The In Vitro Analysis of the Coagulation Mechanism of Activated Factor VII Using Thrombelastogram

2002 ◽  
Vol 88 (11) ◽  
pp. 768-772 ◽  
Author(s):  
Chiharu Kawaguchi ◽  
Yae Hanesaka ◽  
Akira Yoshioka ◽  
Yukihiro Takahashi
Keyword(s):  
Platelet Rich Plasma ◽  
Coagulation Factor ◽  
Factor Vii ◽  
Factor V ◽  
Coagulation Factor Vii ◽  
Activated Factor Vii ◽  
Coagulation Mechanism ◽  
Vitro Analysis ◽  
Hemostatic Effects

SummaryWe investigated the effects of addition of recombinant activated coagulation factor VII (rFVIIa) to coagulation factor-deficient plasma and whole blood, using thrombelastograms (TEGs). The addition of rFVIIa to factor II-or X-deficient plasma did not correct hemostatic parameters, whereas it produced partial responses in factor V-, VIII-or IX-deficient plasma and good responses in factor VII-, XI-or XII-deficient plasma. Furthermore, the addition of rFVIIa and platelets (30-100 X 103/µl) to platelet-poor plasma produced marked corrections, producing TEGs similar to those of platelet-rich plasma. These results indicate that factors II and X are essential for the hemostatic effects of rFVIIa, and that factors V and VIII promote these effects. We believe that TEGs are, at present, one of the most useful tools for evaluating in vitro hemostatic effects of rFVIIa.

The Effect of Dextran of Various Molecular Weight on the Coagulation in Dogs

1961 ◽  
Vol 06 (01) ◽  
pp. 015-024 ◽  
Author(s):  
Sven Erik Bergentz ◽  
Oddvar Eiken ◽  
Inga Marie Nilsson
Keyword(s):  
Molecular Weight ◽  
Body Weight ◽  
High Molecular Weight ◽  
Bleeding Time ◽  
Factor Vii ◽  
Low Molecular Weight ◽  
Factor V ◽  
Coagulation Mechanism ◽  
Coagulation Defects

Summary1. Infusions of low molecular weight dextran (Mw = 42 000) to dogs in doses of 1—1.5 g per kg body weight did not produce any significant changes in the coagulation mechanism.2. Infusions of high molecular weight dextran (Mw = 1 000 000) to dogs in doses of 1—1.5 g per kg body weight produced severe defects in the coagulation mechanism, namely prolongation of bleeding time and coagulation time, thrombocytopenia, pathological prothrombin consumption, decrease of fibrinogen, prothrombin and factor VII, factor V and AHG.3. Heparin treatment of the dogs was found to prevent the decrease of fibrinogen, prothrombin and factor VII, and factor V otherwise occurring after injection of high molecular weight dextran. Thrombocytopenia was not prevented.4. In in vitro experiments an interaction between fibrinogen and dextran of high and low molecular weight was found to take place in systems comprising pure fibrinogen. No such interaction occurred in the presence of plasma.5. It is concluded that the coagulation defects induced by infusions of high molecular weight dextran are due to intravascular coagulation.

Elevated Levels of Factor VII Activity in the Postprandial State: Effect of the Factor VII Arg-GIn Polymorphism

1994 ◽  
Vol 72 (05) ◽  
pp. 734-739 ◽  
Author(s):  
Angela Silveira ◽  
Fiona Green ◽  
Fredrik Karpe ◽  
Margareta Blombäck ◽  
Steve Humphries ◽  
...  
Keyword(s):  
Coagulation Factor ◽  
Plasma Lipoproteins ◽  
Factor Vii ◽  
Percentage Increase ◽  
Fat Intake ◽  
Postprandial Lipaemia ◽  
Coagulation Factor Vii ◽  
Postprandial State ◽  
Activated Factor Vii ◽  
Before And After

SummaryA genetic polymorphism (Arg/Gln353) of coagulation factor VII was recently identified and shown to be associated with differences in basal factor VII coagulant activity. Postprandial lipaemia seems to exert an acute but evanescent effect on the activity of factor VII, and the influence of the Arg/Gln353 polymorphism on factor VII activation during postprandial lipaemia was therefore studied in male post-infarction patients [age 48.8 ± 3.3 years (mean ± SD)] with Arg/Arg (n = 23) and Arg/Gln (n = 8) genotypes. Factor VII antigen (VIlag) and activity along with plasma lipoproteins were determined before and after intake of a mixed meal-type of oral fat load. Patients with the Arg/Gln genotype had basal VIlag and activated factor VII (Vila) levels 75% and 48%, respectively, of those of patients homozygous for the Arg allele. In absolute terms, Vila increased more in homozygotes for the Arg allele (AO-6 h Vila 1.76 ± 1.48 ng/ml) than in heterozygotes (0.60 ± 0.27 ng/ml) in response to fat intake, but the percentage increase in Vila molecules did not differ significantly between subjects with Arg/Arg and Arg/Gln genotypes (37 ± 32% versus 27 ± 15%). This suggests that the influence of the Arg/Gln polymorphism on factor VII activity is mainly accounted for by differences in the basal factor VII protein level between genotypes. Since most of our lives are spent in the postprandial state, possession of the factor VII-Gln353 allele is likely to confer protection against coronary heart disease by reducing the amount of Vila produced in response to fat intake.

scholarly journals Gene-based FVIIa prophylaxis modulates the spontaneous bleeding phenotype of hemophilia A rats

2019 ◽  
Vol 3 (3) ◽  
pp. 301-311 ◽  
Author(s):  
Shannon M. Zintner ◽  
Juliana C. Small ◽  
Giulia Pavani ◽  
Lynn Dankner ◽  
Oscar A. Marcos-Contreras ◽  
...  
Keyword(s):  
Statistical Power ◽  
Hemophilia A ◽  
Tolerance Induction ◽  
Coagulation Factor ◽  
Factor Vii ◽  
Spontaneous Bleeding ◽  
Coagulation Factor Vii ◽  
On Demand ◽  
Activated Factor Vii

Abstract A sizable proportion of hemophilia inhibitor patients fails immune tolerance induction and requires bypass agents for long-term bleed management. Recombinant human-activated coagulation Factor VII (rhFVIIa) is an on-demand bypass hemostatic agent for bleeds in hemophilia inhibitor patients. Prophylactic use of rhFVIIa may enable sustained hemostatic management of inhibitor patients, but the critical relationship of rhFVIIa circulating levels and clinical outcome in that setting remains unclear. To address this in vivo, we used the rat hemophilia A (HA) model that exhibits spontaneous bleeds and allows longitudinal studies with sufficient statistical power. We simulated activated Factor VII (FVIIa) prophylaxis by adeno-associated virus (AAV) gene transfer of a rat FVIIa transgene. Compared with naive HA animals, rat FVIIa continuous expression affected the overall observed bleeds, which were resolved with on-demand administration of recombinant rat FVIIa. Specifically, although 91% of naive animals exhibited bleeds, this was reduced to 83% and 33% in animals expressing less than 708 ng/mL (<14 nM) and at least 708 ng/mL (≥14 nM) rat FVIIa, respectively. No bleeds occurred in animals expressing higher than 1250 ng/mL (>25 nM). Rat FVIIa expression of at least 708 ng/mL was also sufficient to normalize the blood loss after a tail vein injury. Continuous, AAV-mediated rat FVIIa transgene expression had no apparent adverse effects in the hemostatic system of HA rats. This work establishes for the first time a dose dependency and threshold of circulating FVIIa antigen levels for reduction or complete elimination of bleeds in a setting of FVIIa-based HA prophylaxis.

The Clotting Trigger Is an Important Determinant for the Coagulation Pathway In Vivo or In Vitro—Inference from Data Review

2020 ◽  
Author(s):  
Shu He ◽  
Honglie Cao ◽  
Charlotte Thålin ◽  
Jan Svensson ◽  
Margareta Blombäck ◽  
...  
Keyword(s):  
Coagulation Factor ◽  
Coagulation Cascade ◽  
Factor Vii ◽  
Enzymatic Reactions ◽  
Rotational Thromboelastometry ◽  
Coagulation Factor Vii ◽  
Charged Surfaces ◽  
Coagulation Pathway

AbstractBlood coagulation comprises a series of enzymatic reactions leading to thrombin generation and fibrin formation. This process is commonly illustrated in a waterfall-like manner, referred to as the coagulation cascade. In vivo, this “cascade” is initiated through the tissue factor (TF) pathway, once subendothelial TF is exposed and bound to coagulation factor VII (FVII) in blood. In vitro, a diminutive concentration of recombinant TF (rTF) is used as a clotting trigger in various global hemostasis assays such as the calibrated automated thrombogram, methods that assess fibrin turbidity and fibrin viscoelasticity tests such as rotational thromboelastometry. These assays aim to mimic in vivo global coagulation, and are useful in assessing hyper-/hypocoagulable disorders or monitoring therapies with hemostatic agents. An excess of rTF, a sufficient amount of negatively charged surfaces, various concentrations of exogenous thrombin, recombinant activated FVII, or recombinant activated FIXa are also used to initiate activation of specific sub-processes of the coagulation cascade in vitro. These approaches offer important information on certain specific coagulation pathways, while alterations in pro-/anticoagulants not participating in these pathways remain undetectable by these methods. Reviewing available data, we sought to enhance our knowledge of how choice of clotting trigger affects the outcome of hemostasis assays, and address the call for further investigations on this topic.

The use of activated recombinant coagulation factor VII during haemarthroses and synovectomy in a patient with congenital severe factor V deficiency

Haemophilia ◽  
2005 ◽  
Vol 11 (2) ◽  
pp. 167-170 ◽  
Author(s):  
R. Gonzalez-Boullosa ◽  
R. Ocampo-Martinez ◽  
M. J. Alarcon-Martin ◽  
M. Suarez-Rodriguez ◽  
L. Dominguez-Viguera ◽  
...  
Keyword(s):  
Coagulation Factor ◽  
Factor Vii ◽  
Factor V ◽  
Coagulation Factor Vii ◽  
Factor V Deficiency

Glyco-engineered HEK 293-F cell lines for the production of therapeutic glycoproteins with human N-glycosylation and improved pharmacokinetics

Glycobiology ◽  
2021 ◽  
Author(s):  
Rico Uhler ◽  
Ruth Popa-Wagner ◽  
Mario Kröning ◽  
Anja Brehm ◽  
Paul Rennert ◽  
...  
Keyword(s):  
Cell Line ◽  
Therapeutic Proteins ◽  
Coagulation Factor ◽  
Factor Vii ◽  
Hek 293 ◽  
Knock Out ◽  
Hek 293 Cells ◽  
Coagulation Factor Vii ◽  
293 Cells

Abstract N-glycosylated proteins produced in human embryonic kidney 293 (HEK 293) cells often carry terminal N-acetylgalactosamine (GalNAc) and only low levels of sialylation. On therapeutic proteins, such N-glycans often trigger rapid clearance from the patient bloodstream via efficient binding to asialoglycoprotein receptor (ASGP-R) and mannose receptor (MR). This currently limits the use of HEK 293 cells for therapeutic protein production. To eliminate terminal GalNAc, we knocked-out GalNAc transferases B4GALNT3 and B4GALNT4 by CRISPR/Cas9 in FreeStyle 293-F cells. The resulting cell line produced a coagulation factor VII-albumin fusion protein without GalNAc but with increased sialylation. This glyco-engineered protein bound less efficiently to both the ASGP-R and MR in vitro and it showed improved recovery, terminal half-life and area under the curve in pharmacokinetic rat experiments. By overexpressing sialyltransferases ST6GAL1 and ST3GAL6 in B4GALNT3 and B4GALNT4 knock-out cells, we further increased factor VII-albumin sialylation; for ST6GAL1 even to the level of human plasma-derived factor VII. Simultaneous knock-out of B4GALNT3 and B4GALNT4, and overexpression of ST6GAL1 further lowered factor VII-albumin binding to ASGP-R and MR. This novel glyco-engineered cell line is well-suited for the production of factor VII-albumin and presumably other therapeutic proteins with fully human N-glycosylation and superior pharmacokinetic properties.

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