Principles of Hemostasis, Part II

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The primary principle of hemostasis is to minimize blood loss at sites of vessel injury by forming a thrombus (clot) and at the same time maintaining blood flow. To achieve this, there is a highly regulated, fine-tuned interaction of multiple processes, involving the blood vessel wall (principally the endothelium), platelets and non-cellular blood constituents.

The core elements of hemostasis include blood vessel constriction, platelet activation, coagulation and fibrinolysis. All of these processes are initiated at the same time in response to blood vessel injury. Damage to the endothelial cell lining of the blood vessel results in exposure of collagen which, under normal circumstances, is not in contact with the blood. When circulating platelets come into contact with collagen they are activated and start to stick to the damaged surface. At the same time, a substance called tissue factor is also exposed which activates the coagulation cascade resulting in the formation of a fibrin clot. Platelet adhesion and activation is conventionally known as primary hemostasis and the process of coagulation and fibrin formation is termed secondary hemostasis.

Normal hemostasis depends on the interaction of the following:

1. blood vessel
2. platelets
3. coagulation system
4. fibrinolytic system

A defect in one or more of these systems will result in either a bleeding disorder or a tendency to clot. Despite the complexities of the control of hemostasis, ultimately the outcome is, quite simply, a matter of balance between clot formation and clot breakdown, both of which in turn are influenced by blood flow (or stasis), the vessel wall and the constituents of the blood (Fig. 1, pdf).

This article discusses the physiology of coagulation and fibrinolysis. To learn about the interaction of blood vessels and platelets in hemostasis, read “Principles of Hemostasis, Part I”.

Coagulation

Coagulation is the process of converting soluble fibrinogen into insoluble fibrin in order to form a stable blood clot around the platelets that were initially trapped at the site of injury. The plasma contains a large number of proteins called coagulation factors that participate in a sequence of enzymatic reactions that result in the formation of large amounts of insoluble fibrin. This sequential reaction is called the coagulation cascade. The clotting factors involved are factors XI, X, IX, VIII, VII, V and II. Factor II, also called prothrombin, becomes thrombin when activated.

For normal hemostasis to occur, a sufficient amount of thrombin needs to be rapidly generated in order for fibrinogen to be converted to fibrin. Fibrin is relatively ‘watertight’ and therefore further blood loss is prevented. Fibrin is also relatively resistant to fibrinolytic breakdown (see “Fibrinolysis”). When the surface of the endothelium is damaged, coagulation is immediately activated by the exposure of tissue factor (TF), which is present below the endothelial cells in the vessel wall.

The coagulation cascade – The coagulation cascade, as we understand it to exist today, is shown in Figure 2 (pdf). The process of coagulation starts with the presence of TF, which binds and activates FVII to FVIIa. The activated clotting factors are enzymes that sequentially cleave each other in a highly regulated fashion. This process happens wherever TF has become exposed. Once the TF:FVIIa complex is formed, it has a potent positive feedback loop on itself thereby increasing the generation of more TF:FVIIa several thousand fold. TF:FVIIa in turn forms a complex with FX and converts it to FXa. It is also able to bind and activate FIX to FIXa, although the intensity of this reaction is far less potent. FXa, together with the co-factor FVa, converts prothrombin to thrombin (FIIa). This initial generation of thrombin is very short-lived as TF:FVIIa is very rapidly inactivated by tissue factor pathway inhibitor (TFPI). Furthermore, thrombin activates FXIII to FXIIIa which in turn is responsible for irreversibly cross-linking fibrin, creating insoluble polymers and a stable clot.

It can be said that the primary role of the initial small quantity of thrombin generated is to activate platelets that are within and in proximity to the growing thrombus. Thrombin produced from the secondary positive feedback activation loop of FXI by thrombin is responsible for the sustained burst of thrombin that generates the stable fibrin clot.

The importance of vitamin K and calcium – An essential element in the success of the coagulation cascade is the assembly of the various proteases in an orderly fashion on the surface of the platelet. The key to this process is the presence of GLA (γ-carboxyglutamic acid) domains of the vitamin K dependent clotting factors which are factors II, VII, IX and X. The GLA domains of these clotting factors are the key to the assembly of these complexes on platelet surfaces as they provide the critical link. Calcium ions, which are positively charged, bind to various GLA domains and to the phospholipid of the platelet, both of which are negatively charged, thereby localizing the clotting factors to the platelet surface. The calcium ions essentially form a bridge between the clotting factor and the phospholipid on the activated platelet surface (Fig. 3 on next page). This is how the clotting process is kept localized at the original site of endothelial injury. An absence of functional vitamin K will result in reduced ability for the coagulation factors to assemble on the platelet surface and compromise clot formation.

Fibrinolysis

In the absence of adequate quantities of thrombin, a stable clot cannot be formed, resulting in bleeding. Conversely, an unregulated production of thrombin will result in pathological thrombosis. A variety of mechanisms that collectively inhibit thrombin production have been well described. These include:

• Removal of activated clotting factors by blood flow past the clot
• Inactivation of clotting factors by circulating natural anticoagulants (protein C, protein S and antithrombin)
• Fibrinolysis

Fibrinolysis is the process of controlled clot breakdown. As soon as clot formation is triggered, so is the fibrinolytic pathway activated. The primary enzyme involved here is plasmin which degrades fibrin into fibrin breakdown products (FDPs) which includes D-Dimers.

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