The Effect of the Phospholipase Inhibitor Mepacrine on Platelet Aggregation, the Platelet Release Reaction and Fibrinogen Binding to the Platelet Surface

Summary

We have examined whether inhibition by mepacrine of freeing of arachidonic acid from platelet phospholipids inhibits platelet aggregation to collagen, thrombin or ADP, and the release reaction induced by thrombin or collagen. Loss of arachidonic acid was monitored by measuring the amount of ¹⁴ C freed from platelets prelabelled with ¹⁴ C-arachidonic acid. Mepacrine inhibited ¹⁴ C loss by more than 80% but did not inhibit thrombin-induced platelet aggregation and had a small effect on release. ADP-induced platelet aggregation did not cause ¹⁴ C loss. Mepacrine inhibited ADP-induced platelet aggregation by inhibiting the association of fibrinogen with platelets during aggregation. The effect of mepacrine on fibrinogen binding could be considerably decreased by washing the platelets but the inhibition of ¹⁴ C loss persisted. Platelets pretreated with mepacrine and then washed show restoration of aggregation to collagen. Thus, mepacrine has two effects; 1. it inhibits phospholipases, 2. it inhibits fibrinogen binding. Freeing of arachidonic acid is not necessary for platelet aggregation or the release reaction.

Keywords

Platelets - Phospholipases - Mepacrine - Fibrinogen binding

• References

• 1 Charo IF, Feinman RR, Detwiler TT. Interrelations of platelet aggregation and secretion. J Clin Invest 1977; 60: 866-873
• 2 Shuman MA, Botney M, Fenton JJ. Thrombin-induced platelet secretion: Further evidence for a specific pathway. J Clin Invest 1979; 63: 1211-1218
• 3 Packham MA, Guccione MM, Greenberg JJ, Kinlough-Rathbone RL, Mustard JJ. Release of¹⁴ C-serotonin during initial platelet changes induced by thrombin, collagen, or A23187. Blood 1977; 50: 915-926
• 4 Kinlough-Rathbone RL, Packham MM, Reimers H-J, Cazenave J-P, Mustard JJ. Mechanisms of platelet shape change, aggregation and release induced by collagen, thrombin or A23187. J Lab Clin Med 1977; 90: 707-719
• 5 Bills TK, Smith JJ, Silver MM. Metabolism of [¹⁴ C]-arachidonic acid by human platelets. Biochim Biophys Acta 1976; 424: 303-314
• 6 Blackwell GJ, Duncombe WW, Flower RR, Parsons MM, Vane JJ. The distribution and metabolism of arachidonic acid in rabbit platelets during aggregation and its modification by drugs. Br J Pharm 1977; 59: 353-366
• 7 Rittenhouse-Simmons S. Production of diglyceride from phosphatidylinositol in activated human platelets. J Clin Invest 1979; 63: 580-587
• 8 Bell RL, Kennerly DD, Stanford N, Majerus PP. Diglyceride lipase: a pathway for arachidonate release from human platelets. Proc Natl Acad Sci (USA) 1979; 76: 3238-3241
• 9 Malmsten C, Hamberg M, Svensson J, Samuelsson B. Physiological role of an endoperoxide in human platelets: Hemostatic defect due to platelet cyclo-oxygenase deficiency. Proc. Natl Acad Sci (USA) 1975; 72: 1446-1450
• 10 Hamberg M, Svensson J, Samuelsson B. Thromboxanes: A new group of biologically active compounds derived from prostaglandin endoperoxides. Proc Natl Acad Sci (USA) 1975; 72: 2994-2998
• 11 Lapetina AG, Chandrabose KK, Cuatrecasas P. Ionophore A23187 and thrombin-induced platelet aggregation: Independence from cyclo-oxygenase products. Proc Natl Acad Sci (USA) 1978; 75: 818-822
• 12 Vargaftig BB, Fougue F, Chignard M. Interference of bromophenacyl bromide with platelet phospholipase A₂activity induced by thrombin and by the ionophore A23187. Thromb Res 1980; 17: 91-102
• 13 Vargaftig BB, Dao HaiHai. Selective inhibition by mepacrine of the release of ‘rabbit aorta contracting substance’ evoked by the administration of bradykinin. J Pharm Pharmac 1972; 24: 159-161
• 14 Flower RJ, Blackwell GG. The importance of phospholipase-A₂in prostaglandin biosynthesis. Biochem Pharmac 1976; 25: 285-291
• 15 Mustard JF, Perry DD, Kinlough-Rathbone RL, Packham MM. Factors responsible for ADP-induced release reaction of human platelets. Am J Physiol 1975; 228: 1757-1765
• 16 Packham MA, Warrior EE, Glynn MM, Senyi AA, Mustard JJ. Alteration of the response of platelets to surface stimuli by pyrazole compounds. J Exp Med 1967; 126: 171-188
• 17 Mustard JF, Packham MM, Kinlough-Rathbone RL, Perry DD, Regoeczi E. Fibrinogen and ADP-induced platelet aggregation. Blood 1978; 52: 453-466
• 18 Ardlie NG, Perry DD, Packham MM, Mustard JJ. Influence of apyrase on stability of suspensions of washed rabbit platelets. Proc Soc Exp Biol Med 1971; 136: 1021-1023
• 19 Mustard JF, Perry DD, Ardlie NN, Packham MM. Preparation of suspensions of washed platelets from humans. Br J Haemat 1972; 22: 193-204
• 20 Greenberg J, Packham MM, Cazenave J-P, Reimers H-J, Mustard JJ. Effects on platelet function of removal of platelet sialic acid by neuraminidase. Lab Invest 1975; 32: 476-484
• 21 Molnar J, Lorand L. Studies on apyrases. Arch Biochem Biophys 1961; 93: 353-363
• 22 Greenberg JP, Packham MM, Guccione MM, Harfenist EE, Orr JJ, Kinlough-Rathbone RL, Perry DD, Mustard JJ. The effect of pretreatment of human or rabbit platelets with chymotrypsin on their responses to human fibrinogen and aggregating agents. Blood 1979; 54: 753-765
• 23 Mustard JF, Packham MM. Platelet aggregation and the platelet release reaction in thromboembolism. Can Med Assoc J 1970; 103: 859-863
• 24 Kinlough-Rathbone RL, Cazenave J-P, Packham MM, Mustard JJ. Effect of inhibitors of the arachidonate pathway on the release of granule contents from rabbit platelets adherent to collagen. Lab Invest 1980; 42: 28-34