parallel plate flow, and other non-physiologic in vitro configurations.

Relative antithrombotic and antihemostatic effects of protein C activator versus low-molecular-weight heparin in primates.

The anticoagulant and anti-inflammatory enzyme, activated protein C (APC), naturally controls thrombosis without affecting hemostasis. We therefore evaluated whether the integrity of primary hemostasis was preserved during limited pharmacological antithrombotic protein C activator (PCA) treatment in baboons. The double-mutant thrombin (Trp215Ala/Glu217Ala) with less than 1% procoagulant activity was used as a relatively selective PCA and compared with systemic anticoagulation by APC and low-molecular-weight heparin (LMWH) at doses that inhibited fibrin deposition on thrombogenic segments of arteriovenous shunts.

Flow and thrombosis at orifices simulating mechanical heart valve leakage regions.

Background: While it is established that mechanical heart valves (MHVs) damage blood elements during leakage and forward flow, the role in thrombus formation of platelet activation by high shear flow geometries remains unclear. In this study, continuously recalcified blood was used to measure the effects of blood flow through orifices, which model MHVs, on the generation of procoagulant thrombin and the resulting formation of thrombus. The contribution of platelets to this process was also assessed.

Hemostatic effect of activated factor VII without promotion of thrombus growth in melagatran-anticoagulated primates.

Introduction: Pharmacological enhancement of coagulation using activated prothrombin complex concentrate (APCC) or activated factor VII (FVIIa) might be useful hemostatic approaches to bleeding emergencies during anticoagulant therapy. However, any such intervention should not increase thrombotic risk. We therefore investigated their hemostatic and prothrombotic potential during propagation of large arterial-type thrombin in anticoagulated baboons.

Limited generation of activated protein C during infusion of the protein C activator thrombin analog W215A/E217A in primates.

Anticoagulation with activated protein C (APC) reduces the mortality of severe sepsis. We investigated whether the circulating protein C (PC) pool could be utilized for sustained anticoagulation by endogenous APC. To generate APC without procoagulant effects, we administered the anticoagulant thrombin mutant W215A;E217A (WE) to baboons.

Factor VIIa inhibitors: chemical optimization, preclinical pharmacokinetics, pharmacodynamics, and efficacy in an arterial baboon thrombosis model.

Highly selective and potent factor VIIa-tissue factor (fVIIa.TF) complex inhibitors were generated through structure-based design. The pharmacokinetic properties of an optimized analog (9) were characterized in several preclinical species, demonstrating pharmacokinetic characteristics suitable for once-a-day dosing in humans.

Soluble thrombomodulin is antithrombotic in the presence of neutralising antibodies to protein C and reduces circulating activated protein C levels in primates.

We studied whether there was a relationship between the anticoagulant effects of recombinant human soluble thrombomodulin (rhsTM) and activation of protein C in a primate model of acute vascular graft thrombosis in 11 baboons (Papio species). Baboons were pretreated with 0.1, 1 and 5 mg/kg of rhsTM, with or without co-injection of a neutralising monoclonal antibody to protein C (HPC4) in the 1 mg/kg rhsTM group.

A selective, slow binding inhibitor of factor VIIa binds to a nonstandard active site conformation and attenuates thrombus formation in vivo.

The serine protease factor VIIa (FVIIa) in complex with its cellular cofactor tissue factor (TF) initiates the blood coagulation reactions. TF.FVIIa is also implicated in thrombosis-related disorders and constitutes an appealing therapeutic target for treatment of cardiovascular diseases.

In vivo platelet redistribution and acute transient thrombocytopenia after eptifibatide injection in baboons.

Background: The occurrence of thrombocytopenia has been reported during clinical eptifibatide (Integrilin) therapy, but the exact mechanism is not yet established to explain the varied duration and severity of thrombocytopenia associated with glycoprotein (GP) IIb/IIIa inhibitors. We assessed the redistribution of platelets in juvenile baboons during acute transient thrombocytopenia that was observed after eptifibatide injection.

Methods: Eptifibatide was administered intravenously to eight baboons by infusion at 20 microg/kg/min or a bolus injection of 10 mg.

Platelet activation and secretion in patients with major depression, thoracic aortic atherosclerosis, or renal dialysis treatment.

Relatively little is known concerning the magnitude of alterations of platelet activation and secretion markers of patients with major depression when compared to patients at increased risk for, or with current, clinically significant atherosclerosis. Markers of in vivo platelet stimulation and secretion were measured under basal conditions in normal comparison subjects (n = 12) and three patient groups: patients diagnosed with DSM-IV major depression (n = 15), dialysis-dependent patients (n = 12), and patients with severe thoracic aortic atherosclerosis (n = 10). In comparison to normal comparison subjects, depressed patients and patients with thoracic aortic atherosclerosis exhibited the greatest platelet stimulation as detected by increased anti-LIBS platelet binding.

Bileaflet aortic valve prosthesis pivot geometry influences platelet secretion and anionic phospholipid exposure.

The clinical histories of the Medtronic Parallel (MP) and St. Jude Medical (SJM) Standard valves suggest pivot geometry influences the thrombogenic characteristics of bileaflet prostheses. This work studied the effects of various pivot geometries on markers of platelet damage in a controlled, in vitro apparatus.

The sensitivity of indicators of thrombosis initiation to a bileaflet prosthesis leakage stimulus.

Background And Aim Of The Study: The recent clinical history and experimental studies of the Medtronic Parallel (MP) valve suggest that bileaflet valve leakage flow is a primary initiator of thrombosis. These studies investigated the effects of physiologic leakage flow through a MP valve on various markers of blood damage.

Methods: A centrifugal pump was used to drive whole, human blood anticoagulated with PPACK through a circuit containing a MP 27 mm valve in the closed position (experimental runs) or a MP 27 mm valve in the open position (control runs).