THROMBIN-INDUCED THROMBOXANE SYNTHESIS BY HUMAN PLATELETS - PROPERTIES OF AN ANION-BINDING EXOSITE I-INDEPENDENT RECEPTOR

These studies have examined the effects of thrombin-related agonists i n stimulating thromboxane production by human platelets. The results p resented show that (1) the maximal response elicited by thrombin recep tor agonist peptide (TRAP) stimulation was 40% to 50% of that seen wit h thrombin or the thrombin mutant Thrombin Quick I; (2) pretreatment o f platelets with prostaglandin E-1 or genistein resulted in differenti al inhibition of thromboxane production in response to TRAP compared w ith either enzyme agonist; (3) an antibody to the thrombin receptor cl eavage site that inhibits increases in intracellular [Ca2+] only parti ally reduced thromboxane production in response to 5 nmol/L thrombin a nd 15 nmol/L Thrombin Quick I; (4) preincubation with 20 mu mol/L TRAP resulted in desensitization to further stimulation by 100 mu mol/L TR AP, but not by 100 nmol/L thrombin; and (5) the response to thrombin a fter TRAP desensitization was completely inhibited by the tyrosine kin ase inhibitor genistein and was independent of an intracellular [Ca2+] flux. The cumulative results may be explained by the existence of two proteolytically activated receptors that result in thromboxane produc tion in response to thrombin. One is the thrombin receptor/substrate, PAR-1. Stimulation through the second receptor/substrate depends on a genistein-sensitive step, is independent of an intracellular Ca2- flux , and is initiated by a thrombin-activated receptor that does not depe nd on interaction with anion-binding exosite I, as previously indicate d by the relative activity of Thrombin Quick I in stimulating platelet aggregation and thromboxane production. The proposed second thrombin receptor on platelets represents an additional member of the class of proteolytically activated receptors.