scholarly journals Unfractionated heparin inhibits thrombin-catalysed amplification reactions of coagulation more efficiently than those catalysed by factor Xa

1989 ◽  
Vol 257 (1) ◽  
pp. 143-150 ◽  
Author(s):  
F A Ofosu ◽  
J Hirsh ◽  
C T Esmon ◽  
G J Modi ◽  
L M Smith ◽  
...  
Keyword(s):  
Factor Viii ◽  
Antithrombin Iii ◽  
Factor Xa ◽  
Activate Factor ◽  
Factor V ◽  
Inhibitory Effects ◽  
Exogenous Factor ◽  
Additional Support ◽  
Prothrombin Activation ◽  
Inhibit Factor

We have proposed previously that the steps in coagulation most sensitive to inhibition by heparin are the thrombin-dependent amplification reactions, and that prothrombinase is formed in heparinized plasma only after Factor Xa activates Factor VIII and Factor V. These propositions were based on the demonstration that both heparin and Phe-Pro-Arg-CH2Cl completely inhibited 125I-prothrombin activation for up to 60 s when contact-activated plasma (CAP) was replenished with Ca2+. Furthermore, the addition of thrombin to CAP before heparin or Phe-Pro-Arg-CH2Cl completely reversed their inhibitory effects. Additional support for the above hypotheses is provided in this study by demonstrating that, when the activity of thrombin is suppressed by heparin (indirectly) or by Phe-Pro-Arg-CH2Cl (directly), exogenous Factor Xa reverses the ability of these two agents to inhibit prothrombin activation. Prothrombin activation was initiated by adding Factor Xa (1 nM) or thrombin (1 or 10 nM) simultaneously with CaCl2 to CAP. In the absence of heparin or Phe-Pro-Arg-CH2Cl, prothrombin activation was seen 15 s later in either case. Heparin failed to delay, and Phe-Pro-Arg-CH2Cl delayed for 15 s, prothrombin activation in CAP supplemented with Factor Xa. In contrast, heparin and Phe-Pro-Arg-CH2Cl completely inhibited prothrombin activation for at least 45 s in CAP supplemented with 1 nM-thrombin. Heparin failed to delay prothrombin activation in CAP supplemented with 10 nM-thrombin, whereas Phe-Pro-Arg-CH2Cl completely inhibited prothrombin activation in this plasma for 45 s. These results suggest that in CAP: (1) Factor Xa can effectively activate Factor VIII and Factor V when the proteolytic activity of thrombin is suppressed; (2) heparin-antithrombin III is less able to inhibit Factor Xa than thrombin; (3) suppression of the thrombin-dependent amplification reactions is the primary anticoagulant effect of heparin.

HEPARIN IS NOT AN EFFICIENT INHIBITOR OF THE FACTOR Xa-DEPENDENT ACTIVATION OF FACTOR V AND FACTOR VIII

1987 ◽  
Author(s):  
F A Ofosu ◽  
G J Modi ◽  
M R Buchanan ◽  
J Hirsh ◽  
M A Blajchman
Keyword(s):  
Factor Viii ◽  
Antithrombin Iii ◽  
Specific Inhibitor ◽  
Factor Xa ◽  
Factor V ◽  
Factor X ◽  
Inhibitory Effects ◽  
Efficient Inhibitor ◽  
System 1 ◽  
Prothrombin Activation

We have previously proposed that the steps in coagulation most sensitive to inhibition by heparin are the thrombin-dependent activation of factor V and factor VIII. This observation was based on the demonstration that therapeutic concentrations of heparin or 1μM of the thrombin specific inhibitor, phe-pro-arg CH2Cl (PPACK) completely inhibited the activation of prothrombin when contact-activated plasma (CAP) was recalcified for up to 1 min. Under similar conditions, heparin and PPACK only partially inhibited the activation of factor X. Moreover, the addition of thrombin (lOnM) to CAP 1 min before that of heparin or PPACK reversed their inhibitory effects. We now provide further support for our hypothesis by showing that when the activity of thrombin is suppressed by heparin or PPACK, efficient activation of radiolabelled prothrombin occurs only when the factor Xa then present activates factor V and factor VIII. We compared the effects of HEP of PPACK on the following four systems for initiating the activation of prothrombin: (1) CAP; (2) CAP + lOnM thrombin; (3) CAP + InM Xa and (4) unactivated plasma + InM Xa + InM Va + coagulant phospholipids. In each system, the enzymes were added 1 min before the heparin or PPACK. In the absence of heparin or PPACK, all four systems generated the same amount of thrombin activity in 45s. Complete inhibition of prothrombin activation by heparin and PPACK was observed only in system 1 which did not contain exogenous thrombin or factor Xa. No inhibition by heparin or PPACK was observed when thrombin or factor Xa was added to CAP in systems (2) and (3). Only partial inhibition was observed in system (4) which contained exogenous prothrombi-nase complex. Factor Xa thus provides an effective by-pass mechanism for the activation of factor VIII and factor V in plasma containing therapeutic concentrations of heparin. Our data provide further evidence that the heparin-antithrombin III system is not effective in inactivating factor Xa. These results support the hypothesis that in unactivated normal plasma, the primary anticoagulant effect of heparin is the inhibition of the thrombin-dependent activation of factor V and factor VIII.

scholarly journals Characterization of an inducible endothelial cell prothrombin activator

Blood ◽  
1996 ◽  
Vol 88 (8) ◽  
pp. 2989-2994 ◽  
Author(s):  
L Liu ◽  
GM Rodgers
Keyword(s):  
Thrombin Generation ◽  
Interleukin 1 ◽  
Factor Xa ◽  
Apparent Molecular Weight ◽  
Factor V ◽  
Factor X ◽  
Exogenous Factor ◽  
Km Value ◽  
Prothrombin Activation

In vivo prothrombin activation is thought to occur via a factor Xa/factor V-dependent mechanism. We investigated whether human venous endothelial cells (EC) could be induced to express a prothrombin activator. EC treated with lipopolysaccharide (LPS) or interleukin-1 activated prothrombin in the absence of exogenous factors Xa and V. This activity resided in the membrane fraction of EC and was not inhibited by an antibody to factor V. The apparent Km value was 3.3 +/- 0.3 mumol/L. Comparative studies of thrombin generation using a model system of phospholipid and factors Xa/V versus LPS-treated EC were performed to quantitate the effects of known inhibitors to factor Xa. The factor Xa inhibitor DEGR-chloromethyl ketone and an antibody to factor X inhibited prothrombin activation. However, the EC activator did not hydrolyze a factor Xa chromogenic substrate, and recombinant tick anticoagulant peptide did not suppress activity of the prothrombin activator. The apparent molecular weight of the EC activator was approximately 30 kD. Exogenous factor V enhanced the activity of the EC activator, such that in the presence of factor V, the apparent K(m) value was 1.28 +/- 0.10 mumol/L. Additionally, LPS-treated EC activated exogenous factor V. This activator has several characteristics of a previously described inducible murine monocyte prothrombin activator and may contribute to thrombin generation associated with pathologic stimuli.

Heparin and Low Molecular Weight Heparins Inhibit Prothrombinase Formation but not its Activity in Plasma

1994 ◽  
Vol 72 (06) ◽  
pp. 862-868 ◽  
Author(s):  
Frederick A Ofosu ◽  
J C Lormeau ◽  
Sharon Craven ◽  
Lori Dewar ◽  
Noorildan Anvari
Keyword(s):  
Factor Xa ◽  
Catalytic Efficiency ◽  
Thrombin Inhibitor ◽  
Lag Phase ◽  
Factor V ◽  
Factor X ◽  
Normal Plasma ◽  
Plasma Factor ◽  
Prothrombin Activation ◽  
Plasma Heparin

SummaryFactor V activation is a critical step preceding prothrombinase formation. This study determined the contributions of factor Xa and thrombin, which activate purified factor V with similar catalytic efficiency, to plasma factor V activation during coagulation. Prothrombin activation began without a lag phase after a suspension of coagulant phospholipids, CaCl2, and factor Xa was added to factor X-depleted plasma. Hirudin, a potent thrombin inhibitor, abrogated prothrombin activation initiated with 0.5 and 1.0 nM factor Xa, but not with 5 nM factor Xa. In contrast, hirudin did not abrogate prothrombin activation in plasmas pre-incubated with 0.5,1.0 or 5 nM α-thrombin for 10 s followed by the coagulant suspension containing 0.5 nM factor Xa. Thus, thrombin activates plasma factor V more efficiently than factor Xa. At concentrations which doubled the clotting time of contact-activated normal plasma, heparin and three low Mr heparins also abrogated prothrombin activation initiated with 0.5 nM factor Xa, but not with 5 nM factor Xa. If factor V in the factor X-depleted plasma was activated (by pre-incubation with 10 nM a-thrombin for 60 s) before adding 0.5,1.0, or 5 nM factor Xa, neither hirudin nor the heparins altered the rates of prothrombin activation. Thus, none of the five anticoagulants inactivates prothrombinase. When 5 or 10 pM relipidated r-human tissue factor and CaCl2 were added to normal plasma, heparin and the three low Mr heparins delayed the onset of prothrombin activation until the concentration of factor Xa generated exceeded 1 nM, and they subsequently inhibited prothrombin activation to the same extent. Thus, hirudin, heparin and low Mr heparins suppress prothrombin activation solely by inhibiting prothrombinase formation.

Measurement of factor Xa-antithrombin III in plasma: relationship to prothrombin activation in vivo

1995 ◽  
Vol 90 (3) ◽  
pp. 669-680 ◽  
Author(s):  
ISABELLE GOUIN-THIBAULT ◽  
LORI DEWAR ◽  
MYRON KULCZYCKY ◽  
MARION STERNBACH ◽  
FREDERICK A. OFOSU
Keyword(s):  
Antithrombin Iii ◽  
Factor Xa ◽  
Prothrombin Activation

scholarly journals Feiba Immuno: A Preparation with Factor Eight Inhibitor Bypassing Activity

1977 ◽  
Author(s):  
F. Elsinger
Keyword(s):  
Factor Viii ◽  
Factor Xa ◽  
Coagulation Factors ◽  
Active Principle ◽  
Factor Viii Inhibitor ◽  
Factor V ◽  
In Vitro Experiments ◽  
Factor X ◽  
Viii Inhibitor

FEIBA IMMUNO is a preparation in which a new activity is generated capable of bypassing factor VIII. The preparation which is used to treat patients with inhibitors (especially inhibitors to factor VIII) is standardized in FEIBA units, i.e. in terms of its in vitro capacity to shorten the activated PTT of a factor VIII inhibitor plasma.It could be concluded from different in vitro experiments that none of the classic’ activated coagulation factors is responsible for the factor VIII bypassing reaction; FEIB-activity seems to be correlated to a new complex of coagulation factors.To get an answer to the question which coagulation factors are essential for FEIB-activity, we tried to generate this activity from different deficient plasmas; from these experiments the following conclusions could be drawn:, the presence of at least factors VII, IX, and X is essential for the generation of the molecular species responsible for factor VIII as well as factor X bypassing activity, but factor V is not bypassed. This activity is not factor Xa itself. Factors VIII and V are not necessary for the generation of this active principle, but factor V is finally needed for its bypassing action.

Effect of heparin on chronically induced intravascular coagulation in dogs

1975 ◽  
Vol 229 (2) ◽  
pp. 449-454 ◽  
Author(s):  
Owen CA ◽  
EJ Bowie
Keyword(s):  
Factor Viii ◽  
Prothrombin Time ◽  
Antithrombin Iii ◽  
Plasma Fibrinogen ◽  
Factor V ◽  
Thrombin Time ◽  
Turnover Rates ◽  
Clotting Factors ◽  
Intravascular Coagulation ◽  
Absolute Concentrations

When intermediate-strength thromboplastin was continuously infused into dogs for 10 days or more, platelet counts decreased sharply and factor VIII concentrations decreased by more than 50%. There was little change in plasma fibrinogen, prothrombin, factor V, antithrombin III, plasminogen, prothrombin time, and thrombin time values. When heparin was infused (25-50 U/kg per h) along with the same thromboplastin, there was no change in onset or degree of thrombocytopenia. However, the decrease in factor VIII was abolished and there were significant increases in fibrinogen, prothrombin, and factor V. The absolute concentrations of the various clotting factors seemed to give no indication of their turnover rates. Unexplained is the remarkable heparin tolerance that developed in these dogs.

scholarly journals Experimental Studies on Factor VIII Inhibitor Bypassing Activity

1977 ◽  
Author(s):  
H. Vinazzer
Keyword(s):  
Factor Viii ◽  
Calcium Chloride ◽  
Experimental Studies ◽  
Factor Xa ◽  
Factor Viii Inhibitor ◽  
Factor V ◽  
Factor X ◽  
Tissue Thromboplastin ◽  
Viii Inhibitor ◽  
Thrombin Formation

The exact action of factor VIII inhibitor bypassing activity (FEIBA) is still unclear. For this reason, a series of experimental studies was carried out. Procoagulant activities were examined by standard one-stage methods while factor Xa and thrombin were measured by chromogenic substrates. Activities of factors II, VII, IX, and X were similar to PPSB fractions. In addition, low factor V activity and a phospholipid were detected. No activated factor X was present in FEIBA but there was a trace amount of 2.1 NIH units of thrombin per 100 FEIBA units. On addition of calcium chloride slow thrombin formation could be observed which however, reached 1100 NIH units of thrombin per 100 FEIBA units within an incubation time of 10 min. The velocity of thrombin formation was greatly enhanced by addition of a PTT reagent and of thromboplastin respectively. Factor Xa on the other hand, was neither formed after addition of calcium chloride nor by a PTT reagent. Tissue thromboplastin however, activated Xa from FEIBA in the same manner as a PTT reagent plus barium sulfate plasma. From these results, the conclusion could be drawn that thrombin could readily be made available from FEIBA while activation of Xa either needed the complete endogenous pathway or the presence of tissue thromboplastin. The procoagulant activity of FEIBA therefore, could be attributed to direct thrombin formation. By this process, an activation of the clotting mechanism in plasmas deficient in endogenous coagulation factors, and a complete independence from the presence or absence of a specific antibody could be explained.

scholarly journals Synthetic factor VIII peptides with amino acid sequences contained within the C2 domain of factor VIII inhibit factor VIII binding to phosphatidylserine

Blood ◽  
1990 ◽  
Vol 75 (10) ◽  
pp. 1999-2004 ◽  
Author(s):  
PA Foster ◽  
CA Fulcher ◽  
RA Houghten ◽  
TS Zimmerman
Keyword(s):  
Factor Viii ◽  
Factor Xa ◽  
Amino Acid Sequences ◽  
Enzyme Linked Immunosorbent Assay ◽  
C2 Domain ◽  
Factor X ◽  
Phospholipid Binding ◽  
Carboxyl Terminal ◽  
Inhibit Factor

Abstract The effective activation of factor X by factor IXa requires the co- factor activity of activated factor VIII (FVIII). Factor Xa formation is also dependent on the presence of negatively charged phospholipid. A phospholipid binding domain of FVIII has been reported to be present on the FVIII light chain. Recent observations on a subset of human FVIII inhibitors have implicated the carboxyl-terminal C2 domain of FVIII as containing a possible phospholipid binding site. The purpose of this study was to investigate directly the role of the C2 domain in phospholipid binding. Twenty-six overlapping peptides, which span the entire C2 domain of FVIII, were synthesized. The ability of these peptides to inhibit the binding of purified human FVIII to immobilized phosphatidylserine was evaluated in an enzyme-linked immunosorbent assay. Three overlapping synthetic FVIII peptides, 2303–2317, 2305- 2332, and 2308–2322, inhibited FVIII binding to phosphatidylserine by greater than 90% when tested at a concentration of 100 mumols/L. A fourth partially overlapping peptide, 2318–2332, inhibited FVIII binding by 65%. These results suggest that the area described by these peptides, residues 2303 to 2332, may play an important role in the mediation of FVIII binding to phospholipid.

scholarly journals Activation of human factor VII by activated factors IX and X

Blood ◽  
1982 ◽  
Vol 60 (5) ◽  
pp. 1143-1150 ◽  
Author(s):  
DR Masys ◽  
SP Bajaj ◽  
SI Rapaport
Keyword(s):  
Human Factors ◽  
Factor Viii ◽  
Active Site ◽  
Human Factor ◽  
Antithrombin Iii ◽  
Factor Ix ◽  
Factor Vii ◽  
Activate Factor ◽  
Factor X ◽  
Activated Factor Vii

Factor VII clotting activity increases about five-fold when blood is clotted in glass. Prior studies suggested that this results from activation induced by activated factor IX (IXa). However, in purified systems containing phospholipid and calcium, activated factor X (Xa) is known to activate factor VII rapidly. Therefore, we studied activation of factor VII by IXa and X, in systems using purified human factors. Concentrations of IXa and Xa were calculated from total activated protein concentrations rather than from active site concentrations. In the presence of phospolipid and calcium, both IXa and Xa activated factor VII 25-fold; however, Xa was roughly 800 times more efficient than IXa. Without added phospholipid, activation of factor VII by both Xa and IXa was markedly slowed, and Xa was roughly 20 times more efficient than IXa. When both phospholipid and calcium were omitted, activation of factor VII by either enzyme was negligible. Adding normal prothrombin, but not decarboxylated prothrombin, substantially slowed activation of factor VII by both Xa and IXa. Adding thrombin-activated factor VIII and antithrombin-III did not change rates of factor VII activation by either enzyme. These results from purified systems do not provide an explanation for the prior data from plasma systems.

scholarly journals Clinical guidelines for cryosupernatant transfusions

2020 ◽  
Vol 65 (3) ◽  
pp. 351-359
Author(s):  
G. M. Galstyan ◽  
T. V. Gaponova ◽  
F. S. Sherstnev ◽  
A. A. Kupryashov ◽  
N. I. Olovnikova ◽  
...  
Keyword(s):  
Factor Viii ◽  
Antithrombin Iii ◽  
Fresh Frozen Plasma ◽  
Blood Component ◽  
Factor Viii Inhibitor ◽  
Factor V ◽  
Fresh Frozen ◽  
Viii Inhibitor ◽  
Von Willebrand ◽  
Frozen Plasma

Introduction. Cryosupernatant is blood component. Cryosupernatant is the supernatant plasma removed during the preparation of cryoprecipitate. Aim. To provide information on the composition and methods of production, storage, transportation and clinical use of Cryosupernatant. General fi ndings. In comparison with fresh frozen plasma (FFP) and cryoprecipitate, Cryosupernatant plasma is depleted in factor VIII, fi brinogen factor von Willebrand (VWF). Cryosupernatant is defi cient in high molecular weight multimers of VWF, but contains VWF metalloproteinase. The concentrations of factor V, antithrombin III, albumin and immunoglobulins are the same as in FFP and cryoprecipitate. The indications for Cryosupernatant transfusions are massive blood loss in patients with factor VIII inhibitor, plasma exchange in patients with thrombotic thrombocytopenic purpura. For children the doses of Cryosupernatant should be 10-15 mL/kg.

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