Acetylated Prothrombin as a Substrate in the Measurement of the Procoagulant Activity of Platelets: Elimination of the Feedback Activation of Platelets by Thrombin

Jesty, Jolyon; Bluestein, Danny

doi:10.1006/abio.1999.4148

Cited by 100 publications

(152 citation statements)

References 25 publications

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“…16 The prothrombinase complex assembles on the activated platelet surface in the presence of factor Xa bound to the two essential cofactors provided by activated platelets: factor Va and anionic phospholipids. 14 The thrombin, which is generated in the assay, is a powerful platelet agonist that reactivates the platelets in a positive feedback loop.…”

Section: In Vitro Platelet Activation State (Pas) Assay Measurementsmentioning

confidence: 99%

DPIV Prediction of Flow Induced Platelet Activation—Comparison to Numerical Predictions

et al. 2007

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Abstract-Flow induced platelet activation (PA) can lead to platelet aggregation, deposition onto the blood vessel wall, and thrombus formation. PA was thoroughly studied under unidirectional flow conditions. However, in regions of complex flow, where the platelet is exposed to varying levels of shear stress for varying durations, the relationship between flow and PA is not well understood. Numerical models were developed for studying flow induced PA resulting from stress histories along Lagrangian trajectories in the flow field. However, experimental validation techniques such as Digital Particle Image Velocimetry (DPIV) were not extended to include such models. In this study, a general experimental tool for PA analysis by means of continuous DPIV was utilized and compared to numerical simulation in a model of coronary stenosis. A scaled up (5:1) 84% eccentric and axisymetric coronary stenosis model was used for analysis of shear stress and exposure time along particle trajectories. Flow induced PA was measured using the PA State (PAS) assay. An algorithm for computing the PA level in pertinent trajectories was developed as a tool for extracting information from DPIV measurements for predicting the flow induced thrombogenic potential. CFD, DPIV and PAS assay results agreed well in predicting the level of PA. In addition, the same trend predicted by the DPIV was measured in vitro using the Platelet Activity State (PAS) assay, namely, that the symmetric stenosis activated the platelets more as compared to the eccentric stenosis.

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Section: In Vitro Platelet Activation State (Pas) Assay Measurementsmentioning

confidence: 99%

DPIV Prediction of Flow Induced Platelet Activation—Comparison to Numerical Predictions

et al. 2007

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“…Purified GFPs were prepared from whole blood as described in Sheriff et al (6) (Fig. 1) and diluted to a standard concentration of 20 000 platelets/μL in a HEPES-modified Ca 2+ -free Tyrode's buffer (Platelet Buffer) containing 0.1% fatty acid-free bovine serum albumin, with 3 mM Ca 2+ added 10 min prior to experiments (7,22). Equilibrated GFPs were maintained with gentle agitation at room temperature and used for flow loop experiments within 6 h of gel filtration.…”

Section: Preparation Of Platelet Samplesmentioning

confidence: 99%

“…The PAS assay is a laboratory method able to measure the near real-time rate of formation of thrombin-a universal thrombogenic marker-by activated platelets and to correlate with their activation level (6). The test is performed with gel-filtered platelets (GFPs, i.e., platelets deprived of plasma proteins), and the assay makes use of acetylated prothrombin (Ac-FIIa) as the thrombin substrate (7). The PAS assay allows measuring the prothrombotic tendency of activated platelets, that is, the platelet procoagulant activity, by detecting the thrombin production rate of activated platelets, without inducing aggregation or clot formation during the test.…”

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confidence: 99%

“…Mechanisms leading to platelet aggregation are precluded during the test, mainly due to the removal of plasma coagulation factors (mediating aggregation in whole blood) from GFP preparations (7). Furthermore, Ac-FIIa, while retaining the ability to produce thrombin upon activation by GFP-bound prothrombinase complex, is characterized by low (<0.1%) clotting activity (7). Moreover, the thrombin produced by Ac-FIIa is inactive on fibrinogen (7) and thus cannot mediate the conversion of fibrinogen to fibrin.…”

mentioning

confidence: 99%

“…In addition, the use of Ac-FIIa allows inhibiting the physiologic thrombinmediated positive feedback on PA. In detail, the thrombin produced by the conversion of Ac-FIIa does not feed back on the factor Xa (FXa) complex to further activate platelets (7). Hence, with PAS assay, the dose-and time-PA dynamics can be characterized, in response to differing levels (i.e., varying the dose and/or the exposure time) of the agonist, ensuring a one-to-one relationship between the inducer and the actual level of PA, which is proportional to the thrombin production rate.…”

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On the Use of the Platelet Activity State Assay for the In Vitro Quantification of Platelet Activation in Blood Recirculating Devices for Extracorporeal Circulation

et al. 2016

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Abstract:We designed an experimental setup to characterize the thrombogenic potential associated with blood recirculating devices (BRDs) used in extracorporeal circulation (ECC). Our methodology relies on in vitro flow loop platelet recirculation experiments combined with the modified-prothrombinase platelet activity state (PAS) assay to quantify the bulk thrombin production rate of circulated platelets, which correlates to the platelet activation (PA) level. The method was applied to a commercial neonatal hollow fiber membrane oxygenator. In analogous hemodynamic environment, we compared the PA level resulting from multiple passes of platelets within devices provided with phosphorylcholine (PC)-coated and noncoated (NC) fibers to account for flow-related mechanical factors (i.e., fluid-induced shear stress) together with surface contact activation phenomena. We report for the first time that PAS assay is not significantly sensitive to the effect of material coating under clinically pertinent flow conditions (500 mL/min), while providing straightforward information on shear-mediated PA dynamics in ECC devices. Being that the latter is intimately dependent on local flow dynamics, according to our results, the rate of thrombin production as measured by the PAS assay is a valuable biochemical marker of the selective contribution of PA in BRDs induced by device design features. Thus, we recommend the use of PAS assay as a means of evaluating the effect of modification of specific device geometrical features and/or different design solutions for developing ECC devices providing flow conditions with reduced thrombogenic impact.

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Thrombelastography and Other Novel Techniques

Tantry

Doraiswamy

Stokken

et al. 2014

Antiplatelet Therapy in Cardiovascular Disease

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Acetylated Prothrombin as a Substrate in the Measurement of the Procoagulant Activity of Platelets: Elimination of the Feedback Activation of Platelets by Thrombin

Cited by 100 publications

References 25 publications

DPIV Prediction of Flow Induced Platelet Activation—Comparison to Numerical Predictions

DPIV Prediction of Flow Induced Platelet Activation—Comparison to Numerical Predictions

On the Use of the Platelet Activity State Assay for the In Vitro Quantification of Platelet Activation in Blood Recirculating Devices for Extracorporeal Circulation

Thrombelastography and Other Novel Techniques

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