Thus, platelet reactivity towards haemostatic, immune or other stimuli acting see more at CLEC-2 (such as the tumour cell-expressed ligand, podoplanin) or FcγRIIa (such as antiplatelet autoantibodies) could directly or indirectly affect the haemostatic response in individuals. As
discussed below, human platelet GPVI is also irreversibly downregulated by ectodomain shedding, and FcγRIIa is mutually regulated by a separate proteolytic inactivation – ligands acting at either receptor also lead to both ADAM10-mediated ectodomain shedding of GPVI and the calpain-mediated intracellular inactivation of FcγRIIa [51]. In primary haemostasis, major downstream consequences of ligand engagement of GPIbα/GPVI by VWF/collagen is the secretion of autocrine-acting dense granule contents, ADP and thromboxane, which induce G-protein receptor-mediated secondary find more platelet activation, and amplify signals leading to activation of the platelet-specific integrin, αIIbβ3 (GPIIb-IIIa) (Fig. 1). Similarly, studies with Nbeal2-deficient mice, which lack normal α-granule protein content, indicate that secretion of the protein constituents of α-granules is critical in regulating platelet adhesion and overall haemostasis and vascular thrombosis
[52]. Fibrinogen or VWF binding to αIIbβ3 cross-links platelets enabling platelet aggregation. The receptor αIIbβ3 is constitutively expressed on the platelet surface, but requires signalling-dependent activation whereby the complex rapidly becomes competent to bind ligand (inside-out signalling); in addition, ligand binding induces outside-in signals
which further control platelet shape change, signalling and contraction of the formed thrombus. Recent studies have shed new light on the details underpinning this functional regulation of αIIbβ3 [53, 54], and importantly, Thymidylate synthase begin to distinguish between the roles of this receptor in haemostasis vs. thrombosis. In this mechanism, the cytoskeletal protein talin, involved in inside-out signals and ligand binding, is displaced by the G protein, Gα13, which mediates outside-in signalling, regulating functions such as platelet spreading. In turn, this is followed by reciprocal displacement of Gα13 by talin which then controls contraction of the thrombus or other functions. Experimentally, inhibition of Gα13 binding selectively blocks outside-in signalling, impacting upon occlusive arterial thrombus formation but not bleeding time as an indicator of haemostasis [54]. A further example of a new mechanism for downregulation of αIIbβ3 involves the platelet protein disulphide isomerase, ERp57, which appears in increased amounts on the surface of activated platelets in a β3-dependent manner, and also attenuates thrombus growth in mice [55].