Question

Describe the components of the clotting cascade which are NOT directly involved in the sequence. What do they do?

Answer 1

Number and/or name	Function
I (fibrinogen)	Forms clot (fibrin)
II (prothrombin)	Its active form (IIa) activates I, V, VII, VIII, XI, XIII, protein C, platelets
III (tissue factor or tissue thromboplastin)	Co-factor of VIIa (formerly known as factor III)
IV (calcium)	Required for coagulation factors to bind to phospholipid (formerly known as factor IV)
V (proaccelerin, labile factor)	Co-factor of X with which it forms the prothrombinase complex
VI	Unassigned – old name of Factor Va
VII (stable factor, proconvertin)	Activates IX, X
VIII (Antihemophilic factor A)	Co-factor of IX with which it forms the tenase complex
IX (Antihemophilic factor B or Christmas factor)	Activates X: forms tenase complex with factor VIII
X (Stuart-Prower factor)	Activates II: forms prothrombinase complex with factor V
XI (plasma thromboplastin antecedent)	Activates IX
XII (Hageman factor)	Activates factor XI, VII, prekallikrein and plasminogen
XIII (fibrin-stabilizing factor)	Crosslinks fibrin
von Willebrand factor	Binds to VIII, mediates platelet adhesion
prekallikrein (Fletcher factor)	Activates XII and prekallikrein; cleaves HMWK
high-molecular-weight kininogen (HMWK) (Fitzgerald factor)	Supports reciprocal activation of XII, XI, and prekallikrein
fibronectin	Mediates cell adhesion
antithrombin III	Inhibits IIa, Xa, and other proteases
heparin cofactor II	Inhibits IIa, cofactor for heparin and dermatan sulfate ("minor antithrombin")
protein C	Inactivates Va and VIIIa
protein S	Cofactor for activated protein C (APC, inactive when bound to C4b-binding protein)
protein Z	Mediates thrombin adhesion to phospholipids and stimulates degradation of factor X by ZPI
Protein Z-related protease inhibitor (ZPI)	Degrades factors X (in presence of protein Z) and XI (independently)
plasminogen	Converts to plasmin, lyses fibrin and other proteins
alpha 2-antiplasmin	Inhibits plasmin
tissue plasminogen activator (tPA)	Activates plasminogen
urokinase	Activates plasminogen
plasminogen activator inhibitor-1 (PAI1)	Inactivates tPA & urokinase (endothelial PAI)

Coagulation, also known as clotting, is the process by which blood changes from a liquid to a gel, forming a blood clot. It potentially results in hemostasis, the cessation of blood loss from a damaged vessel, followed by repair. The mechanism of coagulation involves activation, adhesion and aggregation of platelets, as well as deposition and maturation of fibrin.

Platelet activation

When the endothelium is damaged, the normally isolated, underlying collagen is exposed to circulating platelets, which bind directly to collagen with collagen-specific glycoprotein Ia/IIa surface receptors. This adhesion is strengthened further by von Willebrand factor (vWF), which is released from the endothelium and from platelets; vWF forms additional links between the platelets' glycoprotein Ib/IX/V and A1 domain. This localization of platelets to the extracellular matrix promotes collagen interaction with platelet glycoprotein VI. Binding of collagen to glycoprotein VI triggers a signaling cascade that results in activation of platelet integrins. Activated integrins mediate tight binding of platelets to the extracellular matrix. This process adheres platelets to the site of injury

Activated platelets release the contents of stored granules into the blood plasma. The granules include ADP, serotonin, platelet-activating factor (PAF), vWF, platelet factor 4, and thromboxane A2 (TXA2), which, in turn, activate additional platelets. The granules' contents activate a Gq-linked protein receptor cascade, resulting in increased calcium concentration in the platelets' cytosol. The calcium activates protein kinase C, which, in turn, activates phospholipase A2 (PLA2). PLA2 then modifies the integrin membrane glycoprotein IIb/IIIa, increasing its affinity to bind fibrinogen. The activated platelets change shape from spherical to stellate, and the fibrinogen cross-links with glycoprotein IIb/IIIa aid in aggregation of adjacent platelets (completing primary hemostasis).

The coagulation cascade of secondary hemostasis has two initial pathways which lead to fibrin formation. These are the contact activation pathway (also known as the intrinsic pathway), and the tissue factor pathway (also known as the extrinsic pathway), which both lead to the same fundamental reactions that produce fibrin. It was previously thought that the two pathways of coagulation cascade were of equal importance, but it is now known that the primary pathway for the initiation of blood coagulation is the tissue factor (extrinsic) pathway. The pathways are a series of reactions, in which a zymogen (inactive enzyme precursor) of a serine protease and its glycoprotein co-factor are activated to become active components that then catalyze the next reaction in the cascade, ultimately resulting in cross-linked fibrin. Coagulation factors are generally indicated by Roman numerals, with a lowercase a appended to indicate an active form.

The coagulation factors are generally serine proteases (enzymes), which act by cleaving downstream proteins. The exceptions are tissue factor, FV, FVIII, FXIII. Tissue factor, FV and FVIII are glycoproteins, and Factor XIII is a transglutaminase. The coagulation factors circulate as inactive zymogens. The coagulation cascade is therefore classically divided into three pathways. The tissue factor and contact activation pathways both activate the "final common pathway" of factor X, thrombin and fibrin.

Tissue factor pathway (extrinsic)

The main role of the tissue factor pathway is to generate a "thrombin burst", a process by which thrombin, the most important constituent of the coagulation cascade in terms of its feedback activation roles, is released very rapidly. FVIIa circulates in a higher amount than any other activated coagulation factor. The process includes the following steps:

1. Following damage to the blood vessel, FVII leaves the circulation and comes into contact with tissue factor (TF) expressed on tissue-factor-bearing cells (stromal fibroblasts and leukocytes), forming an activated complex (TF-FVIIa).

2. TF-FVIIa activates FIX and FX.

3. FVII is itself activated by thrombin, FXIa, FXII and FXa.

4. The activation of FX (to form FXa) by TF-FVIIa is almost immediately inhibited by tissue factor pathway inhibitor (TFPI).

5. FXa and its co-factor FVa form the prothrombinase complex, which activates prothrombin to thrombin.

6. Thrombin then activates other components of the coagulation cascade, including FV and FVIII (which forms a complex with FIX), and activates and releases FVIII from being bound to vWF.

7. FVIIIa is the co-factor of FIXa, and together they form the "tenase" complex, which activates FX; and so the cycle continues. ("Tenase" is a contraction of "ten" and the suffix "-ase" used for enzymes.)

Contact activation pathway (intrinsic)

The contact activation pathway begins with formation of the primary complex on collagen by high-molecular-weight kininogen (HMWK), prekallikrein, and FXII (Hageman factor). Prekallikrein is converted to kallikrein and FXII becomes FXIIa. FXIIa converts FXI into FXIa. Factor XIa activates FIX, which with its co-factor FVIIIa form the tenase complex, which activates FX to FXa. The minor role that the contact activation pathway has in initiating clot formation can be illustrated by the fact that patients with severe deficiencies of FXII, HMWK, and prekallikrein do not have a bleeding disorder. Instead, contact activation system seems to be more involved in inflammation, and innate immunity. Despite this, interference with the pathway may confer protection against thrombosis without a significant bleeding risk.

Final common pathway

The division of coagulation in two pathways is arbitrary, originating from laboratory tests in which clotting times were measured either after the clotting was initiated by glass, the intrinsic pathway; or clotting was initiated by thromboplastin (a mix of tissue factor and phospholipids), the extrinsic pathway.

Further, the final common pathway scheme implies that prothrombin is converted to thrombin only when acted upon by the intrinsic or extrinsic pathways, which is an oversimplification. In fact, thrombin is generated by activated platelets at the initiation of the platelet plug, which in turn promotes more platelet activation.

Thrombin functions not only to convert fibrinogen to fibrin, it also activates Factors VIII and V and their inhibitor protein C (in the presence of thrombomodulin); and it activates Factor XIII, which forms covalent bonds that crosslink the fibrin polymers that form from activated monomers.

The coagulation cascade is maintained in a prothrombotic state by the continued activation of FVIII and FIX to form the tenase complex, until it is down-regulated by the anticoagulant pathways.

Cofactors

Various substances are required for the proper functioning of the coagulation cascade:

Calcium and phospholipid

Calcium and phospholipid (a platelet membrane constituent) are required for the tenase and prothrombinase complexes to function. Calcium mediates the binding of the complexes via the terminal gamma-carboxy residues on FXa and FIXa to the phospholipid surfaces expressed by platelets, as well as procoagulant microparticles or microvesicles shed from them. Calcium is also required at other points in the coagulation cascade.

Vitamin K

Vitamin K is an essential factor to a hepatic gamma-glutamyl carboxylase that adds a carboxyl group to glutamic acid residues on factors II, VII, IX and X, as well as Protein S, Protein C and Protein Z. In adding the gamma-carboxyl group to glutamate residues on the immature clotting factors, Vitamin K is itself oxidized. Another enzyme, Vitamin K epoxide reductase (VKORC), reduces vitamin K back to its active form. Vitamin K epoxide reductase is pharmacologically important as a target of anticoagulant drugs warfarin and related coumarins such as acenocoumarol, phenprocoumon, and dicumarol. These drugs create a deficiency of reduced vitamin K by blocking VKORC, thereby inhibiting maturation of clotting factors. Vitamin K deficiency from other causes (e.g., in malabsorption) or impaired vitamin K metabolism in disease (e.g., in liver failure) lead to the formation of PIVKAs (proteins formed in vitamin K absence), which are partially or totally non-gamma carboxylated, affecting the coagulation factors' ability to bind to phospholipid.

Protein C

Protein C is a major physiological anticoagulant. It is a vitamin K-dependent serine protease enzyme that is activated by thrombin into activated protein C (APC). Protein C is activated in a sequence that starts with Protein C and thrombin binding to a cell surface protein thrombomodulin. Thrombomodulin binds these proteins in such a way that it activates Protein C. The activated form, along with protein S and a phospholipid as cofactors, degrades FVa and FVIIIa. Quantitative or qualitative deficiency of either (protein C or protein S) may lead to thrombophilia (a tendency to develop thrombosis). Impaired action of Protein C (activated Protein C resistance), for example by having the "Leiden" variant of Factor V or high levels of FVIII, also may lead to a thrombotic tendency.

Antithrombin

Antithrombin is a serine protease inhibitor (serpin) that degrades the serine proteases: thrombin, FIXa, FXa, FXIa, and FXIIa. It is constantly active, but its adhesion to these factors is increased by the presence of heparan sulfate (a glycosaminoglycan) or the administration of heparins (different heparinoids increase affinity to FXa, thrombin, or both). Quantitative or qualitative deficiency of antithrombin (inborn or acquired, e.g., in proteinuria) leads to thrombophilia.

Tissue factor pathway inhibitor (TFPI)

Tissue factor pathway inhibitor (TFPI) limits the action of tissue factor (TF). It also inhibits excessive TF-mediated activation of FVII and FX.

Plasmin

Plasmin is generated by proteolytic cleavage of plasminogen, a plasma protein synthesized in the liver. This cleavage is catalyzed by tissue plasminogen activator (t-PA), which is synthesized and secreted by endothelium. Plasmin proteolytically cleaves fibrin into fibrin degradation products that inhibit excessive fibrin formation.

Prostacyclin

Prostacyclin (PGI2) is released by endothelium and activates platelet Gs protein-linked receptors. This, in turn, activates adenylyl cyclase, which synthesizes cAMP. cAMP inhibits platelet activation by decreasing cytosolic levels of calcium and, by doing so, inhibits the release of granules that would lead to activation of additional platelets and the coagulation cascade.