Comparison of in Vivo and in Vitro Effects of Platelet Function Suppressing Drugs

Mapping Intimacies ◽

10.1055/s-0039-1689321 ◽

1975 ◽

Cited By ~ 2

Author(s):

M. Buchanan ◽

J. Hirsh

Keyword(s):

Platelet Function ◽

Prosthetic Heart Valve ◽

Inhibitory Effect ◽

Potent Effect ◽

Platelet Survival ◽

In Vitro Effects ◽

Antithrombotic Efficacy ◽

Induced Aggregation

Comparison of in vivo and in vitro platelet function tests to assess the antithrombotic efficacy of drugs which suppress platelet function have been contradictory. For example, aspirin (ASA) has a potent effect in vitro but little effect when tested on platelet survival in prosthetic heart valve replacements whereas dipyridamole (DIP) has little or no effect in vitro but a marked effect on platelet survival. We have compared in parallel the in vivo and in vitro effects of a number of drugs which suppress platelet function in an animal model. Rabbits were infused with homologous 51Cr-labelled platelets and then given either ASA (10-200 mg/kg), DIP (1-20 mg/kg) or sulfinpyrazone (SUF) (30-200 mg/kg) intravenously. One hour later PRP from each rabbit was tested by ADP and collagen-induced aggregation. Then each rabbit was infused with an identical final concentration of collagen and the subsequent recovery of 51Cr platelet radioactivity was monitored. In untreated rabbits collagen infusion produced 30% reduction of 51Cr platelets which returned to within 85% of a procollagen level by 5 min. ASA (⪖ 10 mg/kg) inhibited in vitro collagen-induced aggregation while a dose of 100 mg/kg of ASA was necessary to achieve the same inhibitory effect in vivo. On the other hand, DIP (1-20 mg/kg) had no inhibitory effect on in vitro platelet aggregation whereas it inhibited aggregation in vivo. The results of SUF were similar in vitro and in vivo. These results suggest that the effectiveness of drugs on platelet function may be affected by centrifugation, addition of anticoagulant or removal of red cells. This may explain the discrepancies reported between the in vivo and in vitro effectiveness of such drugs.

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‘In Vivo’ and ‘In Vitro’ Effects of Some Vasoactive Drugs on Platelet Function and on Coagulation/Fibrinolysis System

10.1055/s-0039-1680783 ◽

1977 ◽

Author(s):

A.G. Dettori ◽

O. Ponari ◽

C. Manotti ◽

A. Megha ◽

M. Pini

Keyword(s):

Platelet Function ◽

Vasoactive Drugs ◽

Platelet Function Tests ◽

Lysis Time ◽

Low Concentrations ◽

Euglobulin Lysis Time ◽

In Vitro Effects ◽

Induced Aggregation

Three substances widely used as vasoactive drugs are known to have an inhibiting effect on platelet aggregation ‘in vitro’. We investigated the changes induced on thrombelastogram, routine clotting tests, euglobulin lysis time (ELT), platelet count, aggregation, and adhesiveness by i, v. administration of these drugs to man. The same indices were also studied ‘in vitro’ by adding comparable concentrations of the substances to human blood or plasma.Aminophilline did not produce any significant variation in ADP-or collagen-induced aggregation either ‘in vitro’ (50 to 200 μg/ml) or ‘in vivo’ (240 mg). A trend to disaggregation was seen only in a few cases. Shorter ELT were found 30 and 120 minutes after injection.A papaverine derivative (Metaverinum, 150 mg) showed a similar ‘in vivo’ pattern: minor changes in platelet function tests and a moderate activation of fibrinolysis were seen. The drug acted ‘in vitro’ as a powerful inhibitor of aggregation (from 30 µg/ml)while fibrinolysis was only activated at the highest concentration (120 µg/ml).Bencyclan, capable of inhibiting platelet function ‘in vitro’ at very low concentrations (0.25µM) did not show similar effects ‘in vivo’ (50 mg) apart from a reduced platelet adhesiveness to glass.

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ALTERATION OF THE RESPONSE OF PLATELETS TO SURFACE STIMULI BY PYRAZOLE COMPOUNDS

Journal of Experimental Medicine ◽

10.1084/jem.126.1.171 ◽

1967 ◽

Vol 126 (1) ◽

pp. 171-188 ◽

Cited By ~ 112

Author(s):

M. A. Packham ◽

E. S. Warrior ◽

M. F. Glynn ◽

A. S. Senyi ◽

J. F. Mustard

Keyword(s):

Platelet Function ◽

Blood Platelets ◽

Platelet Response ◽

Thrombin Inhibition ◽

Platelet Survival ◽

Coated Surfaces ◽

Antigen Antibody ◽

Induced Aggregation

Sulfinpyrazone and phenylbutazone block the aggregating action of collagen, antigen-antibody complexes, and gamma globulin-coated surfaces on blood platelets. These drugs do not block the action of ADP or thrombin. Inhibition of surface-induced aggregation appears to be the result of a decreased response of the platelets to surface stimuli, giving rise to diminished release of platelet constituents, such as ADP and serotonin. The intravenous infusion of these drugs produced results similar to those found in the in vitro experiments. Administration of phenylbutazone in doses sufficient to produce marked suppression of the platelet-collagen reaction impaired hemostatic plug formation at the ends of transected mesenteric vessels in rabbits. Since platelet function is considered a factor influencing platelet survival, the effect of phenylbutazone on platelet survival was examined. It was found that phenylbutazone prolonged platelet survival to more than twice the normal time and reduced platelet turnover by nearly 50%. These studies show that drugs which suppress platelet response to surface stimuli alter platelet function in vivo.

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Sex related differences in platelet function: the effect of aspirin

Blood ◽

10.1182/blood.v59.3.625.bloodjournal593625 ◽

1982 ◽

Vol 59 (3) ◽

pp. 625-627

Author(s):

JG Kelton ◽

CJ Carter ◽

A Santos ◽

J Hirsh

Keyword(s):

Platelet Function ◽

Animal Experiments ◽

Inhibitory Effect ◽

Thromboxane B2 ◽

Equal Dose ◽

Before And After ◽

Male Patients ◽

Induced Aggregation

There is evidence from clinical studies and animal experiments that aspirin has a greater antithrombotic activity in males compared to females. We investigated platelet function in vitro and in vivo in rabbits before and after the administration of a dose of aspirin (5 mg/kg) which inhibited collagen stimulated thromboxane B2 generation. Infusion of collagen into untreated animals resulted in a 38 +/- 4% (m +/- SE, n = 13) decrease in platelet count (assessed by whole blood radioactivity) in the male animals, and a 27 +/- 3% (m +/- SE, n = 13) in the female animals. Pretreatment with aspirin resulted in a significant inhibitory effect in the male but not the female animals (p less than 0.05). The male animals had significantly greater thromboxane B2 generation in vivo than did the female animals following an equal dose of collagen (males, 2.64 +/- 0.7 ng/ml thromboxane B2, n = 14; females, 1.67 +/- 0.4 ng/ml thromboxane B2, n = 15, p less than 0.05). In contrast no sex related difference in the inhibitory effect of aspirin on maximal collagen induced aggregation was found when platelets were studied in vitro. The greater reactivity of male patients in vivo may be accounted for by the observed increase in thromboxane B2 generation. This might also explain the greater thrombotic tendency of males, and the observed difference in the antithrombotic effect of aspirin in males and females.

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Sex related differences in platelet function: the effect of aspirin

Blood ◽

10.1182/blood.v59.3.625.625 ◽

1982 ◽

Vol 59 (3) ◽

pp. 625-627 ◽

Cited By ~ 15

Author(s):

JG Kelton ◽

CJ Carter ◽

A Santos ◽

J Hirsh

Keyword(s):

Platelet Function ◽

Animal Experiments ◽

Inhibitory Effect ◽

Thromboxane B2 ◽

Equal Dose ◽

Before And After ◽

Male Patients ◽

Induced Aggregation

Abstract There is evidence from clinical studies and animal experiments that aspirin has a greater antithrombotic activity in males compared to females. We investigated platelet function in vitro and in vivo in rabbits before and after the administration of a dose of aspirin (5 mg/kg) which inhibited collagen stimulated thromboxane B2 generation. Infusion of collagen into untreated animals resulted in a 38 +/- 4% (m +/- SE, n = 13) decrease in platelet count (assessed by whole blood radioactivity) in the male animals, and a 27 +/- 3% (m +/- SE, n = 13) in the female animals. Pretreatment with aspirin resulted in a significant inhibitory effect in the male but not the female animals (p less than 0.05). The male animals had significantly greater thromboxane B2 generation in vivo than did the female animals following an equal dose of collagen (males, 2.64 +/- 0.7 ng/ml thromboxane B2, n = 14; females, 1.67 +/- 0.4 ng/ml thromboxane B2, n = 15, p less than 0.05). In contrast no sex related difference in the inhibitory effect of aspirin on maximal collagen induced aggregation was found when platelets were studied in vitro. The greater reactivity of male patients in vivo may be accounted for by the observed increase in thromboxane B2 generation. This might also explain the greater thrombotic tendency of males, and the observed difference in the antithrombotic effect of aspirin in males and females.

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The Effect of Barbituric Acid Derivatives on Platelet Function in Vitro and in Vivo

Thrombosis and Haemostasis ◽

10.1055/s-0038-1649080 ◽

1973 ◽

Vol 30 (02) ◽

pp. 315-326

Author(s):

J. Heinz Joist ◽

Jean-Pierre Cazenave ◽

J. Fraser Mustard

Keyword(s):

Platelet Aggregation ◽

Platelet Function ◽

Barbituric Acid ◽

Platelet Rich Plasma ◽

Inhibitory Effect ◽

Primary Response ◽

Sodium Pentobarbital ◽

Barbituric Acid Derivatives

SummarySodium pentobarbital (SPB) and three other barbituric acid derivatives were found to inhibit platelet function in vitro. SPB had no effect on the primary response to ADP of platelets in platelet-rich plasma (PRP) or washed platelets but inhibited secondary aggregation induced by ADP in human PRP. The drug inhibited both phases of aggregation induced by epinephrine. SPB suppressed aggregation and the release reaction induced by collagen or low concentrations of thrombin, and platelet adherence to collagen-coated glass tubes. The inhibition by SPB of platelet aggregation was readily reversible and isotopically labeled SPB did not become firmly bound to platelets. No inhibitory effect on platelet aggregation induced by ADP, collagen, or thrombin could be detected in PRP obtained from rabbits after induction of SPB-anesthesia.

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Effect Of Solvents On Indium-111 Oxine Transport In Human Platelets And On Platelet Function In Vitro

10.1055/s-0038-1653277 ◽

1981 ◽

Author(s):

T Tsukada

Keyword(s):

Platelet Function ◽

Kinetic Constant ◽

Inhibitory Effect ◽

Human Platelets ◽

Polysorbate 80 ◽

Autologous Plasma ◽

Indium 111 ◽

Induced Aggregation ◽

Water Space

Mechanism of Indium-111 oxine(In) transport in human platelets in buffered saline and the effect of In-labeling on platelet function were studied using In dissolved in 25% of ethanol in saline (In-ES) or 0.01% of polysorbate 80 in HEPES buffer(In-PH). Increase in temperature up to 37° C progressively enhanced the transport of In-ES, while transport of In-PH reached to plateau at 15°C. A states of equilibrium was not reached during 2 hr incubation at 22°C in In-ES. Uptake of In-PH reached to plateau after only 15 min of incubation. Distribution of In taken up by platelets in InES was 57% in cytosol and 27% in stroma, while in In-PH 69% in stroma and 22% in cytosol. 88% of In in cytosol was bound to lipids(46% in cholesterol and 27% in PS+PI). 82% of In in stroma was found in PS+PI fraction.The fact that the ratio of free In between the platelet water space and the outside medium after 30 min of incubation at up to 0.1 uM of In exceeded unity, suggests satura- , ble component of In transport prevails at this concentration in In-ES and In-PH. Kinetic constant could be calculated, Kt= 2nM, Vmax= 2.5 pmol/min/ml in In-ES, and Kt= InM, Vmax=0.7 pmol/min/ml in In-PH.Elution of In from radiolableled platelets in autologous plasma incubated at 37°C for 5 hr was less than 10% in the case of In-ES and 56% in the case of In-PH. Less than 3% of labeled-In was eluated from platelets in collagen-induced aggregation and 4-7% of In was eluated in thrombin-induced aggregation.Although 0.3% of ethanol and/or 6nM of oxine have no inhibitory effect of platelet aggregation, collagen-induced aggregation and release reaction of In-labeled platelet was impaired. 0.003% of polysorbate 80 itself abolished completely the aggregability of platelets by collagen or thrombin.It is concluded In-PH is unsuitable for platelet labeling. In-111 oxine also seems to have problems which Cr-51 has, i.e. inhomogenous distribution of In in a platelet population, elution of In from labeled platelets in circulation.

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Inhibition of Human Platelet Aggregation by the Proteolytic Effect of Streptokinase (SK). Role of the Factor VIII Related Antigen

10.1055/s-0039-1689380 ◽

1975 ◽

Cited By ~ 1

Author(s):

R. Castillo ◽

S. Maragall ◽

J. A. Guisasola ◽

F. Casals ◽

C. Ruiz ◽

...

Keyword(s):

Platelet Aggregation ◽

Factor Viii ◽

Platelet Rich Plasma ◽

Gel Filtration ◽

Inhibitory Effect ◽

Factor Viii Related Antigen ◽

Human Factor Viii ◽

Induced Aggregation

Defective ADP-induced platelet aggregation has been observed in patients treated with streptokinase. This same effect appears “in vitro” when adding SK to platelet rich plasma (PRP). Classic hemophilia and normal platelet poor plasmas (PPP) treated with SK inhibit the aggregation of washed platelets; plasmin-treated normal human serum also shows an inhibitory effect on platelet aggregation. However, von Willebrand SK-treated plasmas do not inhibit the aggregation of washed platelets. The same results appear when plasmas are previously treated with a rabbit antibody to human factor VIII.This confirms that the antiaggregating effect is mainly linked to the digested factor VIII related antigen.The inhibition of ADP-induced platelet aggregation has been proved in gel filtration-isolated and washed platelets from SK-treated PRP.Defective ristocetin-induced platelet aggregation has also been observed- This action does not appear in washed platelets from SK-treated PRP in presence of normal PPP, but it does in presence of SK-treated PPP, which suggests that the inhibition of the ristocetin-induced aggregation is due to the lack of factor VIII and not to the factor VIII-related products.Heparin, either “in vivo” or “in vitro”, has corrected the antiaggregating effect of SK.

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Pharmacological Evaluation of Ticlopidine, A Novel Inhibitor of Platelet Function

10.1055/s-0039-1680487 ◽

1977 ◽

Author(s):

M. Johnson ◽

P. L. Walton ◽

R. C. Cotton ◽

C. J. L. Strachau

Keyword(s):

Survival Time ◽

Platelet Function ◽

Thrombus Formation ◽

Antiplatelet Agent ◽

Dose Effect ◽

Onset Of Action ◽

Duration Of Action ◽

Platelet Survival ◽

Induced Aggregation

Ticlopidine (T), 5-(2-chlorobenzyl)-4,5,6,7-tetrahydrothieno(3,2-C)pyridine hydrochloride (a product of Parcor Research) has been evaluated as an antiplatelet agent in various animal species and in human volunteers. T was inactive in vitro, but inhibited platelet aggregation induced by ADP, collagen, thrombin, arachidonic acid and prostaglandin (PG) endoperoxide, when administered orally to mice, rats, rabbits, guinea pigs, pigs, dogs and baboons. Platelet adhesiveness was reduced but platelet survival time was normal in treated animals. Basal PG synthesis and platelet ultra-structure were unaffected by T.T protected against acute thrombocytopenia and death from pulmonary embolism induced by i. v. injection of ADP or collagen. Thrombus formation in experimental models of extra-corporeal circulation and deep venous thrombosis was also impaired.In man, a single oral dose of 500mg was shown to be a potent inhibitor of ADP, collagen, adrenaline, ristocetin, bovine fibrinogen and 5HT-induced aggregation. A dose-effect relationship was apparent, 250 and 500mg resulting in ~47 and 75% inhibition of ADP-induced aggregation respectively. Inhibition was sustained by chronic daily dosing.There was a delay in the onset of action of T in vivo, but which then persisted after withdrawal for at least 48 hours, with no evidence of rebound hyperactivity. The duration of action of T correlated with platelet survival time, suggesting an irreversible modification of platelet function. T is a potent platelet inhibitor, exhibiting a novel mode of action and lack of agonist specificity, which may be of value in the treatment of thrombotic conditions.

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Inhibition of platelet function by organic nitrate vasodilators

Blood ◽

10.1182/blood.v55.4.649.bloodjournal554649 ◽

1980 ◽

Vol 55 (4) ◽

pp. 649-654

Author(s):

AI Schafer ◽

RW Alexander ◽

RI Handin

Keyword(s):

Myocardial Ischemia ◽

Platelet Function ◽

Coronary Circulation ◽

Platelet Reactivity ◽

Inhibitory Effect ◽

Organic Nitrate ◽

Artery Disease ◽

In Vitro Effects ◽

External Calcium

There is evidence that platelet activation in the coronary circulation may be important in the pathogenesis of myocardial ischemia. Since organic nitrate vasodilators are commonly used in coronary artery disease, we have studied the in vitro effects of these drugs on platelet function. Nitroglycerin, isosorbide dinitrate, and their biotransformation product, inorganic nitrite, inhibited platelet aggregation with collagen, epinephrine, arachidonate, and ionophore, and blocked both primary and secondary aggregation in response to ADP. Nitroglycerin was studied in more detail. Its inhibitory effect was reversible and not dependent on external calcium concentration. It inhibited arachidonic acid oxygenation as measured by the arachidonate- induced oxygen burst and malonaldehyde production. These effects were not due to an increase in intracellular cyclic AMP. This unusual generalized inhibition of platelet function by nitroglycerin possibly contributes to its beneficial effect in myocardial ischemia in part by attenuating platelet reactivity in the coronary circulation.

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