scholarly journals A novel nucleotide-based thrombin inhibitor inhibits clot-bound thrombin and reduces arterial platelet thrombus formation

Blood ◽  
1994 ◽  
Vol 83 (3) ◽  
pp. 677-682 ◽  
Author(s):  
WX Li ◽  
AV Kaplan ◽  
GW Grant ◽  
JJ Toole ◽  
LL Leung
Keyword(s):  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
Thrombin Inhibitors ◽  
Thrombin Inhibitor ◽  
Shear Rates ◽  
High Shear Rates ◽  
Dependent Inhibition ◽  
Platelet Thrombus

Abstract A novel thrombin inhibitor based on single-stranded (ss) deoxynucleotides with the sequence GGTTGGTGTGGTTGG (thrombin aptamer) has been recently discovered. In this study, we tested its efficacy in inhibiting clot-bound thrombin activity and platelet thrombus formation in an ex vivo whole artery angioplasty model. The thrombin aptamer showed a specific dose-dependent inhibition of thrombin-induced platelet aggregation (0.5 U/mL) in human platelet-rich plasma, with an IC50 of approximately 70 to 80 nmol/L. In an in vitro clot-bound thrombin assay system, heparin, used at clinically relevant concentrations of 0.2 U/mL and 0.4 U/mL, was ineffective in inhibiting clot-bound thrombin (6.5% and 34.9% inhibition at 0.2 U/mL and 0.4 U/mL, respectively). In contrast, the thrombin aptamer at an equivalent anticoagulant concentration inhibited clot-bound thrombin (79.7% inhibition). In an ex vivo whole artery angioplasty model, the thrombin aptamer markedly suppressed the generation of fibrinopeptide A (FPA), whereas heparin at 2 U/mL was ineffective. Compared with a scrambled ssDNA control, the thrombin aptamer reduced platelet deposition by 34.5% +/- 5% (mean +/- SEM, n = 4, P = .09) at low shear rates (approximately 200 s-1) and 61.3% +/- 11% (mean +/- SEM, n = 4, P = .05) at high shear rates (approximately 850 s-1). Thrombin aptamers based on ssDNA molecules represent a new class of thrombin inhibitors with potent anticoagulant and antithrombotic properties.

scholarly journals A novel nucleotide-based thrombin inhibitor inhibits clot-bound thrombin and reduces arterial platelet thrombus formation

Blood ◽  
1994 ◽  
Vol 83 (3) ◽  
pp. 677-682 ◽  
Author(s):  
WX Li ◽  
AV Kaplan ◽  
GW Grant ◽  
JJ Toole ◽  
LL Leung
Keyword(s):  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
Thrombin Inhibitors ◽  
Thrombin Inhibitor ◽  
Shear Rates ◽  
High Shear Rates ◽  
Dependent Inhibition ◽  
Platelet Thrombus

A novel thrombin inhibitor based on single-stranded (ss) deoxynucleotides with the sequence GGTTGGTGTGGTTGG (thrombin aptamer) has been recently discovered. In this study, we tested its efficacy in inhibiting clot-bound thrombin activity and platelet thrombus formation in an ex vivo whole artery angioplasty model. The thrombin aptamer showed a specific dose-dependent inhibition of thrombin-induced platelet aggregation (0.5 U/mL) in human platelet-rich plasma, with an IC50 of approximately 70 to 80 nmol/L. In an in vitro clot-bound thrombin assay system, heparin, used at clinically relevant concentrations of 0.2 U/mL and 0.4 U/mL, was ineffective in inhibiting clot-bound thrombin (6.5% and 34.9% inhibition at 0.2 U/mL and 0.4 U/mL, respectively). In contrast, the thrombin aptamer at an equivalent anticoagulant concentration inhibited clot-bound thrombin (79.7% inhibition). In an ex vivo whole artery angioplasty model, the thrombin aptamer markedly suppressed the generation of fibrinopeptide A (FPA), whereas heparin at 2 U/mL was ineffective. Compared with a scrambled ssDNA control, the thrombin aptamer reduced platelet deposition by 34.5% +/- 5% (mean +/- SEM, n = 4, P = .09) at low shear rates (approximately 200 s-1) and 61.3% +/- 11% (mean +/- SEM, n = 4, P = .05) at high shear rates (approximately 850 s-1). Thrombin aptamers based on ssDNA molecules represent a new class of thrombin inhibitors with potent anticoagulant and antithrombotic properties.

The Effect of Platelet PlA Polymorphism on Experimental Thrombus Formation in Man Depends on Blood Flow and Thrombogenic Substrate

2001 ◽  
Vol 85 (06) ◽  
pp. 1097-1103 ◽  
Author(s):  
Kjell Sakariassen ◽  
Hélène Grandjean ◽  
Claire Thalamas ◽  
Bernard Boneu ◽  
Pierre Sié ◽  
...  
Keyword(s):  
Blood Flow ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Flow Properties ◽  
Shear Rates ◽  
Perfusion Chamber ◽  
Platelet Receptor ◽  
Platelet Deposition ◽  
Coronary Syndromes ◽  
Platelet Thrombus

SummaryA number of studies have reported conflicting data on the association of the PlA1/PlA2 polymorphism of the GPIIIa gene and coronary syndromes. We have investigated the effect of this polymorphism on experimental platelet thrombus formation in man. Forty healthy male volunteers were genotyped for the PlA1/PlA2 polymorphism. Thrombus formation was induced ex vivo by exposing a tissue factor (TF) or a collagencoated coverslip in a parallel plate perfusion chamber to native blood for 2 and 4 min. The shear rates at these surfaces were 650 and 2,600 s–1. Platelet and fibrin deposition was quantified by immunoenzymatic methods. The frequencies of PlA1/PlA1 and PlA1/PlA2 genotypes were 52.5% and 47.5%, respectively. Ex vivo deposition of fibrin on TF was not affected by the PlA1/PlA2 polymorphism. However, the ex vivo platelet deposition at 650 s–1 was higher in blood from PlA1/PlA1 individuals than in PlA1/PlA2 individuals (P = 0.008 at 4 min). On collagen, neither fibrin nor platelet deposition was significantly affected by the PlA1/PlA2 polymorphism. Platelet thrombus formation is significantly influenced by genetic variations in the GPIIIa platelet receptor. This effect depends on the blood flow properties and the nature of the thrombogenic stimulus.

Lu 23051, A New Potent Inhibitor Of Platelet Aggregation

1981 ◽  
Author(s):  
H D Lehmann ◽  
J Gries ◽  
D Lenke
Keyword(s):  
Platelet Aggregation ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Coronary Vessels ◽  
Inhibitory Effect ◽  
Spontaneously Hypertensive ◽  
Dependent Inhibition ◽  
Induced Aggregation

6- [p-(2-(Chiorpropionylamino)phenyl] -4.5-dihydro-5-methyl-3(2H)-pyridazinone, LU 23051, is primarily characterized by its strong inhibition of platelet aggregation under in vitro and in vivo conditions. In vitro there is a concentration-dependent inhibition of ADP and collagen induced aggregation in platelet rich plasma of man, rat and dog. The inhibitory concentration EC 33 % is 0.0010-0.030 mg/1 (man: ADP-0.030, col 1.-0.013 mg/l) depending on species and type of aggregation. When administered orally in ex vivo experiments on rats and dogs the substance is found to have a dose-dependent antiaggregatory effect in the range from 0.1-3.16 mg/kg. The ED 33 % is 0.27-0.63 mg/kg.-In addition after oral administration the substance has a good inhibitory effect in models being based on intravascular platelet aggregation. Thus, a dose of 1 mg/kg inhibits laser-induced aggregation in mesenteric venules of rats. Mortality after i.v. injection of collagen in mice is reduced by 50 % after a dose of 0.02 mg/kg. A dose of 0.039 mg/kg prolongs the bleeding time of rats by 50 %. The aggregation-inhibiting action is of long duration (0.1 mg/kg p.o.∼24 h). The substance does not interfere with clotting.Besides its effect on platelet aggregation LU 23051 acts as vasodilatator as well. Dilatation of coronary vessels by 100 % is seen in isolated guinea-pig hearts at a concentration of 0.1 mg/l. In spontaneously hypertensive rats the substance has an anti hypertensive effect. The ED 20 % is 0.36 mg/kg p.o.The combination of antiaggregatory and vasodilatatory effects opens up interesting aspects with respect to the pharmacotherapeutic use of the new substance

Iohexol, platelet activation and thrombosis

1998 ◽  
Vol 39 (4) ◽  
pp. 349-354 ◽  
Author(s):  
K. S. Sakariassen ◽  
R. M. Barstad ◽  
M. J. A. G. Hamers ◽  
H. Stormorken
Keyword(s):  
Platelet Activation ◽  
Arterial Wall ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Human Model ◽  
Intravascular Thrombosis ◽  
Platelet Thrombus ◽  
In Vitro Platelet Activation ◽  
Platelet Secretion

Background: The nonionic monomer iohexol triggers in vitro platelet secretion of β-thromboglobulin (β-TG). This iohexol platelet activation may promote intravascular thrombosis. We studied this relationship by employing a human model of collagen-induced platelet thrombus formation at arterial flow. The ionic dimer ioxaglate, the nonionic dimer iodixanol, and glucose were included. Methods and Results: In vitro platelet activation as measured by β-TG secretion following a 1-min incubation of native blood with 50 vol% of iohexol was significant. Glucose solutions of 300, 580 and 825 mosmol, corresponding to the osmolalities of respectively iodixanol, ioxaglate and iohexol, increased the β-TG secretion in parallel with the osmolalities. Ioxaglate and iodixanol were virtually inert. Continuous infusion of iohexol or 580 or 825 mosmol glucose (40 vol%) into flowing native blood at an arterial wall shear rate of 2600 s−1 in an ex vivo collagen-induced platelet thrombus formation device triggered pronounced secretion of β-TG. However, the platelet thrombus formation in blood mixed with iohexol was within the same range as that observed with ioxaglate or iodixanol. Increasing glucose osmolality induced increasing β-TG secretion, which paralleled gradually decreasing platelet thrombus formation. Conclusion: Iohexol and 580 or 825 mosmol glucose trigger platelet secretion of β-TG. This secretion is not associated with enhanced collagen-induced platelet thrombus formation at high arterial shear.

Pharmacological Profile of DZ-697b, a Novel Anti-Platelet Agent -Selective Inhibitor of vWF- and Collagen-Induced Platelet Aggregation.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1875-1875 ◽  
Author(s):  
Yoshiyasu Ogihara ◽  
Sumie Muramatsu ◽  
Yuki Kaneda ◽  
Takako Iijima ◽  
Tomoko Shibutani ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Guinea Pigs ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
Bleeding Risk ◽  
Selective Inhibitor ◽  
Pharmacological Profile ◽  
Induced Aggregation

Abstract Introduction: Bleeding risk accompanied with anti-platelet drugs is an ultimate dilemma in the treatment of thrombosis patient. Under high shear condition of blood flow, vWF- and collagen-induced signaling pathways are likely to trigger the platelet adhesion to the injured endothelium, which leads to the activation of platelets and arterial thrombus formation. Thus, the recent studies suggest that the selective inhibitor of these pathways is a new target of anti-platelet drugs with lower bleeding risk. We report here a pharmacological profile of DZ-697b, which selectively inhibits platelet aggregation evoked by ristocetin and collagen in vitro and ex vivo. Materials and methods: Human volunteers blood was processed platelet rich plasma (PRP) or washed platelets. PRP aggregation was induced by ristocetin and collagen. To reveal the selectivity, effect of DZ-697b on U46619 (TXA2 analogue), ADP, thrombin and TRAP induced aggregation in the washed platelets were examined. In guinea pigs and cynomolgus monkeys, effects of DZ-697b given orally were also examined on ex vivo PRP aggregation induced by collagen. To investigate the underlying mechanisms of DZ-697b, changes in phosphorylation of FcR γ chain, a common signaling pathway of both vWF- and collagen-induced platelet aggregation, were studied. Results: DZ-697b potently inhibited both ristocetin- and collagen-induced human PRP aggregation, the IC50 being 0.74 μM and 0.55 μM, respectively. In contrast, DZ-697b even at 50 μM did not show any influences on U46619, ADP, thrombin and TRAP induced platelet aggregation. DZ-697b did not affect ovine COX-1 and COX-2 activities at up to 300 μM. The bioavailability of this compound was more than 80% in monkeys. Oral administration of DZ-697b at 1–3 mg/kg significantly and persistently inhibited collagen induced PRP aggregation in monkeys and guinea pigs. Application of ristocetin, vWF, and collagen significantly increased the intensity of phosphorylation of FcR γ chain in washed platelets, which were inhibited by DZ-697b. Conclusion: DZ-697b is an orally active compound which selectively inhibits ristocetin- and collagen-induced platelet aggregation and seems to be promising as novel anti-platelet drug.

Antithrombogenic Coating of Stents Using a Biodegradable Drug Delivery Technology

1999 ◽  
Vol 82 (07) ◽  
pp. 51-57 ◽  
Author(s):  
R. Herrmann ◽  
G. Schmidmaier ◽  
B. Märkl ◽  
A. Resch ◽  
I. Hähnel ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Blood Clot ◽  
Gold Surface ◽  
Metallic Surface ◽  
Thrombin Inhibitor ◽  
Bare Steel ◽  
Bare Gold

SummaryTo reduce the thrombogenic properties of coronary artery stents, a biodegradable polylactic acid (PLA) stent coating with an incorporated thrombin inhibitor and a platelet aggregation inhibitor has been developed. In an ex vivo human stasis model, its effect on platelets, plasmatic coagulation and its release characteristics were studied using whole blood. Bare steel and bare gold-surface stents were compared to steel and gold-surface stents coated with PLA (30 kDa) containing 5% polyethyleneglycol (PEG)-hirudin and 1% iloprost, with an empty tube as control. Markers of activated coagulation (prothrombin fragment F1-2 and thrombin-antithrombin III complex, TAT), were assayed and the release of drugs from the coating was assessed by aPTT and collagen-induced platelet aggregation. Bare steel and gold stents were completely covered by a blood clot, and high levels of coagulation markers (F1-2 fragment and TAT) were detected. No differences in the thrombogenic properties were found between bare gold or steel stents. Coated stents were free of blood clots and only minor elevations of markers were detected. Release data from in-vitro studies over 90 days showed a gradual release of the drugs with an initial exponential release characteristic for PEG-hirudin, slow release of iloprost and a 10% degradation of the PLA carrier. This drug releasing biodegradable coating effectively reduced thrombus formation independent of the metallic surface.

SELECTIVE ANTAGONISTS OF PAF: INTERFERENCE WITH PLATELET FUNCTION AND EFFECT ON THROMBOGENESIS INDUCED BY SUBENDOTHELIUM.

1987 ◽  
Author(s):  
P Hadvary ◽  
H R Baumgartner
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
Blood Platelets ◽  
Rabbit Aorta ◽  
Constant Infusion ◽  
Induced Aggregation

Platelet activating factor (PAF) is a very potent excitatory agonist of blood platelets but the physiological importance of this mediator in platelet thrombus formation is not known. We investigated the effect of two chemically unrelated selective inhibitors of PAF-induced platelet aggregation on thrombogenesis induced by rabbit aorta subendothelium (SE) using an ex vivo perfusion system.Ro 19-3704 is a highly potent inhibitor structurally related to PAF. This compound inhibits PAF-induced aggregation of rabbit platelets in platelet rich plasma in vitro competitively. Against 4 nM PAF, a concentration resulting in submaximal platelet aggre-gregation velocity, the IC50 was 70 nM. Inhibition was highly selective for PAF-induced aggregation, since aggregation induced by collagen (HORM, 5 yg/ml), ADP (1 yM) or thrombin (0.4 U/ml) was not inhibited even at a concentration as high as 10 yM. Bro-tizolam, a triazolobenzodiazepine reported to be a selective inhibitor of PAF-induced platelet activation, had in our system an IC50 of 200 nM. The selective benzodiazepine antagonist Ro 151788 was without effect on inhibition of PAF-induced platelet activation by brotizolam.Ro 19-3704 was given intravenously to rabbits as a bolus of 0.2 mg/kg followed by constant infusion of 0.02 mg/kg/min. This dosage provoked ex vivo a constant right shift ratio of the dose response curve for PAF-induced aggregation (RSR[PAF]) by a factor of 25 to 35. Brotizolam was given orally at a dose of 100 mg/ kg together with 300 mg/kg of Ro 15-1788 (to antagonize the central effects) 90 minutes before starting the perfusion experiment, resulting in a RSR[PAF] of 35 to 135. ADP induced platelet aggregation was not impaired by either compound. SE was exposed to the non-anticoagulated blood withdrawn from the carotid artery for 3 min at 2600 s-1 and for 20 min at 200 s-1 shear rate. Quantitative morphometric evaluation showed that SE coverage by platelets and by fibrin, thrombus area and thrombus height were all unchanged by the PAF antagonists at low and at high shear rates despite a very substantial inhibition of PAF-induced platelet aggregation. Therefore a major role of PAF in SE-induced thrombogenesis seems unlikely.

scholarly journals Biorheology of occlusive thrombi formation under high shear: in vitro growth and shrinkage

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Britt J. M. van Rooij ◽  
Gábor Závodszky ◽  
Alfons G. Hoekstra ◽  
David N. Ku
Keyword(s):  
Platelet Rich Plasma ◽  
Lag Time ◽  
Flow Chamber ◽  
High Shear ◽  
Shear Rates ◽  
High Shear Rates ◽  
Dependent Behavior ◽  
Platelet Aggregate ◽  
Contraction Angle

Abstract Occlusive thrombi formed under high flow shear rates develop very rapidly in arteries and may lead to myocardial infarction or stroke. Rapid platelet accumulation (RPA) and occlusion of platelet-rich thrombi and clot shrinkage have been studied after flow arrest. However, the influence of margination and shear rate on occlusive clot formation is not fully understood yet. In this study, the influence of flow on the growth and shrinkage of a clot is investigated. Whole blood (WB) and platelet-rich plasma (PRP) were perfused at high shear rates (> 3,000 s−1) through two microfluidic systems with a stenotic section under constant pressure. The stenotic section of the two devices are different in stenotic length (1,000 vs 150 μm) and contraction angle of the stenosis (15° vs 80°). In all experiments, the flow chamber occluded in the stenotic section. Besides a significantly increased lag time and decreased RPA rate for PRP compared to WB (p < 0.01), the device with a shorter stenotic section and steeper contraction angle showed a shear-dependent occlusion and lag time for both PRP and WB. This shear-dependent behavior of the platelet aggregate formation might be caused by the stenotic geometry.

ANTI-THROMBOTIC EFFECTS OF E-5510 IN EXPERIMENTAL THROMBOSIS MODELS

1987 ◽  
Author(s):  
T Fujimori ◽  
T Saeki ◽  
K Harada ◽  
M Sato ◽  
N Ohshima
Keyword(s):  
Blood Flow ◽  
Oral Administration ◽  
Guinea Pigs ◽  
Vessel Wall ◽  
Pulmonary Thromboembolism ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Platelet Thrombus

A new agent developed in our laboratory, 4-cyano-5,5-bis(4-methoxyphenyl)-4-pentenoic acid (E-5510), suppressed various human platelet functions in vitro. The compound also showed quite potent ex vivo anti-platelet effects in many experimentalanimals. It is well known that anti-platelet effects are not always parallel to anti-thrombotic effects. Thus, in order to predict the efficacy of E-5510 in thrombotic disorders, the anti-thrombotic effects were examined in 3 different animal models of thrombosis.(1) Anti-thrombotic effect in an extracorporeal shunt model Two hrs after oral administration of the drug to guinea pigs,an extracorporeal shunt between the right carotid artery and the left jugular vein was performed. The thrombus formation on a silk thread inserted in the shunt tube was quantitated by measuring the time from the onset of circulation to the stenosis of blood flow. E-5510 dose-dependently inhibited thrombus formation, the minimum effective dose being 0.03 mg/kg.(2) Effect on microthrombus formation in mesenteric arteriole In order to evaluate the effect on intravascular platelet thrombus formation, thrombosis was induced in vivo in mesenteric arteriole in guinea pigs with filtered light from a mercury lamp and an intravenous fluorescent dye in an intravital microscope system (M. Sato and N. Ohshima, Thromb. Res.,35, 319, 1984). The thrombus formation was quantitated by measuring the time taken for circulating platelets to begin to adhere to vessel wall and the time taken for blood flow to stop completely due to fully developed thrombus. Both the time required for platelet adhesion to vessel wall and for platelet thrombus formation were significantly prolonged after oral administration of E-5510.(3) Effect on pulmonary thromboembolism Acute pulmonary thromboembolism was induced in mice by a bolus intravenous injection of arachidonic acid, and mortality was determined 3 min later. E-5510 dose-dependently reduced pulmonary thromboembolic mortality, and its ED50 was 0.11 mg/kg. The results described above indicate thatE-5510 may have beneficial effects in clinical treatments of thrombotic disease.

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