Coagulation factors bound to procoagulant platelets concentrate in cap structures to promote clotting

Abstract

AbstractBinding of coagulation factors to phosphatidylserine (PS)-exposing procoagulant-activated platelets followed by formation of the membrane-dependent enzyme complexes is critical for blood coagulation. Procoagulant platelets formed upon strong platelet stimulation, usually with thrombin plus collagen, are large “balloons” with a small (∼1 μm radius) “cap”-like convex region that is enriched with adhesive proteins. Spatial distribution of blood coagulation factors on the surface of procoagulant platelets was investigated using confocal microscopy. All of them, including factors IXa (FIXa), FXa/FX, FVa, FVIII, prothrombin, and PS-sensitive marker Annexin V were distributed nonhomogeneously: they were primarily localized in the “cap,” where their mean concentration was by at least an order of magnitude, higher than on the “balloon.” Assembly of intrinsic tenase on liposomes with various PS densities while keeping the PS content constant demonstrated that such enrichment can accelerate this reaction by 2 orders of magnitude. The mechanisms of such acceleration were investigated using a 3-dimensional computer simulation model of intrinsic tenase based on these data. Transmission electron microscopy and focal ion beam-scanning electron microscopy with Annexin V immunogold-labeling revealed a complex organization of the “caps.” In platelet thrombi formed in whole blood on collagen under arterial shear conditions, ubiquitous “caps” with increased Annexin V, FX, and FXa binding were observed, indicating relevance of this mechanism for surface-attached platelets under physiological flow. These results reveal an essential heterogeneity in the surface distribution of major coagulation factors on the surface of procoagulant platelets and suggest its importance in promoting membrane-dependent coagulation reactions.