Arachidonate Metabolism And Platelet Disaggregation (DA)-Reaggregation (RA)

1981 ◽  
Author(s):  
Gundu H R Rao ◽  
James G White

Previous work has shown that platelets irreversibly aggregated by ADP or thrombin (T) can be dissociated by various agents and that the refractory state of disaggregated cells can be reversed immediately by treatment with epinephrine (E). In the present study we have evaluated the influence of drugs which affect different steps in the process of prostaglandin (PG) synthesis on platelet DA-RA. Aspirin and indomethacin did not cause DA of platelets in the process of aggregation nor did they prevent reversal of the refractory state by E and subsequent RA of previously dissociated platelets. Imidazole, which inhibits conversion of endoperoxide to thromboxane A2, also failed to influence DA or restoration of sensitvity and RA of disaggregated platelets. On the other hand, chemicals which interfere with release of AA from the membrane of activated platelets, such as mepacrine, chlorpromazine and trifluoperazine, caused rapid DA. Products of PG synthesis, such as PGE1, PGD2 and PGI2, which usually inhibit platelet aggregation, also caused rapid DA. The refractory state of platelets dissociated from aggregates by most of these agents could be reversed by E treatment. However, trifluoperazine disaggregated platelets could be reaggregated only by the combination of E and AA. Agents which block the a-adrenergic receptors did not cause dissociation of aggregating platelets, but prevented correction of the refractory state of dissociated platelets by E. Thus interference with AA release, even after aggregation, can cause DA of clumped platelets, but blockade of peroxidase, cyclo-oxygenase and thromboxane synthetase do not cause reversal once it is in progress. A membrane linked mechanism associated with AA availability, but not metabolism, regulates DA and restoration of membrane sensitivity for RA.

1981 ◽  
Author(s):  
C Bonne ◽  
B Martin

“Large spectrum”anti-aggregating activity could be only achieved by agents which increased the c AMP content of platelets. Cyclo-oxygenase inhibitors could only block the Thromboxane (Tx)- dependent pathway of platelet aggregation. Conversely, Tx-synthetase inhibitors could deviate the endoperoxides metabolism to anti-aggregating prostaglandins in particular in the presence of vascular tissues. In this study we have investigated the effect of CBS634 (1-(3-hydroxy- 1 - octenyl)-imidazole nicotinic ester, dichlorhydra- teD, a potent inhibitor of T×A2 synthesis, both on the production of anti-aggregating prosta- glandins and on the simultaneous c AMP synthesis in platelets.Rat aorta fragments pretreated with aspirin were incubated with rat platelet rich plasma in the presence or absence of tested compound, c AMP, T×B2, PGE2 and 6-keto-PGFF1α were determined by radioimmunoassays.In the presence of CBS63U (50μM), T×B2 formation was reduced from 17 ± 2 to 0.2 ng/ml/min. In parallel, PGE production in controls was 1.4 ± 0.6 and 8 ± 1 ng/ml/min in the presence of the drug. On the other hand, 6-keto-PGF1α formation, very low in controls, rose to 4 ±1.2 ng/mV min in the presence of CBS63U. Radiochemical assays performed with c14C)-arachidonate confirmed that metabolic deviation. The increased level of c AMP formed in the presence of T×A2-synthetase inhibitor supports the hypothesis that such a drug could present a “large spectrum”antiaggregating activity.


2010 ◽  
Vol 638 (1-3) ◽  
pp. 5-12 ◽  
Author(s):  
Fernanda C.F. Brito ◽  
Arthur E. Kummerle ◽  
Claire Lugnier ◽  
Carlos A.M. Fraga ◽  
Eliezer J. Barreiro ◽  
...  

1981 ◽  
Author(s):  
K D Butler ◽  
E D Maguire ◽  
A A Turnbull ◽  
R B Wallis ◽  
A M White

The role of thromboxane A2 (TXA2) in haemostasis was investigated with the use of a selective inhibitor of platelet thromboxane synthetase used in conjunction with radioimmunoassay of thromboxane B2 (TXB2). N-Carboxyheptylimidazole is such an inhibitor having no effect on platelet cyclooxygenase. An oral dose of this substance (10 mg/kg) to rats resulted in 85% (P < 0.001) suppression of platelet TXB2 production induced by collagen ex vivo while the ED50 and maximum rate of platelet aggregation were unchanged. It also caused a prolongation of tail bleeding time from 153±13 to 284±22 secs (P < 0.01). The thrombocytopenia resulting from the Arthus reaction in rats was unchanged, and the prothrombin and activated partial thromboplastin coagulation times were not affected by either 10 or 30 mg/kg p.o. It is concluded that the role of TXA2 in prevention of rat tail bleeding is not as an activator of platelet aggregation or blood coagulation. It is more likely that TXA2 prevents bleeding via its potent vaso-constricting properties. In addition the increased bleeding time may be due to change in the equilibrium of other vasoactive prostanoids.


1980 ◽  
Vol 238 (1) ◽  
pp. H87-H92 ◽  
Author(s):  
K. Schror ◽  
E. F. Smith ◽  
M. Bickerton ◽  
J. B. Smith ◽  
K. C. Nicolaou ◽  
...  

Pinane thromboxane A2 (PTA2), a thromboxane A2 analog has been shown to antagonize the vasoconstriction and platelet aggregation induced by thromboxane A2, in addition to specifically inhibiting thromboxane synthetase. Because thromboxane A2 generation would be detrimental in acute myocardial ischemia (MI) by both decreasing coronary blood flow and increasing platelet aggregation, inhibition of thromboxane production and action may be beneficial in myocardial ischemia. In pentobarbital-anesthetized cats, the left anterior descending coronary artery was ligated, and PTA2 (0.5 mumol . kg-1 . h-1) or a Na2CO3 vehicle was infused 30 min post-MI for 270 min. Compared to vehicle-treated MI cats, PTA2 prevented the increase in plasma thromboxane levels seen at 2 through 5 h (P less than 0.005 at 2 through 5 h) and prevented the large increase in plasma CK activities at 4 and 5 h (P less than 0.025). In addition, PTA2 treatment abolished the differences in myocardial CK activities between ischemic and nonischemic regions and prevented the decrease in percent-bound cathepsin D in the ischemic region. Moreover, ECG analysis revealed a decreased incidence of premature beats in PTA2-treated MI cats as compared to MI-vehicle cats. In summary, these data indicate that PTA2 protects the ischemic myocardium and provide further evidence that inhibition of thromboxane formation, in addition to antagonism of its activity, is beneficial during the early stages of acute myocardial ischemia.


1981 ◽  
Author(s):  
G J Johnson ◽  
G H R Rao ◽  
J G White

Epinephrine (E) potentiates arachidonate (A)-induced aggregation of human platelets. A-insensitive dog platelets (AIP), that form thromboxane A2 (T) but do not aggregate when stirred with A alone, aggregate when exposed to E + A. Therefore, we studied the effect of E on T-stimu- lated human platelet aggregation. AIP stirred with A formed T which was confirmed by TLC. 1/100 to 1/200 volume of AIP was removed 30 sec. after A, and transferred to gel- filtered, aspirin-incubated human platelets. Recipient platelet aggregation was proportional to the volume of AIP transferred. The addition of the thromboxane synthetase inhibitor, Azo Analog I, abolished the aggregating activity of AIP. Transfer of an aliquot of AIP that was inadequate to aggregate human gel-filtered, aspirin-incubated platelets resulted in irreversible aggregation in the presence of ≥0.5nM E. E potentiated aggregation when added 3 min. before but not 3 min. after aliquot transfer. T-stimulated aggregation was abolished by the T-antagonist, 13 azapro- stenoic acid (APA), but E added after APA and before T restored aggregation. E potentiation of T-stimulated aggregation was abolished by prior exposure to equimolar yohimbine, dihydroergocryptine and phentolamine, agents that bind to alpha2 adrenergic receptors, but not by prazosin an alpha1 antagonist. Higher concentrations of E reversed the inhibitory effects of the alpha2 adrenergic agents. All of these agents in higher concentrations (1-100μM) also blocked aggregation induced by T alone. Therefore T-induced platelet aggregation is potentiated by E, in concentrations attained in vivo, by a mechanism linked to platelet alpha adrenergic receptors. Platelet alpha2 receptors have a close functional relationship to the postulated T receptor. E may initiate platelet aggregation in vivo when T is formed in quantities inadequate to alone induce aggregation.


2004 ◽  
Vol 91 (06) ◽  
pp. 1158-1167 ◽  
Author(s):  
Chikaho Toma ◽  
Takako Nishiya

SummaryLiposomes with a covalently bound synthetic peptide containing the dodecapeptide sequence HHLGGAKQAGDV, the putative platelet interaction site at γ400-411 of fibrinogen (dodecapeptide-liposomes), were prepared. These liposomes enhanced platelet aggregation and specifically adhered to platelets activated on the collagen surface. Dodecapeptide-liposomes released encapsulated materials upon interacting with platelets activated on the collagen surface.The rate of content release was dependent on the peptide surface density, indicating that the interaction between the dodecapeptide-liposomes and platelets activated on the collagen surface induces clustering of the surfacecoupled ligands at the binding site on the receptor matrix to facilitate release of the internal contents through the liposome membranes. The level of lipid mixing between the dodecapeptide-liposomes and platelets activated on the collagen surface was relatively low, however it was increased in liposome preparations containing octa-arginine, the (R)8GDV sequence, while content release was maintained at the same level as that of the dodecapeptide-liposomes. The level of content release and lipid mixing for liposome preparations containing the RGD sequence as a ligand (RGD-liposomes) upon interacting with platelets activated on the collagen surface was extremely low. Both the level of the content release and lipid mixing, however, were enhanced in liposome preparations containing octa-arginine, the (R)8RGD sequence. Dodecapeptide-liposomes and RGD-liposomes were not internalized by activated platelets. On the other hand, liposomes containing (R)8PPQ, (R)8RGD, or (R)8GDV were internalized by activated platelets, and the extent of internalization was inversely related to ligand affinity to the target.


Blood ◽  
1987 ◽  
Vol 70 (2) ◽  
pp. 475-483 ◽  
Author(s):  
K Niiya ◽  
E Hodson ◽  
R Bader ◽  
V Byers-Ward ◽  
JA Koziol ◽  
...  

Platelet activation altered the binding of three monoclonal antibodies (monovalent Fab' fragment) directed against the glycoprotein (GP) IIb/IIIa complex. An increased binding of two- to threefold occurred after stimulation with thrombin or phorbol myristate acetate (PMA), with slight but significant increase in the dissociation constants (Kd) of two antibodies (LJ-CP8 and LJ-P9). In contrast, no statistically significant changes were observed with ADP-stimulated platelets. The increased binding of LJ-CP3, but not of the other two antibodies, to activated platelets decreased by 30% to 40% in the presence of EDTA at 22 to 25 degrees C. Platelets stimulated by thrombin or PMA bound more fibrinogen than did those stimulated by ADP, and significant differences in the extent but not in the affinity of fibrinogen binding were observed with various platelet agonists. When the pool of GP IIb/IIIa molecules exposed on the surface of unstimulated platelets was reacted with the monoclonal antibody LJ-CP3 to block ADP-induced fibrinogen binding and platelet aggregation, stimulation with thrombin or PMA still induced substantial binding of antibody and fibrinogen, and aggregation ensued. Therefore, platelets exposed to “strong” agonists exhibit an increased number of surface-oriented epitopes associated with GP IIb/IIIa. The GP IIb/IIIa molecules bearing these newly exposed epitopes are functional in that they can bind fibrinogen and mediate platelet aggregation.


1987 ◽  
Author(s):  
K Niiya ◽  
E Hodson ◽  
R Bader ◽  
V Byers-Ward ◽  
E F Plow ◽  
...  

Platelet stimulation altered the binding of three monoclonal antibodies (monovalent Fab’ fragment) directed against the glycoprotein (GP)IIb/IIIa complex. We found that 47,600-60,300 molecules of antibody bound per platelet before stimulation, as compared to 89,200-146,500 molecules per platelet after thrombin stimulation. These changes were observed in parallel with a small but significant increase in the dissociation constant (Kd) of two antibodies. In contrast, no statistically significant changes were observed with ADP-stimulated platelets. The increased binding of LJ-CP3, but not of the other two antibodies, to activated platelets decreased by 3040% in the presence of EDTA at 22-25°C, suggesting the occurrence of divalent-cation mediated, activation-dependent changes in the corresponding GPIIb/IIIa epitope. Platelets stimulated by thrombin bound more fibrinogen than those stimulated by ADP, and significant differences in the extent but not in the affinity of fibrinogen binding were observed with different platelet agonists. When the pool of GPIIb/IIIa molecules exposed on the surface of unstimulated platelets was reacted with monoclonal antibody LJ-CP3 to block ADP-induced fibrinogen binding and platelet aggregation, thrombin stimulation still induced substantial binding and aggregation. This effect of thrombin required exposure of platelets to the active agonist and was not mediated by molecules released by thrombin into the medium. Therefore, platelets activated with “strong” agonists exhibit increased number of surface-oriented epitopes associated with GPIIb/IIIa. The GPIIb/IIIa molecules bearing these newly exposed epitopes are functional in that they bind fibrinogen and mediate platelet aggregation.


Blood ◽  
1987 ◽  
Vol 70 (2) ◽  
pp. 475-483 ◽  
Author(s):  
K Niiya ◽  
E Hodson ◽  
R Bader ◽  
V Byers-Ward ◽  
JA Koziol ◽  
...  

Abstract Platelet activation altered the binding of three monoclonal antibodies (monovalent Fab' fragment) directed against the glycoprotein (GP) IIb/IIIa complex. An increased binding of two- to threefold occurred after stimulation with thrombin or phorbol myristate acetate (PMA), with slight but significant increase in the dissociation constants (Kd) of two antibodies (LJ-CP8 and LJ-P9). In contrast, no statistically significant changes were observed with ADP-stimulated platelets. The increased binding of LJ-CP3, but not of the other two antibodies, to activated platelets decreased by 30% to 40% in the presence of EDTA at 22 to 25 degrees C. Platelets stimulated by thrombin or PMA bound more fibrinogen than did those stimulated by ADP, and significant differences in the extent but not in the affinity of fibrinogen binding were observed with various platelet agonists. When the pool of GP IIb/IIIa molecules exposed on the surface of unstimulated platelets was reacted with the monoclonal antibody LJ-CP3 to block ADP-induced fibrinogen binding and platelet aggregation, stimulation with thrombin or PMA still induced substantial binding of antibody and fibrinogen, and aggregation ensued. Therefore, platelets exposed to “strong” agonists exhibit an increased number of surface-oriented epitopes associated with GP IIb/IIIa. The GP IIb/IIIa molecules bearing these newly exposed epitopes are functional in that they can bind fibrinogen and mediate platelet aggregation.


1979 ◽  
Author(s):  
C. Busch ◽  
C. Ljungman ◽  
L. Birgersson

The monoaminooxidase inhibitor tranylcypromine has been suggested to be a selective inhibitor of prostacyclin synthetase with no inhibitory effect on thromboxane A2 formation in platelets. The substance would then be a suitable tool for discrimination between prostacyclin synthesis and release on one hand and other possible thrombocytophobic properties of the endothelial cell particularly its surface on the other. This study, however, shows that tranylcypromine interferes with platelet aggregation induced by thrombin, ADP, adrenaline and collagen whereas that of arachidonic acid is not affected. The results indicate an inhibition at an earlier common pathway of platelet aggregation than the metabolism of arachidonic acid. It is also suggested that the search for a selective inhibitor of prostacyclin synthesis which does not interfere with platelet functions should continue.


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