ADP Plays a Key Role in Thrombogenesis in Rats

Summary

The relative importance of ADP, arachidonic acid metabolites and serotonin as thrombogenic factors was evaluated in rats by comparing, after oral administration, the effects of two inhibitors of ADP-induced platelet aggregation (ticlopidine and PCR 4099), three cyclo-oxygenase inhibitors (aspirin, triflusal and indobufen) and a selective serotonin 5HT2 receptor antagonist (ketanserin) on platelet aggregation, in four platelet-dependent thrombosis models and on bleeding time. Platelet aggregation induced by ADP and collagen was completely inhibited by ticlopidine and PCR 4099 whereas only the collagen aggregation was reduced by the cyclo-oxygenase inhibitors. Ketanserin or a depletion of platelet serotonin by reserpine did not affect platelet aggregation. Ticlopidine and PCR 4099 greatly prolonged rat tail transection bleeding time. This is probably related to their known ability to inhibit ADP-mediated platelet aggregation. In contrast, the cyclooxygenase inhibitors did not affect bleeding time at all. Reserpine and ketanserin prolonged bleeding time by interfering with the action of serotonin on the vascular wall. Ticlopidine and PCR4099 were very potent antithrombotics in all the models. Aspirin, only at a high dose, inhibited poorly thrombus formation on a silk thread in an arterio-venous shunt, suggesting that the inhibition of cyclo-oxygenase was not responsible. Triflusal was inactive in all models while indobufen slightly reduced thrombus formation in the silk thread and metallic coil models. Ketanserin and reserpine reduced thrombus only in the metallic coil model. Thrombus formation was greatly reduced in fawn-hooded rats, which lack ADP in their platelet dense granules because of a genetic storage pool deficiency. Taken together, the results obtained with the drugs and with the fawn-hooded rats support the concept that ADP plays a key role in thrombogenesis in rats.

Keywords

Fawn-hooded rat - Platelet aggregation - Thrombosis - Ticlopidine - PCR 4099 - Aspirin - Triflusal - Indobufen - Ketanserin

• References

• 1 Packham MA, Mustard JF. Normal and abnormal platelet activity. In: Blood platelet function and medicinal chemistry. A Lasslo (ed) pp. 61 Elsevier Biomedical; New York: 1984
• 2 Hwang DH. Aggregation and inhibition of rat platelets, and the formation of endoperoxide metabolities. Prostaglandins and Medicine 1980; 5: 163-173
• 3 Huzoor-Akbar AnwerK. Evidence that the rat is not an appropriate model to study the role of prostaglandins in normal or abnormal platelet aggregation. Thromb Res 1986; 41: 555-566
• 4 Nishizawa EE, Williams DJ, Connell CL. Arachidonate induced aggregation of rat platelets may not require prostaglandin endoperox- ides or thromboxane A2. Thromb Res 1983; 30: 289-296
• 5 Emms H, Lewis GP. The roles of prostaglandin endoperoxides, thromboxane A2 and adenosine diphosphate in collagen-induced aggregation in man and the rat. Br J Pharmac 1986; 87: 109-115
• 6 Buczko W, Gambino MC, de Gaetano G. Prolongation of rat tail bleeding time by ketanserin: mechanisms of action. European J Pharmacol 1984; 103: 261-268
• 7 Inarrea P, Gomez-Cambronero J, Nieto M, Sanchez Crespo M. Characteristics of the binding of platelet-activating factor to platelets of different animal species. European J Pharmacol 1984; 105: 309-115
• 8 Umetsu T, Sanai K. Effect of l-Methyl-2Mercapto-5-(3-Pyridyl)- Imidazole (KC-6141), an anti-aggregating compound, on experimental thrombosis in rats. Thromb Haemostas 1978; 39: 74-83
• 9 Shand RA, Smith JR, Wallis RB. Expression of the platelet procoagulant activity in vivo in thrombus formation in an extracorporeal shunt in the rat. Thromb Res 1984; 36: 223-232
• 10 Kumada T, Ishihara M, Ogawa H, Abiko Y. Experimental model of venous thrombosis in rats and effects of some agents. Thromb Res 1980; 18: 189-203
• 11 Reyers I, Mussoni L, Donati MB, de Gaetano G. Failure of aspirin at different doses to modify experimental thrombosis in rats. Thromb Res 1980; 669-674
• 12 Bernat A, Vallee E, Maffrand JP, Gordon J. The role of platelets in experimental thrombosis induced by venous stasis in the rat. Thromb Res. Accepted for publication
• 13 Reyers I, Mussoni L, Donati MB, de Gaetano G. Severe thrombocytopenia delays but does not prevent the occlusion of an arterial prosthesis in rats. Thromb Haemostas 1981; 46: 558-560
• 14 Reyers I, Hennissen A, Donati MB, Hornstra G, de Gaetano G. Aspirin and the prevention of experimental arterial thrombosis: difficulty in establishing unequivocal effectiveness. Thromb Res 1985; 54: 619-621
• 15 Dejana E, Villa S, de Gaetano G. Bleediung time in rats: a comparison of different experimental conditions. Thromb Haemostas 1982; 48: 108-11
• 16 Panak E, Maffrand JP, Picard-Fraire C, Vallee E, Blanchard J, Roncucci R. Ticlopidine: a promise for the prevention and treatment of thrombosis and its complications. Haemostasis 1983; 13, suppl (Suppl. 01) 1-54
• 17 Thebault J, McEwen J, McCraw A, Bouloux C, Jacob C, Chigot C, Irvine A, Kindermans M, Maffrand JP, Roncucci R. PCR4099 A new antithrombotic drug, evaluation of tolerance and of pharmacological activity. Thromb Res 1986; Suppl 6: 286
• 18 Sancho MJ, de Castellarnau C, Villa L, Garcia J, Carretero F, Fabra A, Rutilant MLI. Effects of salicylate derivatives on cyclo-oxygenase interaction studies. Biol Clin Hematol 1985; 7: 135-143
• 19 Crow MJ, Salter MC P, Donaldson DR, Rajah SM. Inhibition of platelet function with indobufen: correlation with plasma drug level. Thromb Res 1985; 38: 303-306
• 20 De Clerk F, Van Nueten JM, Reneman RS. Platelet-vessel wall interactions: implication of 5-hydroxytryptamine. A review Agents and actions 1984; 15: 612-626
• 21 Tschopp TB, Zucker MB. Hereditary defect in platelet function in rats. Blood 1972; 40: 217-226
• 22 Born GV R, Cross MJ. The aggregation of blood platelets. J Physiol 1963; 168: 1178-1195
• 23 Cazenave JP, Hemmendinger S, Beretz A, Sutter-Bay A, Launay J. L’agrégation plaquettaire: outil d’investigation clinique et d’étude pharmacologique. Methodologie Ann Biol clin 1983; 41: 167-179
• 24 Cerletti C, Gambino MC, Garattini S, de Gaetano G. Biochemical selectivity of oral versus intravenous aspirin in rats. J Clin Invest 1986; 78: 323-326
• 25 Feliste R, Delebassee D, Simon MF, Chap H, Defreyn G, Vallee E, Douste-Blazy L, Maffrand JP. Ticlopidine and released ADP. Thromb Res. Accepted for publication
• 26 Dunn FW, Soria J, Soria C, Thomaidis A, Lee H, Caen JP. In vivo effect of ticlopidine on fibrinogen-platelet cofactor activity and binding of fibrinogen to platelets. Agents and Actions suppl 1984; 15: 97-104
• 27 Di Minno G, Cerbone AM, Mattioli PL, Turco S, Iovine C, Mancini M. Functionally thrombasthenic state in normal platelets following the administration of ticlopidine. J Clin Invest 1985; 75: 328-338
• 28 Boissel JP. Registry of multicenter clinicals trials. report Thromb Haemostas 1986; 55: 282-291
• 29 Reyers-degli InnocentiI, Poggi A, de Gaetano G. Platelet aggregation and haemolysis induced in rats by intravenous infusion of ADP. Effect of potentially antithrombotic drugs Scand J Haemath 1974; 13: 331-337
• 30 Martin JF, Suggett AJ, Jones PB C. Effect of acetylsalicylic acid on platelet aggregation and thromboxane B2 production in flowing aortic blood in the rat studied with a filter loop technique. Thromb Res 1982; 27: 251-259
• 31 Diamantis W, Kohlhepp WC, Haertlein B, Melton J, Sofia RD. Meseclazone, 5-chlorosalicylic acid and acetylsalicylic acid. Comparison of their effects on in vitro and ex vivo platelet aggregation Thromb Haemostas 1978; 40: 24-36
• 32 Legrand C, Nurden AT. Studies on platelets of patients with inherited platelet disorders suggest that collagen-induced fibrinogen binding to membrane receptors requires secreted ADP but not released alpha- granule proteins. Thromb Haemostas 1985; 54: 603-606
• 33 Buchanan MR, Rischke JA, Hirsh J. Aspirin inhibits platelet function independent of the acetylation of cyclo-oxagenase. Thromb Res 1982; 25: 363-373
• 34 Born GV R. Adenosine diphosphate as a mediator of platelet aggregation in vivo: an editorial view. Circulation 1985; 72: 741-746
• 35 Zawilska KM, Born GV R, Begent NA. Effect of ADP-utilizing enzymes on the arterial bleeding time in rats and rabbits. Br J Haematol 1982; 50: 317-325
• 36 George JH, Nurden AT, Phillips DR. Molecular defects in interactions of platelets with the vessel wall. N Engl J Med 1984; 311: 1084-1098
• 37 Hornstra G. Dietary fats, prostanoids and arterial thrombosis. Mar- tinus Nijhoff publishers 1982.
• 38 Begent N, Born GV R. Growth rate in vivo of platelet thrombi, produced by iontophoresis of ADP, as a function of mean blood flow velocity. Nature 1970; 227: 926-930
• 39 Zawilska K, Izrael V, Chelloul N, Caen J. Prevention de la coagulation intravasculaire disseminee au cours du phenomene de Schwartz- man generalise chez le lapin par un systeme enzymatique consommant l’ADP plasmatique. Act Hematol 1975; 9: 269-286
• 40 Lewis Jr HD, Davis JW, Archibald DG, Phil M, Steinke WE, Smitherman TC, Doherty JE, Schnaper HW, Lewinter MM, Linares E, Pouget JM, Sabharwal SC, Chesler E. Demots H. Protective effects of aspirin against acute myocardial infarction and death in men with unstable angina N Engl J Med 1983; 309: 396-403
• 41 Cairns JA, Gent M, Singer J, Finnie KJ, Frogatt GM, Holder DA, Jablonsky G, Kostuk WJ, Melendez LJ, Myers MG, Sackett DL, Searley BJ, Tanser PH. Aspirin, sulfinpyrazone, or both in unstable angina. Results of a Canadian multicenter trial N Engl J Med 1985; 213: 1369-1375
• 42 Chesebro JH, Fuster V, Elveback LR, Clements IP, Smith HC, Holmes DR, Bardsley WT, Pluth JR, Wallace RB, Puga FJ, Orszulak TA, Piehler JM, Danielson GK, Schaff HV, Frye RL. Effect of dipyridamole and aspirin on late vein-graft patency after coronary bypass operations. N Engl J Med 1984; 310: 209-214
• 43 Colman RW. Platelet activation: role of an ADP receptor. Seminars in Hematology 1986; 23: 119-128
• 44 Timmons S, Kloczewiak M, Hawiger J. ADP-dependent common receptor mechanism for binding of von Willebrand factor and fibrinogen to human platelets. Proc Natl Acad Sci USA 1984; 81: 4935-4939
• 45 Weiss HJ, Turitto VT, Baumgartner HR. Platelet adhesion and thrombus formation on subendothelium in platelets deficient in glycoproteins Ilb-IIIa, lb, and Storage Granules. Blood 1986; 67: 322-330
• 46 Escolar G, Bastida E, Castillo R, Ordinas A. Ticlopidine inhibits platelet thrombus formation studied in a flowing system. Thromb Res 1987; 45: 561-571
• 47 Arcan JC, Destors JM. Controlled clinical trial of ticlopidine in patients suffering from intermittent claudication. Thromb Res 1986; suppl VI: 315