Platelet-derived factor Va/Va Leiden cofactor activities are sustained on the surface of activated platelets despite the presence of activated protein C.
We investigated the role of the thrombin-activated platelet in modulating the rate and extent of activated protein C (APC)-catalyzed inactivation of platelet-derived factor Va and factor VaLeiden. Platelet-derived factor Va and factor VaLeiden were inactivated by APC at near identical rates; however, complete inactivation of the cofactors was never achieved. Greater residual cofactor activity remained when using thrombin-activated platelets compared with that observed with synthetic phospholipid vesicles and platelet-derived microparticles, suggesting that thrombin-activated platelets protect the cofactors from APC-catalyzed inactivation. This apparent protection was not due to (1) an insufficient number of membrane binding sites for APC or factor Va; (2) the destruction of these sites; or (3) the presence of a platelet-associated APC inhibitor. Results from a plasma-based clotting assay (with or without APC) with platelets or PCPS vesicles added to induce clot formation indicated that, even in the presence of high concentrations of APC, platelets offered protection of the cofactor by delaying cleavage at Arg506. This resulted in incomplete proteolysis of the heavy chain, suggesting that platelets can also protect plasma-derived factor Va from APC-catalyzed inactivation. However, additional experiments indicated that the plasma-derived cofactor, bound to thrombin-activated platelets, was completely inactivated by APC, suggesting that the plasma and platelet-derived cofactor pools represent different substrates for APC. Collectively, these results indicate that platelets sustain procoagulant events by providing a membrane surface that delays cofactor inactivation and by releasing a cofactor molecule that displays an APC resistant phenotype. Thus, at sites of arterial injury, the factor VLeiden mutation may not as readily predict arterial thrombosis, because the normal and variant platelet-derived cofactors are equally resistant to APC at the activated platelet surface.
Camire
R M
RM
Department of Biochemistry, University of Vermont, College of Medicine, Burlington, VT, USA.
Kalafatis
M
M
Simioni
P
P
Girolami
A
A
Tracy
P B
PB
D001792:Blood Platelets / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D005165:Factor V / Q000378:metabolism; D015943:Factor Va / Q000378:metabolism*; D006801:Humans; D015539:Platelet Activation / Q000187:drug effects; D011486:Protein C / Q000378:metabolism*; D013917:Thrombin / Q000494:pharmacology
D011486:Protein C; C095381:factor V Leiden; D015943:Factor Va; D005165:Factor V; D013917:Thrombin