Platelet Plug Formation

PLATELET PLUG FORMATION

Platelets (thrombocytes) are small, anucleate fragments of megakaryocytes that adhere to exposed collagen of damaged endothelium, get activated, and aggregate to form a platelet plug (Figure 34.20). Formation of the platelet plug is referred to as primary hemostasis. In a normal adult there are 150,000–450,000 platelets per μl of blood. They have a lifespan of up to 10 days, after which they are taken up by the liver and spleen and destroyed. Clinical laboratory tests to measure platelet number and activity are available.

Figure 34.20 Size comparison of platelets, erythrocytes, and a leukocyte.

A. Adhesion

Adhesion of platelets to exposed collagen at the site of vessel injury is mediated by the protein von Willebrand factor (VWF). VWF binds to collagen, and platelets bind to VWF via glycoprotein Ib (GPIb), a component of a membrane receptor complex (GPIb– V–IX) on the platelet surface (Figure 34.21). Binding to VWF stops the forward movement of platelets. [Note: Deficiency in the receptor for VWF results in Bernard-Soulier syndrome, a disorder of decreased platelet function and number.] VWF is a glycoprotein that is released from platelets. It also is made and secreted by endothelial cells. In addition to mediating the binding of platelets to collagen, VWF also binds to and stabilizes FVIII in the blood. Deficiency of VWF results in von Willebrand disease (VWD), the most common inherited defect in the ability to clot (coagulopathy). VWD coagulopathy results from decreased binding of platelets to collagen and a deficiency in FVIII (due to increased degradation). Platelets can also bind directly to collagen via the membrane receptor GPVI. Once adhered, platelets get activated. [Note: Damage to the endothelium also exposes FIII, initiating the extrinsic pathway of blood clotting and activation of FX (see Figure 34.8).]

Figure 34.21 Binding of platelets via the receptor glycoprotein Ib (GPIb) to von Willebrand factor (VWF). VWF is bound to the exposed collagen at a site of injury.

Figure 34.8 The extrinsic or tissue factor (TF) pathway. Binding of FVII to exposed TF (FIII) activates FVII. [Note: The pathway is quickly inhibited by tissue factor pathway inhibitor (TFPI).] F = factor; Gla = γ-carboxyglutamate; PL = phospholipid; a = active.

B. Activation

Once adhered to areas of injury, platelets get activated. Platelet activation involves morphologic (shape) changes and degranulation, the process by which platelets secrete the contents of their α and δ (or dense) storage granules. Activated platelets also expose PS on their surface. Thrombin is the most potent platelet activator. Thrombin binds to and activates protease-activated receptors, a type of G protein– coupled receptor (GPCR), on the surface of platelets (Figure 34.22) . Thrombin is primarily associated with Gq proteins, resulting in activation of phospholipase C and a rise in diacylglycerol (DAG) and inositol trisphosphate (IP3). [Note: Thrombomodulin, through its binding of thrombin, decreases the availability of thrombin for platelet activation (see Figure 34.18).]

Figure 34.22 Platelet activation by thrombin. [Note: Protease-activated receptors are a type of G protein-coupled receptor.] PIP2 = phosphoinositol bisphosphate; DAG = diacylglycerol; IP3 = inositol trisphosphate;TXA2 = thromboxane A2; ADP = adenosine diphosphate; PDGF = platelet-derived growth factor; VWF = von Willebrand factor; F = factor.

Figure 34.18 Formation and action of the APC complex. Gla = γ-carboxyglutamate; a = active; F = factor.

1. Degranulation: DAG activates protein kinase C, a key event for degranulation. IP₃ causes the release of Ca²⁺ (from dense granules). The Ca²⁺ activates phospholipase A₂, which cleaves membrane phospholipids to release arachidonic acid, the substrate for the synthesis of thromboxane A₂ (TXA₂) in activated platelets by cyclooxygenase-1 (COX-1). TXA₂ causes vasoconstriction, augments degranulation, and binds to platelet GPCRs, causing activation of additional platelets. Recall that aspirin irreversibly inhibits COX and, consequently, TXA₂ synthesis and is referred to as an “antiplatelet” drug. Degranulation also results in release of serotonin and adenosine diphosphate (ADP) from dense granules. Serotonin causes vasoconstriction. ADP binds to GPCRs on the surface of platelets, activating additional platelets. [Note: Some antiplatelet drugs, such as clopidogrel, are ADP-receptor antagonists.] Platelet-derived growth factor (involved in wound healing), VWF, FV, FXIII, and fibrinogen are among other proteins released from α granules. [Note: Platelet-activating factor (PAF), an ether phospholipid synthesized by a variety of cell types including endothelial cells and platelets, binds PAF receptors (GPCRs) on the surface of platelets and activates them.]

2. Shape change: The change in shape of activated platelets from discoidal to spherical with pseudopod-like processes that facilitate platelet–platelet and platelet–surface interactions (Figure 34.23) is initiated by the release of Ca²⁺ from dense granules. Ca²⁺ bound to calmodulin mediates the activation of myosin light-chain kinase that phosphorylates the myosin light chain, resulting in a major reorganization of the platelet cytoskeleton.

Figure 34.23 Activated platelets undergo Ca²⁺-initiated shape change.

C. Aggregation

Activation causes dramatic changes in platelets that lead to their aggregation. Structural changes in a surface receptor (GPIIb/IIIa) expose binding sites for fibrinogen. Bound fibrinogen molecules link activated platelets to one another (Figure 34.24), with a single fibrinogen able to bind two platelets. The fibrinogen is converted to fibrin by thrombin and then covalently cross-linked by FXIIIa coming from both the blood and the platelets. [Note: The exposure of PS on the surface of activated platelets allows formation of the Xase complex (VIIIa, IXa, X, and Ca²⁺) with subsequent formation of FXa and generation of thrombin.] Fibrin strengthens the platelet plug. [Note: Rare defects in the platelet receptor for fibrinogen result in Glanzmann thrombasthenia (decreased platelet function), whereas autoantibodies to this receptor are a cause of immune thrombocytopenia (decreased platelet number).]

Unnecessary activation of platelets is prevented because 1) an intact vascular wall is separated from the blood by a monolayer of endothelial cells, preventing the contact of platelets with collagen; 2) endothelial cells synthesize prostaglandin I₂ (PGI₂, or prostacyclin) and nitric oxide, each of which causes vasodilation; and 3) endothelial cells have a cell-surface ADPase that converts ADP to AMP.

Figure 34.24 Linking of platelets by fibrinogen via the receptor glycoprotein (GP) IIb/IIIa. [Note: The shapes in the fibrinogen molecule represent the two D and one E domains.] GPIb = glycoprotein Ib receptor; VWF = von Willebrand factor.