Properties of Procoagulant Platelets

Abstract

Objective To define and characterize the subpopulation of platelets capable of regulating the functional interactions of factors Va (FVa) and Xa (FXa) on the thrombin-activated platelet surface. Methods and Results Flow cytometric analyses were used to define and characterize platelet subpopulations. At a concentration of thrombin known to elicit maximal platelet activation, platelet-derived FVa release, and prothrombinase assembly/function, only a subpopulation of platelets were positive for FVa and FXa binding. An additional subpopulation bound lower levels of FVa, but little, if any, FXa. Fluorescence microscopy analyses confirmed these data. Phenotypically, platelets capable of binding FXa were more highly reticulated and demonstrated significantly increased expression of several key adhesion molecules, including P-selectin, GPIbα, and integrins αIIb and β3. This platelet subpopulation was also defined by the expression of a non-dissociable, membrane-bound pool of functional platelet-derived FVa which comprised ~35–50% of the total membrane-bound cofactor. Conclusions The ability of activated platelets to support thrombin generation is defined by a subpopulation of platelets expressing a non-dissociable pool of platelet-derived FVa and increased adhesive receptor density. This subpopulation is hypothesized to play a significant role in regulating both normal hemostasis and pathological thrombus formation since the adherent properties of platelets and their ability to mount and sustain a procoagulant response are crucial steps in both of these processes.

Superactivated Platelets

“…11 In vitro flow experiments identified superactivated platelets after adhesion to collagen matrices as well, suggesting that collagen alone, perhaps in combination with shear stress, may be sufficient to produce this platelet phenotype. 7 Similarly, Fager et al 12 showed that high concentration of thrombin is sufficient to generate superactivated platelets in comparable proportions and with comparable procoagulant activity to platelets activated by thrombin and convulxin. Stimulation with thrombin coupled with an activator of the platelet Fc Receptor, FcγRIIA, similarly generates a population of superactivated platelets (although it was also noted that this agonist combination consistently produced a smaller proportion of superactivated platelets than stimulation with collagen and thrombin).…”

“…However, there may be more than higher levels of PS exposure to account for enhanced binding of FVa to the membrane surface. It was noted by Fager et al 12 that when superactivated platelets were washed, a significant amount of platelet-derived FVa could not be removed from the platelet surface, leading the authors to conclude that FVa was tethered to the membrane surface by the heavy chain of the cofactor that was bound to the surface, potentially through a glycosylphophtidylinositol anchor. An alternative hypothesis was proposed by Dale et al, 26 which was based on the observation that not only was FVa bound on the platelets' surface but there was retention of other α-granule contents as well, including von Willebrand factor, fibrinogen, and thrombospondin, and therefore described the superactivated platelets as coated.…”

“…28 Several investigators have noted that young (reticulated) platelets have a higher probability of becoming a superactivated platelet and suggest a role of platelet aging in the capacity for superactivation; however, the mechanism linking platelet aging and capacity to become superactivated has not been determined. 8,12 Although the characteristics of superactivated platelets have been described both in vitro and in vivo, individual differences in SPP in both healthy and disease states lack robust clinical data. Some associations between cerebrovascular disease risk have been made, including hemorrhagic transformation and spontaneous intracranial hemorrhage in patients with low SPP.…”

Antiplatelet agents: Is twice a day a better strategy?

“…The population of circulating platelets is not homogeneous [33], immature platelets comprise the youngest component of the circulating platelet pool, and appear to participate most actively in the thrombotic process [34]. They contain measurable amounts of cytosolic messenger ribonucleic acid (mRNA) that is translationally active [35].…”

Analysis of procoagulant phosphatidylserine‐exposing platelets by imaging flow cytometry

“…Flow cytometric analysis of PS‐exposing platelets in suspension using fluorescently‐labeled annexin A5, first described over two decades ago,20 has become widely used in studies of procoagulant platelets and platelet‐derived extracellular vesicles (EVs) 4, 21, 22, 23, 24, 25. A limitation of traditional flow cytometry is the lack of images of individual analyzed cells.…”