ATP- and ADP-Induced Rat Platelet Aggregation: Significance of Plasma in ATP-Induced Aggregation

1979 ◽  
Vol 42 (05) ◽  
pp. 1580-1588 ◽  
Author(s):  
Ethan J Haskel ◽  
Kailash C Agarwal ◽  
Robert E Parks
Keyword(s):  
Creatine Kinase ◽  
Platelet Aggregation ◽  
Human Plasma ◽  
Platelet Rich Plasma ◽  
Creatine Phosphate ◽  
Rat Plasma ◽  
Human Platelets ◽  
Induced Aggregation ◽  
Aggregation Of Platelets ◽  
Rat Platelets

SummaryATP caused platelet aggregation in rat platelet-rich plasma (PRP) but in contrast strongly inhibited ADP-induced human platelet aggregation. ADP-induced aggregation of rat platelets suspended in human plasma was strongly inhibited by ATP, whereas human platelets in rat plasma were aggregated by ADP. The ATP analog β,γ-methylene ATP which is not dephosphorylated did not induce aggregation in rat PRP. Adenosine, AMP, 2- chloroadenosine, α,β-methylene ADP and β,γ-methylene ATP each inhibited ATP-induced aggregation of platelets in rat PRP to a similar extent as ADP-induced aggregation. A solution containing creatine kinase and creatine phosphate (which converts ADP to ATP) rapidly reversed both ADP- and ATP-induced aggregation in rat PRP; preincubation with this solution completely inhibited rat platelet aggregation induced by both ADP and ATP. Adenosine-8-14C-triphosphate ([14C]-ATP) conversion to [14C]-ADP was about five-fold faster in rat plasma than in human plasma. Addition of creatine phosphate to rat PRP strongly inhibited ATP-induced aggregation, while creatine or creatine kinase slightly potentiated aggregation by ATP. Creatine phosphate, creatine, or creatine kinase individually had minimal and varying effects on ADP-induced rat platelet aggregation. These results suggest that the observed phenomenon of ATP-induced aggregation in rat PRP is caused by a higher activity of creatine kinase in rat plasma than in human plasma, which converts the added ATP to ADP, a potent aggregator.

EFFECTS OF INHIBITORS ON COLLAGEN INDUCED PLATELET AGGREGATION IN SIX DIFFERENT SPECIES.

1987 ◽  
Author(s):  
J W C M Jansen
Keyword(s):  
Platelet Aggregation ◽  
Guinea Pig ◽  
Species Differences ◽  
Platelet Rich Plasma ◽  
Phosphodiesterase Inhibitors ◽  
Human Platelets ◽  
Cyclooxygenase Inhibitors ◽  
Pharmacological Studies ◽  
Aggregation Of Platelets ◽  
Rat Platelets

One approach to the development of antithrombotics is inhibition of platelet aggregation. The pharmacological approach often used is to test compounds on collagen induced platelet aggregation measured in platelet rich plasma. Therefore we have compared inhibitors with different mechanism of action on aggregation of platelets from six different species commonly used in pharmacological studies. Aggregation was induced with submaximal amounts of collagen (Hormone Chemie).Inhibitors of the cyclooxygenase system, aspirin and indomethacin, were very potent in inhibiting aggregation of platelets from humans guinea pig and dog (IC50 20-60 and 1-3 ¼M resp.). Aggregation of pig and rat platelets was poorly inhibited by both of these compounds (IC5: 700-900 ¼M), whereas platelets from mice showed intermediate sensitivety (IC50 ca.100 ¼M).The combined lipoxygenase/cyclooxygenase inhibitor BW755C, was extremely active on platelets of guinea pig (IC50 1 ¼M) and was poorly active in mice platelets (IC50 300 ¼M). In the other species the inhibitory activity ranged from 20-80 ¼M.The phosphodiesterase inhibitors, papaverine and BL3459 inhibited aggregation in all species (IC50 50-100 and 1-5 ¼M resp.). Dipyridamole inhibited aggregation also in all species but with lower activity (IC50 > 100 ¼M).Conclusion: remarkable species differences are present with respect to inhibition of collagen induced platelet aggregation by the various compounds e.g. rat and porcine platelet aggregation was hardly inhibited by cyclooxygenase inhibitors. The effects of the compounds on human platelets are comparable to the effects on canine plateletes.

The Effect of Red Blood Cells on the ADP-induced Aggregation of Human Platelets in Flow Through Tubes

1990 ◽  
Vol 63 (01) ◽  
pp. 112-121 ◽  
Author(s):  
David N Bell ◽  
Samira Spain ◽  
Harry L Goldsmith
Keyword(s):  
Platelet Aggregation ◽  
Shear Rate ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Platelet Rich Plasma ◽  
Mean Transit Time ◽  
White Blood Cells ◽  
Aggregate Size ◽  
Human Platelets ◽  
Induced Aggregation

SummaryThe effect of red blood cells, rbc, and shear rate on the ADPinduced aggregation of platelets in whole blood, WB, flowing through polyethylene tubing was studied using a previously described technique (1). Effluent WB was collected into 0.5% glutaraldehyde and the red blood cells removed by centrifugation through Percoll. At 23°C the rate of single platelet aggregtion was upt to 9× greater in WB than previously found in platelet-rich plasma (2) at mean tube shear rates Ḡ = 41.9,335, and 1,920 s−1, and at both 0.2 and 1.0 µM ADP. At 0.2 pM ADP, the rate of aggregation was greatest at Ḡ = 41.9 s−1 over the first 1.7 s mean transit time through the flow tube, t, but decreased steadily with time. At Ḡ ≥335 s−1 the rate of aggregation increased between t = 1.7 and 8.6 s; however, aggregate size decreased with increasing shear rate. At 1.0 µM ADP, the initial rate of single platelet aggregation was still highest at Ḡ = 41.9 s1 where large aggregates up to several millimeters in diameter containing rbc formed by t = 43 s. At this ADP concentration, aggregate size was still limited at Ḡ ≥335 s−1 but the rate of single platelet aggregation was markedly greater than at 0.2 pM ADP. By t = 43 s, no single platelets remained and rbc were not incorporated into aggregates. Although aggregate size increased slowly, large aggregates eventually formed. White blood cells were not significantly incorporated into aggregates at any shear rate or ADP concentration. Since the present technique did not induce platelet thromboxane A2 formation or cause cell lysis, these experiments provide evidence for a purely mechanical effect of rbc in augmenting platelet aggregation in WB.

Interactions of Purified Rat Factor VIII/von Willebrand Factor with Rat and Human Platelets – Effect of Albumin and Ristocetin

1984 ◽  
Vol 52 (01) ◽  
pp. 057-059 ◽  
Author(s):  
E Dejana ◽  
M Furlan ◽  
B Barbieri ◽  
M B Donati ◽  
E A Beck
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Rat Plasma ◽  
Human Platelets ◽  
High Concentrations ◽  
Low Sensitivity ◽  
Von Willebrand ◽  
Induced Aggregation ◽  
Willebrand Factor ◽  
Rat Platelets

SummaryRat platelets do not respond to ristocetin in their own plasma nor do they aggregate in the presence of bovine or porcine factor VIII von Willebrand factor (F VIII R:WF) or human F VIII R:WF in presence of ristocetin. However, rat plasma supports ristocetin induced aggregation of washed human platelets. In this study we report on purification of rat F VIII R:WF from cryoprecipitate. Similarly to porcine or bovine material, purified rat F VIII R:WF induced aggregation of human washed fixed platelets. This effect was enhanced by addition of ristocetin and was not modified by addition of albumin. Rat washed platelets were aggregated by ristocetin in the presence of rat or human F VIII R:WF provided that high concentrations of ristocetin are added in a system essentially free of extraneous proteins. Increasing concentrations of albumin dramatically reduced the ability of ristocetin to aggregate rat platelets while human platelet aggregation by human or rat F VIII R:WF was only moderately affected.These studies show that rat F VIII R:WF can interact with rat and human platelets. The lack of response of rat platelets to ristocetin in their own plasma is most likely due to a low sensitivity of rat platelets to this drug and to an inhibitory activity of plasma proteins on this reaction.

scholarly journals Preclinical studies of RUC-4, a novel platelet αIIbβ3 antagonist, in non-human primates and with human platelets

2019 ◽  
Vol 3 (2-3) ◽  
pp. 65-74 ◽  
Author(s):  
Spandana Vootukuri ◽  
Jihong Li ◽  
Mark Nedelman ◽  
Craig Thomas ◽  
Jiang-Kang Jiang ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
Human Platelets ◽  
Initial Slope ◽  
Receptor Blockade ◽  
St Segment Elevation ◽  
St Segment ◽  
Aggregation Inhibition ◽  
Induced Aggregation ◽  
Dose Dependent

AbstractIntroduction:We are developing the novel αIIbβ3 antagonist, RUC-4, for subcutaneously (SC)-administered first-point-of-medical-contact treatment for ST segment elevation myocardial infarction (STEMI).Methods:We studied the (1) pharmacokinetics (PK) of RUC-4 at 1.0, 1.93, and 3.86 mg/kg intravenous (IV), intramuscular (IM), and SC in non-human primates (NHPs); (2) impact of aspirin on RUC-4 IC50in human platelet-rich plasma (PRP); (3) effect of different anticoagulants on the RUC-4 IC50in human PRP; and (4) relationship between αIIbβ3 receptor blockade by RUC-4 and inhibition of ADP-induced platelet aggregation.Results:(1) All doses of RUC-4 were well tolerated, but animals demonstrated variable temporary bruising. IM and SC RUC-4 reached dose-dependent peak levels within 5–15 minutes, with T1/2s between 0.28 and 0.56 hours. Platelet aggregation studies in NHPs receiving IM RUC-4 demonstrated >80% inhibition of the initial slope of ADP-induced aggregation with all three doses 30 minutes post-dosing, with subsequent dose-dependent loss of inhibition over 4–5 hours. (2) The RUC-4 IC50for ADP-induced platelet aggregation was unaffected by aspirin treatment (40±9 nM vs 37±5 nM;p= 0.39). (3) The RUC-4 IC50was significantly higher in PRP prepared from D-phenylalanyl-prolyl-arginyl chloromethyl ketone (PPACK)-anticoagulated blood compared to citrate-anticoagulated blood using either thrombin receptor activating peptide (TRAP) (122±17 vs 66±25 nM;p= 0.05;n= 4) or ADP (102±22 vs 54±13;p<0.001;n= 5). (4) There was a close correspondence between receptor blockade and inhibition of ADP-induced platelet aggregation, with aggregation inhibition beginning with ~40% receptor blockade and becoming nearly complete at >80% receptor blockade.Discussion:Based on these results and others, RUC-4 has now progressed to formal preclinical toxicology studies.

Inhibition Of Platelet Aggregation, Malonyldialdehyde And Thromboxane Formation By Hydroperoxides Of Arachidonic Acid

1981 ◽  
Author(s):  
D Aharonv ◽  
J B Smith ◽  
M J Silver
Keyword(s):  
Arachidonic Acid ◽  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
Human Platelets ◽  
Lipoxygenase Pathway ◽  
Dose Dependent Fashion ◽  
Induced Aggregation ◽  
Layer Chromatography ◽  
Dose Dependent ◽  
Thromboxane Formation

The arachidonate hydroperoxides 12-HPETE and 15-HPETE were biosynthesized from arachidonic acid using partially purified human platelet lipoxygenase or soybean lipoxidase respectively, and isolated by thin layer chromatography. Both compounds inhibited the arachidonic acid- induced aggregation of washed human platelets, suspended in calcium-free Krebs Henseleit solution, in a dose dependent fashion at concentrations between 1 and 50 uM. No inhibition was seen with up to 100 uM of these hydroperoxides when platelet -rich plasma was used. 12-HPETE (in micromolar concentrations) inhibited the formation of both thromboxane B2 (radioimmunoassay) and malonyldialdehyde (spectrophotometrie assay) when washed platelets were incubated with arachidonic acid. The 12-hydroxide, 12-HETE also inhibited platelet aggregation and thromboxane formation, but was less potent than 12-HPETE. We suggest that arachidonate hydroperoxide generated in platelets via the lipoxygenase pathway modulates platelet aggregation induced by arachidonic acid by inhibiting thromboxane formation.

Effect of Heparin on Aggregation, Release Reaction and Thromboxane A2synthesis in Human Platelets

1979 ◽  
Author(s):  
H.Y.K. Chuang ◽  
S.F. Mohammad ◽  
R.G. Mason
Keyword(s):  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
Thromboxane A2 ◽  
Rabbit Aorta ◽  
Human Platelets ◽  
Appreciable Effect ◽  
Layer Chromatography ◽  
Aggregation Of Platelets ◽  
Platelet Functions

Studies on the effect of heparin on platelet functions have resulted in conflicting observations: heparin has been reported to cause aggregation of platelets, potentiate aggregation induced by various aggregating agents, or cause inhibition of aggregation. Using paritally purified heparin (beef lung or porcine mucosa) we observed that addition of heparin to citrated platelet rich plasma(C-PRP)potentiated the aggregation of platelets induced by ADP, epinephrine, or arachidonic acid. Presence of heparin in C-PRP results in complete inhibition of thrombin induced effects and partial inhibition of platelet aggregation induced by collagen. Presence of heparin in C-PRP also resulted in release of significantly higher concentrations of 14C-serotonin when platelets were challenged by appropriate aggregating agents. Those concentrations of heparin that resulted in potentiation of aggregation had no appreciable effect on c-AiMP or c-GMP levels of platelets. However, the presence of heparin results in a significant elevation of thromboxane A2 as determined by contraction of rabbit aorta or after conversion to thromboxane B2 by thin layer chromatography. These observations are of interest since increased production of thromboxane A2 in the presence of heparin may explain in part, the potentiation of platelet aggregation in vitro or thrombocytopenia observed frequently in patients receiving heparin intravenously Supported in part by grants HL22583 & 20679 from NHLBI of NIH.

scholarly journals Zinc-induced platelet aggregation is mediated by the fibrinogen receptor and is not accompanied by release or by thromboxane synthesis

Blood ◽  
1985 ◽  
Vol 66 (1) ◽  
pp. 213-219 ◽  
Author(s):  
P Heyns A du ◽  
A Eldor ◽  
R Yarom ◽  
G Marx
Keyword(s):  
Platelet Aggregation ◽  
Dense Body ◽  
Platelet Rich Plasma ◽  
Adenosine Monophosphate ◽  
Adenosine Diphosphate ◽  
Creatine Phosphate ◽  
Calcium Flux ◽  
Fibrinogen Receptor ◽  
Induced Aggregation

Abstract We demonstrate that zinc (0.1 to 0.3 mmol/L) induces aggregation of washed platelet suspensions. Higher concentrations (1 to 3 mmol/L) of zinc were needed to aggregate platelets in platelet-rich plasma obtained from blood anticoagulated with low-molecular-weight heparin, probably due to the binding of zinc to the plasma proteins. Zinc- induced aggregation of normal washed platelets required added fibrinogen and no aggregation occurred with thrombasthenic platelets or with normal platelets pretreated with a monoclonal antibody (10E5) that blocks the platelet fibrinogen receptor. These data indicate that the platelet membrane fibrinogen receptor-glycoproteins IIb and IIIa mediate the effect of zinc. Zinc-induced aggregation was blocked by the agent TMB-8, which interferes with the internal calcium flux, and by prostacyclin, which elevates platelet cyclic adenosine monophosphate levels. Zinc-induced aggregation was not accompanied by thromboxane synthesis or by the secretion of dense-body serotonin and was not affected by preexposure of platelets to acetylsalicylic acid. Experiments with creatine phosphate/creatine phosphokinase showed that the zinc effect on platelets was independent of extracellular adenosine diphosphate (ADP). Zinc had an additive effect when platelet aggregation was stimulated with subthreshhold concentrations of collagen or ADP. Together with the known effects of nutritional zinc on in vivo bleeding, on platelet aggregation, and on lipid metabolism, the results suggest that zinc may have an important bearing on normal hemostasis, thrombosis, and atherosclerosis.

Inhibition of ADP-Induced Platelet Aggregation by Adenosine Tetraphosphate

1976 ◽  
Vol 36 (02) ◽  
pp. 388-391 ◽  
Author(s):  
Margaret J. Harrison ◽  
R Brossmer
Keyword(s):  
Platelet Aggregation ◽  
Inhibitory Action ◽  
Platelet Rich Plasma ◽  
Release Reaction ◽  
Induced Aggregation ◽  
Aggregation Of Platelets

SummaryIn contrast to previous reports, highly purified adenosine tetraphosphate (AP4) does not induce the aggregation of platelets but inhibits the aggregation and release reaction in platelet-rich plasma promoted by ADP. The inhibitory action of AP4 on the aggregation by ADP is compared with that of AMP and ATP. The data presented suggest a competitive manner of inhibition of the ADP-induced aggregation by AP4.

Effects of Lead Acetate on Platelet Aggregation, Ultrastructure and Serotonin Release

1971 ◽  
Vol 26 (03) ◽  
pp. 455-466 ◽  
Author(s):  
R. B Davis ◽  
G. C Holtz
Keyword(s):  
Platelet Aggregation ◽  
Platelet Function ◽  
Lead Acetate ◽  
Platelet Rich Plasma ◽  
Adenosine Diphosphate ◽  
Blood Platelet ◽  
Human Platelets ◽  
Serotonin Release ◽  
Aggregation Of Platelets

SummaryThe effects of lead on blood platelet function and ultrastructure have been investigated. Lead acetate was injected intravenously in 27 rats and was added to rat and human platelet rich plasma in vitro. In vitro studies showed that concentrations of 2.5 × 10-3 M lead acetate reduced or blocked aggregation of rat and human platelets by adenosine diphosphate, collagen, and thrombin. Radioactive serotonin release from human platelets was inhibited by 10-4 M lead acetate. One hour after the injection of lead, platelet aggregation by thrombin was reduced, but platelet aggregation by adenosine diphosphate and collagen showed little change. Three days after lead, aggregation of platelets by collagen and thrombin was blocked and aggregation by adenosine diphosphate reduced. Thrombocytopenia was present 4 days after intravenous lead acetate. Electron micrographs of platelets showed that the mean number of mitochondria per platelet was increased, whereas alpha granules were reduced. Dense bodies were not significantly changed. Lead acetate affects platelet function in concentrations reported in human bone marrow in lead poisoning, and may relate to the binding of free sulfhydryl groups by lead.

Effects of Ticlopidine and Indobufen on Platelet Aggregation Induced by A23187 and Adrenaline in the Presence of Different Anticoagulants

1989 ◽  
Vol 17 (6) ◽  
pp. 514-520 ◽  
Author(s):  
C. Cimminiello ◽  
M. Milani ◽  
T. Uberti ◽  
G. Arpaia ◽  
G. Bonfardeci
Keyword(s):  
Platelet Aggregation ◽  
Platelet Function ◽  
Sodium Citrate ◽  
Platelet Rich Plasma ◽  
Antiplatelet Agents ◽  
Ionophore A23187 ◽  
Tyrode Solution ◽  
Low Levels ◽  
Induced Aggregation ◽  
Aggregation Of Platelets

As Ca2+ is known to play a fundamental role in platelet function, the effect of combining two platelet aggregating agents (adrenaline and the ionophore A23187) with different effects on Ca2+ was studied at levels subthreshold for aggregation using platelet-rich plasma from eight atherosclerotic patients. Adrenaline lowered the A23187 threshold required to induce aggregation. The effects of treating patients with the antiplatelet agents, indobufen and ticlopidine, on A23187 and adrenaline induced aggregation of platelets prepared in hirudin or sodium citrate was also evaluated. Aggregation was also studied using platelets resuspended in Ca2+-free and Ca2+-enriched Tyrode solution. Before treatment hirudin treated platelet-rich plasma, which has physiological extraplatelet Ca2+ levels, was more sensitive to A23187 and adrenaline than was citrated platelet-rich plasma, which has suppressed Ca2+ levels. Ticlopidine significantly raised the concentration of A23187 required to induce aggregation in citrated but not hirudin treated platelet-rich plasma. Indobufen did not significantly affect A23187 induced aggregation. Ticlopidine acts by inhibiting the glycoprotein IIb – IIIa complex on the platelet membranes. Low levels of extracellular Ca2+ and ticlopidine may act synergistically to reduce the aggregatory response of stimulated platelets.

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