Added Value of Blood Cells in Thrombin Generation Testing

Wan, Jun; Konings, Joke; Laat, Bas de; Hackeng, Tilman M.; Roest, Mark

doi:10.1055/a-1450-8300

Cited by 18 publications

(17 citation statements)

References 152 publications

(285 reference statements)

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“…The TG assay was originally designed as a global test of hemostasis, including most of the relevant coagulation factors present in the blood. Additional methods have been developed for the TG assay, to determine the role of platelet count and function by measuring TG in platelet rich plasma ( 35 – 37 ), and to determine the added effect of other blood cells and components by measuring TG in whole blood ( 38 – 40 ). Moreover, thrombomodulin can be added to the TG assay to investigate the function of the anticoagulant activated protein C pathway ( 41 – 43 ), which causes the increased risk of thrombosis in women using oral contraceptives ( 21 , 44 ).…”

Section: Discussionmentioning

confidence: 99%

Assessing the individual roles of FII, FV, and FX activity in the thrombin generation process

Bai

Konings

Ninivaggi

et al. 2022

Front. Cardiovasc. Med.

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Thrombin generation (TG) is known as a physiological approach to assess the hemostatic function. Although it correlates well with thrombosis and bleeding, in the current setup it is not sensitive to the effects of fluctuations in single coagulation factors. We optimized the calibrated automated thrombinography (CAT) method to quantify FII, FV and FX activity within the coagulation system. The CAT assay was fine-tuned for the assessment of FII, FV and FX by diluting the samples in FII-, FV-, or FX-deficient plasma, respectively, and measuring TG. Plasma FII levels correlated linearly with the ETP up to a plasma concentration of 100% FII. FV and FX levels correlated linearly with the peak height up to a plasma level of 2.5% FV and 10% FX, respectively. Sensitized CAT protocols were designed by adding a fixed volume of a pre-diluted patient sample to FII, FV, and FX deficient plasma in TG experiments. This approach makes the TG measurement dependent on the activity of the respective coagulation factor. The ETP or peak height were quantified as readouts for the coagulation factor activity. The intra- and inter-assay variation coefficients varied from 5.0 to 8.6%, and from 3.5 to 5.9%, respectively. Reference values were determined in 120 healthy subjects and the assays were clinically validated in 60 patients undergoing coronary artery bypass grafting (CABG). The sensitized CAT assays revealed that the contribution of FII, FV, and FX to the TG process was reduced after CABG surgery, leading to reduced prothrombin conversion and subsequently, lower TG.

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Section: Discussionmentioning

confidence: 99%

Assessing the individual roles of FII, FV, and FX activity in the thrombin generation process

Bai

Konings

Ninivaggi

et al. 2022

Front. Cardiovasc. Med.

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“…The formation of this complex activates the cleavage of prothrombin to thrombin and fragment 1.2. Excessive and unwanted presence of PS exposure will lead to a prothrombotic state, 10 and lack of a proper removal of PS exposing cells can result in uncontrolled and persistent presentation of self-antigens to the immune system. 11 sPLA2 plays an important role in inflammation.…”

Section: Plasma Membrane Phospholipidsmentioning

confidence: 99%

Hyperinflammation, apoptosis, and organ damage

Kuypers

2022

Exp Biol Med (Maywood)

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The cytokine storm (CS) in hyperinflammation is characterized by high levels of cytokines, extreme activation of innate as well as adaptive immune cells and initiation of apoptosis. High levels of apoptotic cells overwhelm the proper recognition and removal system of these cells. Phosphatidylserine on the apoptotic cell surface, which normally provides a recognition signal for removal, becomes a target for hemostatic proteins and secretory phospholipase A2. The dysregulation of these normal pathways in hemostasis and the inflammasome result in a prothrombotic state, cellular death, and end-organ damage. In this review, we provide the argument that this imbalance in recognition and removal is a common denominator regardless of the inflammatory trigger. The complex reaction of the immune defense system in hyperinflammation leads to self-inflicted damage. This common endpoint may provide additional options to monitor the progression of the inflammatory syndrome, predict severity, and may add to possible treatment strategies.

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“…By adding plasma, the platelet concentrates were diluted to a concentration of 4 × 10 8 platelets/mL (PRP). Since thrombin generation can be initiated by both the extrinsic (factor VII) and intrinsic (factor XII) pathways [ 20 ], factor XIIa inhibitor (corn trypsin inhibitor, Santa Cruz Biotechnology, Heidelberg, Germany) (50 µg/mL PRP) was added. Afterwards, PRP was transferred to a black 96-well plate (ThermoFisher Scientific Inc., Waltham, MA, USA) (40 µL/well) and treated with DPBS as a negative control, recombinant TF as a positive control, or the respective breast cancer cells (1 × 10 5 tumor cells/mL).…”

Section: Methodsmentioning

confidence: 99%

The Heparan Sulfate Proteoglycan Syndecan-1 Triggers Breast Cancer Cell-Induced Coagulability by Induced Expression of Tissue Factor

Hassan

Bückreiß

Efing

et al. 2023

Cells

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Syndecan-1 (Sdc-1) upregulation is associated with poor prognosis in breast cancer. Sdc-1 knockdown results in reduced angiogenesis and the dysregulation of tissue factor (TF) pathway constituents. Here, we evaluate the regulatory mechanisms and functional consequences of the Sdc-1/TF-axis using Sdc-1 knockdown and overexpression approaches in MCF-7 and MDA-MB-231 breast cancer cells. Gene expression was analyzed by means of qPCR. Thrombin generation and cell migration were detected. Cell-cycle progression and apoptosis were investigated using flow cytometry. In MDA-MB-231 cells, IL6, IL8, VEGF, and IGFR-dependent signaling affected TF pathway expression depending on Sdc-1. Notably, Sdc-1 depletion and TF pathway inhibitor (TFPI) synergistically affected PTEN, MAPK, and STAT3 signaling. At the functional level, the antiproliferative and pro-apoptotic effects of TFPI depended on Sdc-1, whereas Sdc-1’s modulation of cell motility was not affected by TFPI. Sdc-1 overexpression in MCF-7 and MDA-MB-231 cells led to increased TF expression, inducing a procoagulative phenotype, as indicated by the activation of human platelets and increased thrombin formation. A novel understanding of the functional interplay between Sdc-1 and the TF pathway may be compatible with the classical co-receptor role of Sdc-1 in cytokine signaling. This opens up the possibility of a new functional understanding, with Sdc-1 fostering coagulation and platelet communication as the key to the hematogenous metastatic spread of breast cancer cells.

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Added Value of Blood Cells in Thrombin Generation Testing

Cited by 18 publications

References 152 publications

Assessing the individual roles of FII, FV, and FX activity in the thrombin generation process

Assessing the individual roles of FII, FV, and FX activity in the thrombin generation process

Hyperinflammation, apoptosis, and organ damage

The Heparan Sulfate Proteoglycan Syndecan-1 Triggers Breast Cancer Cell-Induced Coagulability by Induced Expression of Tissue Factor

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