Cleavage of a 100 kDa membrane protein (aggregin) during thrombin-induced platelet aggregation is mediated by the high affinity thrombin receptors.
Thrombin-induced platelet aggregation is accompanied by cleavage of aggregin, a surface membrane protein (Mr = 100 kDa), and is mediated by the intracellular activation of calpain. We now find that agents that increase intracellular levels of platelet cAMP by stimulating adenylate cyclase, also inhibit thrombin binding and platelet activation by destabilizing thrombin receptors on the platelet surface. Iloprost (a stable analog of PGI2) and forskolin each completely inhibited platelet aggregation by 2 nM thrombin and markedly decreased cleavage of aggregin. Thrombin inactivated by D-phenylalanine-L-prolyl-L-arginine chloromethyl ketone (PPACK-thrombin) binds to the highest affinity site for thrombin on the platelet surface, but thrombin modified by N alpha-tosyl-L-lysine chloromethylketone (TLCK-thrombin) does not. We now demonstrate that preincubation of platelets with PPACK-thrombin blocked platelet aggregation and cleavage of aggregin induced by 2 nM thrombin. In contrast, TLCK-thrombin neither blocked platelet aggregation nor the cleavage of aggregin. These results show that a) platelet aggregation and cleavage of aggregin by thrombin (2nm) involves the occupancy of high affinity alpha-thrombin receptors on the platelet surface, and b) stimulators of adenylate cyclase which increase cAMP, inhibit thrombin-induced platelet aggregation and cleavage of aggregin by mechanisms which include inhibiting the binding of thrombin to its receptors.
Puri
R N
RN
Thrombosis Research Center, Temple University Health Sciences Center, Philadelphia, PA 19140.
Zhou
F X
FX
Colman
R F
RF
Colman
R W
RW
D002154:Calpain / Q000502:physiology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology; D011464:Epoprostenol / Q000494:pharmacology; D016285:Iloprost; D008565:Membrane Proteins / Q000378:metabolism*; D010974:Platelet Aggregation; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000502:physiology*; D018179:Receptors, Thrombin; D013917:Thrombin / Q000037:antagonists & inhibitors / Q000494:pharmacology*
D008565:Membrane Proteins; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D018179:Receptors, Thrombin; C055422:aggregin; D005576:Colforsin; D011464:Epoprostenol; D000242:Cyclic AMP; D013917:Thrombin; D002154:Calpain; D016285:Iloprost