Platelets and Haemostasis

Platelets And Haemostasis

Leaks in the cardiovascular system can lead to loss of blood and must be rapidly plugged. This is the purpose of haemostasis, a complex process that includes formation of the blood clot, a tough mesh of fibrin entrapping platelets and blood cells. Its complexity is in part due to the precarious balance that must be maintained between providing a rapid and effective means of stopping leaks, and inappropriate formation of clots in blood vessels (thrombosis). Thrombosis is associated with many serious conditions such as coronary artery disease. Platelets play a critical role in haemostasis. They circulate in the blood but are not true cells, being small (∼3 μm) vesicle-like structures formed from megakaryocytes in the bone marrow. They have a lifespan of ∼4 days. Platelets have multiple surface receptors and clearly visible dense granules. These contain mediators such as serotonin and adenosine diphosphate (ADP), which are released on activation.

Primary haemostasis

The immediate response to damage of a blood vessel wall is vasocon- striction, which reduces blood flow and thus loss; it is an intrinsic property of the blood vessel. This is followed by a sequence of events that eventually leads to sealing of the wound by a clot (Fig. 9a). Damage to the vessel wall exposes collagen, to which a plasma protein called von Willebrand factor (vWF) binds. Tissue factor (TF) is also exposed (see below). Platelets have glycoprotein (GP) recep- tors which avidly bind to vWF, tethering the platelet. Further receptors including integrins (Chapter 3) bind directly to collagen. Together these cause adhesion of the platelet to the damaged area. Binding to these receptors also initiates platelet activation, partly by increasing intracellular Ca2+. Platelets change shape, put out pseudopodia and make thromboxane A2 (TXA2) via cyclooxygenase (COX). TXA2 stimulates release of serotonin, ADP and other compounds from the platelet granules. TXA2 and serotonin also enhance the vasoconstric- tion. The process propagates because ADP directly activates more platelets via purinergic (P2Y) receptors. It also causes activation of fibrinogen (GPIIb/IIIa) receptors on their surface, which bind to fibrinogen in the plasma causing the platelets to become sticky and aggregate, forming a soft platelet plug (Fig. 9a). This is stabilized during clotting by conversion of the fibrinogen to fibrin. Note that thrombin (see below) is also a potent platelet activator.

Formation of the blood clot (coagulation) The process leading to formation of the blood clot involves sequential conversion of proenzymes to active enzymes (clotting factors; e.g. factor X → Xa) in an amplifying cascade. Most clotting factors are produced in the liver, which requires vitamin K, and many (e.g. thrombin, factor X) require Ca2+ to act. The ultimate purpose is to produce a massive burst of thrombin (factor IIa; a protease which cleaves fibrinogen to fibrin), and thus rapid formation of the clot. The cell-based model of clotting (Fig. 9b) has replaced the older extrinsic and intrinsic pathways. Most of the action in this model only occurs on cell or platelet surfaces (hence its name).

The initial phase of clotting occurs when cells that express a protein called tissue factor (TF; thromboplastin) become exposed to plasma as a result of vascular damage. Such cells include fibroblasts, monocytes and damaged endothelial cells. Factor VIIa from the plasma is then able to bind to TF (TF : VIIa), and this complex consequently activates a key protein in the clotting process, factor X. It is this, when combined with cofactor Va to form prothrombinase, that converts prothrombin to thrombin; importantly, it can only do so when tethered to the cell by phospholipids. However, insufficient thrombin is produced at this stage for clot formation, and the signal has to be amplified.

The amplification phase takes place on the surface of platelets (Fig. 9b). The small amount of thrombin produced above activates nearby platelets, and also cofactor V on their surface. Cofactor VIII is normally bound to plasma vWF, which protects it from degradation. Thrombin cleaves factor VIII from vWF and activates it, when it also binds to the platelet surface. The end product is a large number of activated platelets, each with a large surface area (due to pseudopodia) covered with the active cofactors, all stuck together by fibrinogen (see above).

The scene is now set for the propagation phase. Thrombin acti- vates a short cascade that leads to activation of factor IX (also acti- vated by TF:VIIa). Factor IXa forms a complex with cofactor VIIIa on the platelet surface to form tenase, a much more powerful activator of factor X than TF:VIIa. The large amount of factor Xa thus generated binds to cofactor Va also on the platelet surface to form a similarly large amount of prothrombinase. There is consequently a massive burst of thrombin production, 1000-fold greater than in the initial phase. Thrombin cleaves the fibrinogen bound around the platelets to form fibrin monomers, which spontaneously polymerize to a fibrous mesh of fibrin, entrapping the platelets and other blood cells. The fibrin polymer is finally cross-linked by factor XIIIa (also activated by thrombin) to create a tough network of fibrin fibres and a stable clot. Retraction of entrapped platelets contracts the clot by ∼60%, making it tougher and assisting repair by drawing the edges of the wound together.

Inhibitors  of  haemostasis  and  fibrinolysis Because thrombin both activates and is produced by the mechanisms described above, there is an element of positive feedback, and the whole process is intrinsically unstable. Multiple inhibitory mechanisms counteract this to prevent inappropriate clotting. Undamaged endothelium produces prostacyclin and nitric oxide (Chapter 24), which impede platelet adhesion and activation and so limit them to damaged areas. Plasma antithrombin inhibits thrombin, factor Xa and tenase, and is strongly potentiated by heparin and heparans on endothelial cells. Thrombomodulin (also on endothelial cells) binds thrombin and converts it so it no longer cleaves fibrinogen but instead activates protein C (APC), which with protein S inactivates factors Va and VIIIa, and hence tenase and prothrombinase. Finally, the clot is broken down by plasmin, a process called fibrinolysis. This occurs when plasma plasminogen binds to fibrin, and is converted to plasmin by tissue plasminogen activator (tPA). Plasmin is itself inactivated by α2-antiplasmin.