Thrombin Generation Assessment in Platelet Rich Plasma Offers Additional Criteria for Low Molecular Weight Heparin Antithrombotic Fingerprint Characterization.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4177-4177
Author(s):  
Grigorios T. Gerotziafas ◽  
Galea Vasso ◽  
Jeanine M. Walenga ◽  
Evi Kalodiki ◽  
Mourad Chaari ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Physicochemical Properties ◽  
Thrombin Generation ◽  
Platelet Rich Plasma ◽  
Inhibitory Effect ◽  
Low Molecular Weight ◽  
Generation Process ◽  
Health Authorities ◽  
The Impact

Abstract Abstract 4177 Introduction National and international organizations recognize that low molecular weight heparins (LMWHs) are distinct entities and that they should not be used interchangeably in clinical practice. Physicochemical properties, such as molecular weight anti-Xa/anti-IIa ratios, are used by health authorities to establish LMWHs differentiation. LMWHs are multitargeted drugs in which small differences in physicochemical properties may lead to significantly different inhibition of blood coagulation. In the present in vitro study we have investigated if thrombin generation assay, reflecting the overall coagulation process, is a suitable tool for the LMWH variability evalution. Materials and methods LMWHs (bemiparin, enoxaparin, nadroparin, dalteparin and tinzaparin) and unfractionated heparin (UFH) were added in vitro in normal citrated platelet rich plasma from 10 healthy individuals, at clinically relevant concentrations (ranging from 0.2 to 1 anti-Xa IU/ml). Thrombin generation was studied with Thrombogram-Thrombinoscope® assay using diluted thromboplastin (Innovin®, 1/1000 final dilution, Siemens France). The concentrations doubling the lag-time, the time to Peak, the IC50 for Peak, the endogenous thrombin potential (ETP) and the mean rate index of propagation phase (MRI) were determined for each compound. Results LMWHs compared on their anti-Xa activity basis showed variable inhibitory effect on thrombin generation. At equivalent anti-Xa concentrations tinzaparin was significantly more potent than the other LMWHs, being almost similar to UFH profile. Enoxaparin, nadroparin and dalteparin showed a similar inhibitory activity. Bemiparin had the lesser pronounced inhibitory effect on thrombin generation. The impact of anti-Xa and anti-IIa activities on each phase of thrombin generation process is different. Thrombogram chronometric parameters are mainly influenced by the anti-IIa activity. Similarly, ETP inhibition depends more on anti-IIa rather than anti-Xa activity. The MRI of the propagation phase is more sensitive to the anti-Xa activity of LMWHs. Conclusion Thrombin generation assessment in platelet rich plasma allows to evaluate the anticoagulant fingerprint of LMWHs and to differentiate them on global and functional criteria. Each LMWH has a particular inhibitory impact on each phase of thrombin generation process. These functional criteria need to be standardized and probably required for a better characterization of LMWHs heterogeneity by health authorities. They could be used also to evaluate bioequivalence of generic and original LMWHs. Disclosures: No relevant conflicts of interest to declare.

HIGH CONCENTRATIONS OF HEPARIN ARE MORE INHIBITORY TO PLATE- AGGREGATION IN-VITRO THAN ARE LOW MOLECULAR WEIGHT HEPARINS AND HEPARINOIDS AT THE SAME CONCENTRATION

1987 ◽  
Author(s):  
H Messmore ◽  
B Griffin ◽  
J Seghatchian ◽  
E Coyne
Keyword(s):  
Molecular Weight ◽  
Heparan Sulfate ◽  
Dermatan Sulfate ◽  
Platelet Rich Plasma ◽  
Inhibitory Effect ◽  
Low Molecular Weight ◽  
Low Concentrations ◽  
High Concentrations ◽  
Induced Aggregation

Other investigators have shown that heparin in the usual therapeutic range (0.1-0.5 units/ml) has an enhancing effect on ADP aggregation and an inhibitory effect on collagen and thrombin induced aggregation. The effects of low molecular weight heparin (LMWH)and heparinoids (dermatan sulfate, heparan sulfate) on platelet aggregation have not been as extensivelystudied. We have utilized citrated platelet rich plasma (3.2%citrate-whole blood 1:9) drawn in plastic and adjusted to a final platelet count of 250,000/ul. A Bio-Data 4 channgl aggregometer was utilized with constantstirring at 37 C. The reaction was allowed to run for 20 minutes. Platelet rich plasma was supplemented 1:9 with saline or heparin and various agonists were then added ifno aggregation occurred. ADP, collagen, thrombin, ristocetin and serum from patients with heparin inudced thrombocytopenia (HIT) were utilized as agonists. Heparin was substituted at concentrations of 0.1 to 500 units per ml and various LMWH and heparinoids were substituted in equivalent anti-Xa or gravimetric concentrations. At low concentrations no inhibitory effect on any ofthe agonists was observed with any of the heparins or heparinoids. At concentrations of heparin of 100 u/ml or greater, all agonists were inhibited. At equivalent concentrations of five different LMWH (Cy 216, Cy 222, Pk 10169, Kabi 2165 and pentasaccharide) inhibition did notoccur at all or at very high concentions only. Dermatan sulfate and heparan sulfate inhibited only at high concentrations. HIT serum could not aggregate platelets with dermatan sulfate or pentasaccharide atany concentrations, but it was a good agonist with the other heparins and heparinoids.

HEPARIN-INDUCED THROMBOCYTOPENIA: IN VITRO STUDIES WITH LOW MOLECULAR WEIGHT HEPARINOID, ORG 10172

1987 ◽  
Author(s):  
B H Chong ◽  
F Ismail ◽  
J Cade ◽  
A S Gallus ◽  
S Gordon ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
Inhibitory Effect ◽  
Heparin Therapy ◽  
Cross Reactivity ◽  
Low Molecular Weight ◽  
Heparin Induced Thrombocytopenia ◽  
Normal Platelet

Heparin-induced thrombocytopenia (HIT), an adverse effect of heparin therapy, may be associated with serious thrombosis resulting in severe disability or death. An IgG heparin-dependent antibody may be demonstrated in HIT by platelet aggregation studies with patient serum/plasma and standard (s) heparin. A recent study showed high cross-reactivity of the antibody with low molecular weight (LMW) heparins as most of the 22 patient sera tested gave a positive reaction with various LMW heparins including CY222, CY216, PK10169 and Kabi 2165 (Messmore HL et al, Blood 64(5), 1984 suppl). However, cross-reactivity rate with Org 10172, a LMW heparinoid which has strong anti-Xa but negligible antithrombin activity, is unknown. We tested the plasma of 17 patients with HIT for cross-reactivity with Org 10172. Although all 17 patient plasmas reacted positively with s.heparin (0.2 1.0 IU/ml), only 3 patient plasmas gave a positive but weaker reaction with Org 10172 (0.2-1.0 IU/ml).Further studies were performed to investigate the effect of adding Org 10172 (0.2 to 2.0 anti-Xa U/ml) to a reaction mixture of normal platelet-rich plasma, patient plasma and s.heparin (0.2 IU/ml). With 7 patient plasmas which showed no cross-reactivity with Org 10172, the antibody-induced platelet aggregation was inhibited when the concentration of Org 10172 exceeded 0.5 to 1.0 anti-Xa U/ml. When the study was repeated with other s.heparin concentrations (0.05, 0.1, 0.4 IU/ml), this inhibitory effect was again present provided the ratio of Org 10172 concentration (anti-Xa U/ml) to heparin concentration (IU/ml) exceeds 2.5 to 5. However, this inhibitory effect was not observed with the 3 patient plasmas which showed cross-reactivity with the heparinoid whqp. the same concentrations of Org 10172 were added. This inhibitory effect appeared to be specific for platelet aggregation induced by the heparin-dependent antibody as Org 10172 (<10 anti-Xa U/ml) did not affect platelet aggregation induced by collagen (2 ug/ml) and ADP (2.5 uM). These findings together with our experience of 3 cases of HIT successfully treated with Org 10172 suggest that this LMW heparinoid may be a useful drug for the treatment of HIT.

EFFECT OF VARIOUS HEPARIN(OID)S ON HEPARIN COFACTOR II MEDIATED ANTI-THROMBIN ACTIVITY AND INHIBITION OF THROMBIN GENERATION IN VITRO

1987 ◽  
Author(s):  
T G van Dinther ◽  
F Hol ◽  
D G Meuleman
Keyword(s):  
Molecular Weight ◽  
Thrombin Generation ◽  
Factor Xa ◽  
Weight Fraction ◽  
Blood Platelet ◽  
Low Molecular Weight ◽  
Heparin Cofactor Ii ◽  
Thrombin Activity ◽  
At Iii

The effects of various heparin(oid)s, standard heparin VII (SH), dermatan sulphate (DS), a low molecular weight fraction of heparin (UMW-H), FragminR (FRA), Org 10172 = low molecular weight heparinoid, the fraction of Org 10172 with high affinity for AT-III (HA-10172) and the low affinity fraction (LA-10172) respectively were examined on in vitro thrombin generation and inactivation.Thrombin inactivation in the presence of either heparin cofactor II (HC-II) or anti-thrombin III (AT-III) was assessed with two newly developed assays using the purified cofactors, thrombin and chromogenic substrate S2238 on microtiterplates. Thrombin generation in the presence of HC-II and AT-III was studied using purified factor Xa, prothrombin and blood platelet lysate and the residual thrombin activity was assessed amidolytically.The inhibition of the compounds on thrombin activity are summarized in the tableThe following conclusions can be drawn:- SH, LMW-H, HA-10172 and FRA potentiate the AT-III mediated inactivation of Ha more strongly than the HC-II mediated inactivation.- DS and LA-10172 show the reverse pattern of inactivation, while Org 10172 potentiates both inactivaton pathways to a similar extent.Thrombin generation in the presence of HC-II is inhibited by mw-heparin(oid)s at approx. 2-5 times lower concentrations than the HC-II mediated thrombin inactivation, while the inhibiting effect of SH in both assays is comparable.AT-III mediated thrombin generation inhibition and AT-III mediated thrombin inactivation is comparable as well for SH, LMW-H and FRA. In contrast, Org 10172 and its subfractions are approx. 10 times more potent on AT-III mediated thrombin generation inhibition than on AT-III mediated thrombin inactivation.Org 10172 shows low anti-thrombin activity and this activity is mainly mediated via FC-II.

scholarly journals Heparin-induced thrombocytopenia: studies with a new low molecular weight heparinoid, Org 10172

Blood ◽  
1989 ◽  
Vol 73 (6) ◽  
pp. 1592-1596
Author(s):  
BH Chong ◽  
F Ismail ◽  
J Cade ◽  
AS Gallus ◽  
S Gordon ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Platelet Aggregation ◽  
Platelet Activation ◽  
Reaction Rate ◽  
Platelet Rich Plasma ◽  
Inhibitory Effect ◽  
Cross Reaction ◽  
Low Molecular Weight ◽  
Heparin Induced Thrombocytopenia ◽  
Normal Platelet

Studies were performed to determine the cross-reaction rate of the heparin-dependent antibody with Org 10172, a new low molecular weight heparinoid, and to investigate the effect of Org 10172 on platelet activation induced by the antibody. The plasmas of 17 patients with thrombocytopenia induced by standard heparin were shown, by platelet aggregation studies, to contain the heparin-dependent antibody. Of these 17 patient plasmas, only three cross-reacted with the heparinoid, producing a cross-reaction rate of 18%. When Org 10172 was added to a reaction mixture containing normal platelet-rich plasma, patient plasma, and standard heparin with non-cross-reacting plasmas, it inhibited platelet aggregation and thromboxane B2 production induced by the antibody, provided that the ratio of Org 10172 concentration (anti- Xa U/mL) to standard heparin concentration (IU/mL) exceeded 2.5 to 5.0. This inhibitory effect was observed only with platelet activation mediated by the antibody, but not by collagen (2 micrograms/mL) or ADP (5.0 mumol/L). Additionally, three of 17 patients with serious thrombosis, whose plasma showed no cross-reaction with the heparinoid, received Org 10172 treatment with a good response in each case. These findings suggest that Org 10172 may be a useful drug for the treatment of heparin-induced thrombocytopenia.

In Vitro Characterization of a Homogeneous Low-Molecular Weight Heparin with Reversible Anticoagulant Activity

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4263-4263
Author(s):  
Matthew F Whelihan ◽  
Yongmei Xu ◽  
Jian Liu ◽  
Nigel S. Key
Keyword(s):  
Molecular Weight ◽  
Thrombin Generation ◽  
Conflicts Of Interest ◽  
Anticoagulant Activity ◽  
Low Molecular Weight ◽  
Thrombin Generation Assay ◽  
In Vitro Characterization ◽  
Fixed Concentration ◽  
Contact Pathway

Abstract Introduction. Due to their increased half-life over unfractionated heparin (UFH) and marked decrease in the incidence of heparin induced thrombocytopenia (HIT), low molecular weight heparins (LMWH) are the most widely prescribed heparin in the US. However owing to their incomplete reversibility with protamine, LMWHs (such as Enoxaparin) carry the risk of bleeding. The synthetic pentasaccharide, Fondaparinux, also lacks a specific antidote. We recently published (Xu et al. Nat. Chem. Biol. 2014) on a new class of synthetic LMWH that is not renal-excreted and offers the benefit of reversal by protamine. The new compound, dubbed “Super 12-mer”, is a 3,483 Da dodecasaccharide consisting of an antithrombin (AT) binding moiety with repeating units of IdoA2S-GlcNS6S (S is sulfate) and two 3-O-sulfate groups which afford the ability to bind protamine. We sought to characterize this new compound in a series of biochemical and global coagulation assays to better characterize its efficacy as a new reversible anticoagulant. Methods. Factor (F) Xa-AT inhibition assays were performed in both purified and plasma-based systems. The Super 12-mer was further tested in a purified prothrombinase system, as well as by tissue factor-initiated thrombin generation assays in contact pathway inhibited citrated plasma. Results. In vitro FXa inhibition studies indicated the IC50 to be 2-fold higher (49 ng/mL, 24 nM) than was previously reported. Nevertheless, the Super 12-mer anti FXa activity was approximately 2-fold greater than Enoxaparin at identical concentrations. However, the anti FXa activities of the Super 12-mer and Enoxaparin in plasma-based systems were roughly equivalent. Prothrombinase experiments indicated that both the Super 12-mer and Enoxaparin were equivalent in their ability to inhibit FXa in complex with FVa. When the two heparinoids were compared in a plasma-based thrombin generation assay (TGA), their effects on thrombin generation were nearly identical with a 50% reduction in peak thrombin generation occurring at approximately 325 nM heparinoid. When protamine is titrated against a fixed concentration of Super 12-mer (625 nM), the Super 12-mer displays a complete reconstitution of thrombin generation. Conclusions. In plasma and purified systems, the Super 12-mer displayed virtually identical efficacy in FXa inhibition compared to Enoxaparin. In buffered systems, the Super 12-mer was approximately 2-fold more active than Enoxaparin against FXa suggesting the Super 12-mer may have other binding partners in plasma. Interestingly, FXa inhibition in prothrombinase was essentially identical between the two heparinoids. Unlike Enoxaparin however, the Super 12-mer displayed near complete reversibility with protamine in TGAs. A significant lag in thrombin generation was observed when protamine was added, consistent with a previous report (Ni Ainle et al. Blood 2009) that protamine itself can act as an anticoagulant by interfering with FV activation. These data show that the Super 12-mer has almost identical efficacy to Enoxaparin in terms of FXa inhibition, while displaying significant reversibility with protamine. Taken together with the fact that this compound can be safely used in renal-impaired patients, the Super 12-mer is a promising new heparanoid anticoagulant with a potentially enhanced safety profile. Disclosures No relevant conflicts of interest to declare.

scholarly journals Heparin-induced thrombocytopenia: studies with a new low molecular weight heparinoid, Org 10172

Blood ◽  
1989 ◽  
Vol 73 (6) ◽  
pp. 1592-1596 ◽  
Author(s):  
BH Chong ◽  
F Ismail ◽  
J Cade ◽  
AS Gallus ◽  
S Gordon ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Platelet Aggregation ◽  
Platelet Activation ◽  
Reaction Rate ◽  
Platelet Rich Plasma ◽  
Inhibitory Effect ◽  
Cross Reaction ◽  
Low Molecular Weight ◽  
Heparin Induced Thrombocytopenia ◽  
Normal Platelet

Abstract Studies were performed to determine the cross-reaction rate of the heparin-dependent antibody with Org 10172, a new low molecular weight heparinoid, and to investigate the effect of Org 10172 on platelet activation induced by the antibody. The plasmas of 17 patients with thrombocytopenia induced by standard heparin were shown, by platelet aggregation studies, to contain the heparin-dependent antibody. Of these 17 patient plasmas, only three cross-reacted with the heparinoid, producing a cross-reaction rate of 18%. When Org 10172 was added to a reaction mixture containing normal platelet-rich plasma, patient plasma, and standard heparin with non-cross-reacting plasmas, it inhibited platelet aggregation and thromboxane B2 production induced by the antibody, provided that the ratio of Org 10172 concentration (anti- Xa U/mL) to standard heparin concentration (IU/mL) exceeded 2.5 to 5.0. This inhibitory effect was observed only with platelet activation mediated by the antibody, but not by collagen (2 micrograms/mL) or ADP (5.0 mumol/L). Additionally, three of 17 patients with serious thrombosis, whose plasma showed no cross-reaction with the heparinoid, received Org 10172 treatment with a good response in each case. These findings suggest that Org 10172 may be a useful drug for the treatment of heparin-induced thrombocytopenia.

Abciximab Does Not Inhibit the Increase of Thrombin Generation Produced in Platelet-Rich Plasma In Vitro by Sodium Arachidonate or Tissue Factor

2005 ◽  
Vol 11 (3) ◽  
pp. 271-277 ◽  
Author(s):  
Raul Altman ◽  
Alejandra Scazziota ◽  
Silvina Santoro ◽  
Claudio Gonzalez
Keyword(s):  
Tissue Factor ◽  
Thrombin Generation ◽  
Platelet Rich Plasma ◽  
Coronary Intervention ◽  
Inhibitory Effect ◽  
Platelet Membrane ◽  
Time To Peak ◽  
Coronary Events

Aspirin and platelet membrane glycoprotein (GP) IIb/IIIa blockers are currently used for acute coronary events, and in percutaneous coronary intervention for preventing further coronary outcomes, because they inhibit platelet function. Aspirin also inhibits thrombin generation (TG) in platelet-rich plasma (PRP) activated by sodium arachidonate (AA). The effect of the platelet membrane GP IIb-IIIa (integrin αIIbβ3) blocker abciximab on thrombin generation was studied in vitro using PRP. Thirty healthy volunteers taking no medication, and 28 volunteers who had taken aspirin (160 mg/day for 3-4 days), were included in the protocol. Control or in vivo aspirinated PRP, stimulated or not by AA or tissue factor (TF), was investigated for the inhibitory effect of abciximab pre-incubated for 3 minutes. AA and TF added in vitro activated non-aspirinated PRP: lag-time (LT) and time to peak (TTP) were significantly shortened. Peak TG (PTG) and endogenous thrombin potential (ETG) were increased by AA but not TF; thus, AA seems to be more efficient than TF for TG in this system. Abciximab added in vitro to non-activated, non-aspirinated PRP had no effect on LT, TTP, or ETP, but caused a decrease in PTG that was not statistically significant. Abciximab (3 or 4 μg/mL) added in vitro to AA or TF-activated, non-aspirinated PRP produced no effect on TG, although in aspirinated platelets both LT and time to peak were prolonged. AA as well as TF added in vitro to PRP or in vivo aspirinated PRP increased TG, although AA seems to be more efficient in our assay system. Abciximab, which affects nonaspirinated, nonactivated PRP weakly, has no effect on AA or TF in activated control PRP or in vivo aspirinated PRP.

Inhibition of In Vitro Thrombin Generation: Another Parameter Reinforcing the LMWH Heterogeneity.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 912-912 ◽  
Author(s):  
Grigoris T. Gerotziafas ◽  
Anna D. Petropoulou ◽  
Mohamed Hatmi ◽  
Meyer M. Samama ◽  
Ismail Elalamy
Keyword(s):  
Thrombin Generation ◽  
Platelet Rich Plasma ◽  
In Vitro Study ◽  
Lag Time ◽  
Inhibitory Effect ◽  
Therapeutic Concentration ◽  
Unfractioned Heparin ◽  
Concentration Peak ◽  
Pathway Activation

Abstract Introduction: Low molecular weight heparins (LMWH) are derived from unfractioned heparin (UFH) by depolymerization. Thus, they present biochemical and pharmacological differences and the ratio of anti-Xa/anti-IIa activities varies from one product to another. In this study, we compared in vitro the Thrombin Generation (TG) inhibition potency of various LMWHs and UFH using the Thrombogram-Thrombinoscope® assay. Materials and Methods: TG was assessed after Tissue Factor (TF) pathway activation in Platelet Rich Plasma (PRP) (1.5x105 platelets/μl) using diluted thromboplastin (Dade Innovin®, 1:1000 final dilution). We studied five different LMWHs (Enoxaparin, Dalteparin, Nadroparin, Tinzaparin and Bemiparin), as well as UFH at five different prophylactic and therapeutic anti-Xa final concentrations. These agents were added to control plasma from 14 healthy volunteers with equivalent anti-Xa concentrations. TG was initiated by adding the triggering solution containing CaCl2 and the fluorogenic substrate. The analyzed TG parameters are the lag time, the maximal concentration of thrombin (Cmax), the time to reach Cmax (Tmax), the TG velocity and the endogenous thrombin potential (ETP). Results: Bemiparin had almost no effect on TG, with concentrations below 0.60 IUanti-Xa/ml. Dalteparin, Nadroparin and Enoxaparin showed a similar potency in inhibiting TG at equal anti-Xa concentrations. Tinzaparin proved to be the most active LMWH in inhibiting TG and had a similar potency to UFH. Tinzaparin and UFH, with the lowest anti-Xa/anti-IIa ratio, exerted their inhibitory effect mostly by prolonging lag time and Tmax and by reducing TG velocity, especially at concentration below 0.40 IU anti-Xa/ml. Besides, UFH totally inhibited TG, as expressed by ETP, at a concentration over 0.40 IU anti-Xa/ml. For a given anti-Xa/anti-IIa ratio characterizing each LMWH the IC50 for each parameter was different. The IC50 for the reduction of TG velocity was lower in comparison to the IC50 for the other parameters (Table 1). Conclusion: Our study reinforces the concept of LMWH heterogeneity and the important effect exerted by the additional anti-IIa activity combined with anti-Xa activity. Thus, their characterization can be made through their ability to inhibit TG and not only their anti-Xa/anti-IIa ratio. Nevertheless, in vitro study ignores pharmacokinetic characteristics which are important in clinical practice. Thus, Enoxaparin, at validated prophylactic concentrations (0.20–0.40 IU anti-Xa/ml) had a weak effect on TG parameters, whereas, at peak therapeutic concentrations, it showed an important inhibitory activity similar to Tinzaparin. (Table 2).The use of TG test for the biological monitoring of LMWH requires further evaluation. Table 1. The IC50 for each parameter of TG (anti-Xa IU/ml) Lag time Tmax ETP Cmax Velocity Bemiparin >1 >1 0.98 0.85 0.68 Enoxaparin 0.62 0.58 0.55 0.45 0.38 Nadroparin 0.80 0.75 0.55 0.45 0.38 Dalteparin 0.65 0.65 0.50 0.42 0.38 Tinzaparin 0.35 0.28 0.35 0.25 0.18 UFH 0.05 0.10 0.30 0.25 0.18 Table 2. Effect of Enoxaparin and Tinzaparin on TG at therapeutic concentration peak in vitro Therapeutic Concentration Peak (anti-Xa) Lag Time Tmax ETP Cmax Velocity Enoxaparin 1IU/ml 91% 94% 72% 82% 91% Tinzaparin 0.85 IU/ml >100% >100% 82% 90% 97%

The Relative Antithrombotic Effectiveness of Heparin, a Low Molecular Weight Heparin, and a Pentasaccharide Fragment in an Animal Model

1989 ◽  
Vol 61 (02) ◽  
pp. 204-207 ◽  
Author(s):  
D P Thomas ◽  
R E Merton ◽  
E Gray ◽  
T W Barrowcliffe
Keyword(s):  
Molecular Weight ◽  
Animal Model ◽  
Low Molecular Weight Heparin ◽  
Thrombin Generation ◽  
Thrombus Formation ◽  
Relative Effectiveness ◽  
Low Molecular Weight ◽  
Antithrombotic Agent

SummaryThe antithrombotic efficacy of unfractionated heparin (UFH), a low molecular weight heparin (LMWH) and a synthetic pentasaccharide (PENTA) has been compared in an animal model for stasis thrombosis. We have also compared the relative ability of these three agents to impair thrombin generation in vitro and in vivo, and measured their effects on anti-Xa activity and thrombin clotting times. UFH, LMWH and PENTA all had the capacity to impair thrombogenesis, although there were marked differences in their relative effectiveness. Reduction of thrombin generation to 20% of control values was closely correlated with the prevention of thrombosis after 20 minutes’ stasis, but this was only achieved with UFH. The same dry weight dose of LMWH or PENTA reduced thrombin generation to about half control values, and neither significantly impaired thrombus formation after 20 minutes’ stasis. Impaired thrombin generation correlated better than anti-Xa activity with prevention of stasis thrombosis. In this model, UFH was clearly superior to LMWH and PENTA as an antithrombotic agent.

The Effect of Subcutaneous Injection of Unfractionated and Low Molecular Weight Heparin on Thrombin Generation in Platelet Rich Plasma - A Study in Human Volunteers

1994 ◽  
Vol 72 (05) ◽  
pp. 705-712 ◽  
Author(s):  
A Vicky Bendetowicz ◽  
Hu Kai ◽  
Richard Knebel ◽  
H Caplain ◽  
H Coenraad Hemker ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Low Molecular Weight Heparin ◽  
Thrombin Generation ◽  
Platelet Rich Plasma ◽  
Factor Xa ◽  
Critical Length ◽  
Low Molecular Weight ◽  
High Dose ◽  
Platelet Factor ◽  
Thrombin Inhibition

SummaryWe administered a dose of unfractionated heparin (UFH) and two doses of a low molecular weight heparin (LMWH) to healthy volunteers by SC injection. The doses given were: a) UFH, 5000IU, which represents 8.7 mg of >5,400 MW active heparin (ACLM) and no <5,400 active heparin (BCLM), b) enoxaparin 40 mg (3.4 mg ACLM, 2.2 mg BCLM) and c) enoxaparin 1 mg/kg body weight (on the mean 75 mg, containing 6.4 mg ACLM and 4.1 mg BCLM). We determined the effect on thrombin generation in platelet rich plasma (PRP) between 1 and 8 h after injection. UFH administration caused only a 5-8% inhibition of the thrombin potential (i. e. the area under the thrombin generation curve). Significantly higher inhibition of the thrombin potential was seen after administration of both doses of enoxaparin. To wit 9-26% at the low dose and 29-46% at the high dose. UFH injection caused a prolongation of the lag-time before the thrombin burst. Only with the high dose of enoxaparin the lag-times were significantly more prolonged with enoxaparin than with UFH.Excess amounts of platelet factor 4 (PF4) were able to neutralize completely the anti-thrombin activity in normal plasma spiked with enoxaparin as well as in plasma samples obtained after SC enoxaparin injection. With a large excess of PF4 the anti-factor Xa activity could be inhibited to a maximum of 50%. This indicates that ACLM (above critical length material, MW >5400) is neutralized completely by PF4 whereas BCLM (below critical length material, MW <5400) is not. The anti-thrombin heparin-activity, hence the ACLM fraction of heparin, was shown to have disappeared from the serum of PRP samples. The BCLM fraction was found after coagulation of PRP in concentrations that were indistinguishable from those in the PPP.We conclude that in PRP the activity of the BCLM fraction of injected LMWH remains after platelet activation. The possible role of this activity in thrombin inhibition and in the antithrombotic action of low molecular weight heparins is discussed.

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