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Coagulation Factors
Coagulation Factors Alvin H. Schmaier University of Michigan, Ann Arbor, USA ã 2007 Elsevier Inc. All rights reserved.
Physiologic hemostasis and thrombosis occur in the intravascular compartment. In general, the endothelium functions as a constitutive anticoagulant surface. Circulating platelets form a locus, which, upon activation, coagulation factors assemble and physiologic hemostasis occurs. Activation of platelets results in a hemostatic plug by the adherence of platelets by von Willebrand factor to collagen or glycoprotein Ib-V-IXa on the vessel wall. The close contact of platelets on the vessel wall results in an activation mechanism with inside-out signaling and activation of various platelet receptors. On or about the activating platelet surface, assemblies of coagulation proteins occur leading to their activation and the formation of thrombin. Thrombin is the main clotting enzyme by proteolyzing fibrinogen. It is also a potent platelet activator that proteolyzes its platelet receptors, protease activated receptor 1 and protease activated receptor 4. Activation of platelets induces their aggregation forming a hemostatic plug and the cessation of bleeding (Fig. 1). For the last 40 years, the coagulation system, which is a subset of the hemostatic system, has been known as a series of zymogens that get sequentially activated to serine proteases in a waterfall cascade. This notion, however, is starting to be replaced by a revised hypothesis that the coagulation system is a group of zymogens that when activated become serine proteases in protein assemblies associated with cell membranes. Physiologic hemostasis consists of three protein assemblies. The initiating enzyme of physiologic hemostasis is activated factor VII (factor VIIa) in complex with the protein tissue factor. Tissue factor is ubiquitous in all cells and cryptically present on cell membranes. When expressed, it complexes with factor VI I. The mechanism(s) by which factor VII becomes activated is not completely known, but some factor VIIa appears to be constantly available. The tissue factor-factor VIIa complex has the ability to activate factor IX to factor IXa (intrinsic tenase) or activate factor X by factor Xa (extrinsic tenase). The second critical protein assembly is called tenase. It consists of activated factor IX (factor IXa) in the presence of thrombin activated factor VIII:C (factor VIIIa) that converts zymogen factor X (FX) to activated factor Xa (FXa). The assembly of tenase increases the rate of factor X activation 1.4 X 108 over the rate of factor X activation by factor IXa alone. The third critical protein assembly in physiologic hemostasis is prothrombinase. It consists of the assembly of activated factor X in the presence of thrombin activated factor V converting zymogen prothrombin (Factor II) to thrombin (Factor IIa). The rate of factor Xa activation of factor II alone is increased by 1.7 X 108 in prothrombinase. Once thrombin or factor IIa is formed, the enzyme proteolyzes the substrate fibrinogen to liberate fibrinopeptides A and B and to allow the residual Aalpha, Bbeta, and gamma chains of fibrinogen to associate end-to-end and side-to-side to form soluble cross-linked fibrin into an insoluble fibrin clot. Thrombin-activated factor XIII cross-links fibrin. Thus, physiologic hemostasis can be characterized as a group of zymogen, cofactors, and substrates. There are three groups of proteins that comprise the coagulation proteins: the phospholipid-bound zymogens, the surface-bound zymogens, and the hemostatic cofactors and substrates. The phospholipid-bound zymogens are the so-called vitamin K dependent proteins, factors II (72 kDa), VII (50 kDa), IX (56 kDa), and X (56 kDa). These proteins that have similar structures require vitamin K for an essential carboxylation reaction that takes place on each of these proteins’ glutamic acid residues on their amino terminal ends, 1
2
Coagulation Factors
Fig. 1. Overview of Hemostasis. When a vessel is injured, platelets adhere to the site of injury by von Willebrand factor (VWF). The close contact of platelets leads to their activation. There are multiple platelet receptors, ADP, alpha adrenergic, collagen, platelet activating factor (PAF) , and thrombin. Release of platelet fibrinogen allows for it to bind to the heterodimeric complex of platelet glycoprotein IIb and IIIa (alpha2Bbeta3 integrins). The platelet aggregate forms an initial hemostatic plug. Simultaneous with platelet activation, an assembly of coagulation factor occurs and becomes activated on or about the platelet surface. Physiologic hemostasis is intiated by tissue factor that probably activates factor IX to ; factor IXa, which then activates factor X to factor Xa. Factor Xa activates prothrombin (II) to thrombin which proteolyses fibrinogen (Fb) to form a fibrin clot. Thrombin also activates platelets.
making them alpha-carboxyglutamic acid. This carboxylation reaction allows these proteins to bind to lipid and cell membranes, where they are activated. Without this carboxylation reaction, these proteins do not function properly in the coagulation system. The vitamin K antagonist warfarin inhibits two enzymes essential for vitamin K metabolism for this carboxylation reaction. As a result, all the phospholipid-bound zymogens are abnormal proteins that function as inhibitors to physiologic hemostatic reactions. The importance of the phospholipid-bound, vitamin K-dependent proteins in the coagulation system was emphasized by the creation of each gene deletion in mice. The knockout mouse for factors VII, X, and II are lethal hemorrhagic states. The factor IX knockout, like the clinical human disorder of factor IX deficiency, hemophilia B, is a severe bleeding disorder. It, along with factor VIII deficiency, is probably the most severe bleeding disorder that allows for full gestation and delivery. With the exception of factor IX, the phospholipid-based hemostatic factors VII, X, and II are best measured by the thromboplastin- or tissue factor-induced prothrombin time. The second group of proteins that comprise the so-called coagulation proteins are the surface-bound zymogens of the plasma kallikrein/kinin system. These proteins are factor
Coagulation Factors
XII (Hageman factor), prekallikrein deficiency (Fletcher factor), and factor X I. Deficiencies of factor XII (FXII) (80 kDa) and prekallikrein (PK) (88 kDa) are not associated with bleeding. Factor XI deficiency is associated with a mild bleeding state that becomes manifest after surgery or trauma. Factor XII and prekallikrein along with factor XI (160 kDa) were considered to be part of the physiologic coagulation system because both proteins contribute to the shortening of the coagulation time when blood is collected into glass tubes. This fact is the basis for the surface-activated coagulation assays, the activated partial thromboplastin time (APTT) Proctor and Rapaport (1961) and activated clotting time (ACT). These proteins contribute to these assays because factor XII has the unique property to autoactivate when exposed to an artificial, negatively charged surface like a glass tube Wiggins and Cochrane (1979), Silverberg et al (1980). When factor XII autoactivates, it then activates prekallikrein to plasma kallikrein that then amplifies the activation of more zymogen factor XII to factor XIIa. The activated factor XII then activates factor XI to factor XIa, initiating the cascade of serine proteases that lead to thrombin formation. These reactions are accelerated by their cofactor, high molecular weight kininogen (see below). The fact that this occurs in glass tubes was the basis that allowed for Ratnoff and Davies to propose the coagulation cascade hypothesis forty years ago Ratnoff and Davie (1964), Macfarlane (1964), Roberts (2003) (Fig. 2). The third group of proteins in physiologic hemostasis are the cofactors and substrates for the enzymes of the system. Tissue factor is a 47 kDa protein that is found on the membrane of cells and serves as a critical cofactor and, perhaps, cell-surface receptor for factor VIIa’s activities. The absence of tissue factor is incompatible with life. Factor VIII (antihemophilic factor) (330 kDa) is an essential cofactor when activated by thrombin for factor IXa in tenase. Its deficiency is associated with the most common, severe bleeding disorder, Hemophilia A. Factor V (330 kDa) is an essential cofactor for factor Xa’s activation of prothrombin. Its deficiency is associated with severe hemorrhage in gestation or at birth and is virtually incompatible with life. Both factors VIII and V are structurally similar to each other and ceruloplasm. High molecular weight kininogen (120 kDa) (HK) is a cofactor and substrate of the surface-activated serine proteases factor XIIa, plasma kallikrein, and factor XIa. It is also the parent protein of the biologically active peptide bradykinin. A deficiency of HK gives a long surface-activated coagulation test (APTT,
Fig. 2. The Coagulation Cascade. There are two mechanisms for factor X (X) activation. On a negatively charged surface, factor XII (XII) autoactivates to factor XIIa (XIIa). This activation event is accelerated by prekallikrein (PK) and high molecular weight kininogen (HK). Factor XIIa than activates factor XI (XI) to factor XIa (XIa). Factor XIa activates factor IX (IX) to factor IXa (IXa). Factor IX can also be activated independent of factor XIa by tissue factor (TF) and factor VIIa (VIIa) complex. Factor IXa in the presence of activated factor VIII (VIIIa) activates factor X to factor Xa (Xa). The tissue factor and factor VIIa complex can also activate factor X directly, but the presence of an inhibitor, tissue factor pathway inhibitor, not shown in the figure, probably prevents this pathway under physiologic circumstances. Formed factor Xa in the presence of activated factor V (Va) converts prothrombin (II) to thrombin (IIa). Thrombin then proteolyses fibrinogen to form a fibrin clot.
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Coagulation Factors
ACT) like factor XII without a bleeding state. Thus, it is important to make the distinction between those proteins that contribute to physiologic hemostasis and those proteins (XII, PK, HK) that only contribute to the clinical assays used to assess hemostatic function. The recognition that factor XII, prekallikrein, and high molecular weight kininogen deficiency are not associated with bleeding suggested that another hypothesis for physiologic hemostasis needed to be developed. The first evidence that led to the development of an alternative hypothesis for physiologic hemostasis was the recognition that tissue factor-factor VIIa could directly activate factor IX Osterud and Rapaport (1977). This pathway led to bypass of factor XIa activation of factor I X. Up until recently, see below, factor XI was believed to be activated only by factor XIIa. The second evidence that was essential to develop an alternative hypothesis for physiologic hemostasis with the observation by Gailini and Broze (1991) and Naito and Fujikawa (1991) that thrombin can activate factor XI directly and independent of the proteins of the plasma kallikrein/kinin system. These bypass mechanisms offered physiologically cogent mechanisms for thrombin formation that are independent of factor XII and prekallikrein activation in the presence of high molecular weight kininogen. A revised mechanism for physiologic hemostasis is presented in Fig. 3. In addition to the prothrombotic, thrombin forming hemostatic system, two regulatory systems are present as part of the coagulation system. These two systems include the clot lysing, fibrinolytic system and the proteins that comprise the anticoagulation system. The fibrinolytic system consists of the zymogen plasminogen being activated to the clot lysing enzyme, plasmin, by tissue plasminogen activator (tPA) or single chain urokinase (ScuPA, prourokinase) when activated to two chain urokinase (tcuPA, urokinase). Formed plasmin degrades cross-linked, insoluble and soluble fibrin and soluble fibrinogen into fibrin(ogen) degradation products or lysed clot (Fig. 4).
Fig. 3. Physiologic Hemostatic Mechanism. A current hypothesis for physiologic hemostasis is the initiation of activation by the tissue factor-VIIa complex (TF-VIIa). Since there is sufficient tissue factor pathway inhibitor (TFPI), TF-VIIa activation of factor X to factor Xa (Xa) probably does not occur. Rather TF-VIIa activates factor IX to factor IXa (IXa). Factor IXa in the presence of activated factor VIII (VIIIa) activates factor Xa to Xa. Factor Xa in the presence of activated factor V (Va) activates prothrombin to thrombin (IIa). Thrombin proteolyzes fibrinogen to form fibrin. If more thrombin is needed, it can activate factor XI (XI) to factor XIa (XIa). The XIa then activates factor IX to IXa to amplify the system. Increased IIa can prevent fibrinolysis by activating carboxypeptidase U [thrombin activable fibrinolysis inhibitor TAFIa)] to reduce fibrinolysis of formed fibrin by inactivating plasmin. This mechanism also provides for an alternative means for factor XI activation. Factor XI can be activated by the plasma kallikrein/kinin system, but since deficiencies of factor XII (XII), prekallikrein (PK), and high molecular weight kininogen (HK) are not associated with bleeding defects, this pathway of activation is not physiologic. Physiologic activation of the plasma kallikrein/kinin system probably arises by prekallikrein activation on endothelial cells bound to HK by the enzyme, prolylcarboxypeptidase (PRCP) Shariat-Madar et al (2002). Formed kallikrein (HK-K) bound to HK then activates XII to factor XIIa (XIIa). Factor XIIa can activate PK to kallikrein to amplify activation of this system. Factor XIIa can also activate factor XI to XIa. This latter mechanism may be important in pathophysiologic circumstances such as sepsis.
Coagulation Factors
Fig. 4. The Fibrinolytic System. Zymogen plasminogen is activated to plasmin by tissue plasminogen activator (tPA), single chain urokinase plasminogen activator (ScuPA), or two chain urokinase plasminogen activator (TcuPA). Plasmin then proteolyses fibrin or fibrinogen to degrade these proteins and form fibrin or fibrinogen degradation products (FDP).
The plasma anticoagulation system has two major components. The major in vivo anticoagulation system consists of the protein C-protein S system. Protein C is a 62 kDa vitamin K–dependent zymogen, which binds to an endothelial cell membrane protein called thrombomodulin. Thrombin activates protein C bound to thrombomodulin to make activated protein C. Activated protein C binds to protein S, a 69 kDa vitamin K–dependent protein that allows activated protein C to degrade factors Va and VIIIa to make them inactive to serve as cofactors for the generation of thrombin or factor Xa in prothrombinase or tenase, respectively. Activated protein C also binds to the endothelial cell protein C receptor (EPCR) to proteolyze protease activated receptor 1 on endothelial cells to stimulate the liberation of tPA that activates plasminogen (Fig. 5). The second anticoagulation system in vivo of the coagulation system is the group of SERPINS, serine protease inhibitors. The major SERPIN in plasma of the hemostatic system is antithrombin, a 58 kDa protein that inhibits thrombin and factor Xa as well as factors XIIa, XIa, IXa, VIIa, and plasma kallikrein. The importance of antithrombin is that its gene knockout in mice is incompatible with normal gestation. Another SERPIN, C1 inhibitor is a major regulator of factor XIIa, plasma kallikrein, and factor XIa in plasma.
Fig. 5. The Protein C Anticoagulation System. Protein C is binds to thrombomodulin and is activated by thrombin to activated protein C (APC). APC binds to the endothelial cell protein C receptor (EPCR) to activate protease activated receptor 1 (PAR1) on endothelial cells. This mechanism results in tissue plasminogen activator release from endothelial cells and induction of fibrinolysis. APC in the presence of its cofactor, protein S (PS), inactivates activated forms of factor VIIIa (VIIIa) and factor Va (Va) into inactive forms (VIIIi and Vi, respectively). Inactivation of VIIIa and Va prevents the cofactor activity of these proteins for factor X (X) and factor II (II) activation to factor Xa (Xa) and thrombin (IIa), respectively, to generate more clot. Reduction of thrombin formation and stimulation of fibrinolysis is the combined mechanisms by which the protein C system functions as the major anticoagulant system.
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Coagulation Factors
Plasminogen activator inhibitor-1 (PAI-1) is the major regulator of tPA and tcuP A. PAI-1 is also a good inhibitor of plasma kallikrein and factor XIa. Heparin cofactor II and protease nexin I are two SERPINS that exclusively serve as thrombin inhibitors. Protein Z inhibitor is a SERPIN that in the presence of protein Z, a vitamin K-dependent protein, is an inhibitor of factor Xa. Alpha-2-antiplasmin is the major SERPIN that regulates plasmin. Tissue factor pathway inhibitor (TFPI) is a Kunitz-type serine protease inhibitor of the factor VIIa-tissue factor complex bound to factor X/Xa. Last, the amyloid betaprotein precursor inhibits coagulation factors XIa, IXa, VIIa, and X. The exact physiologic role of this latter Kunitz-type serine protease inhibitor is not known, but it may serve as a cerebral anticoagulant.
Journal Citations Gailani, D., Broze, G.J., 1991. Factor XI activation in a revised model of blood coagulation. Science, 253, 909–912. Macfarlane, R.G., 1964. An enzyme cascade in the blood clotting mechanism, and its function as a biochemical amplifier. Nature, 202, 498–499. Naito, K., Fujikawa, K., 1991. Activation of human blood coagulation factor XI independent of factor XI I. Factor XI is activated by thrombin and factor XIa in the presence of negatively charged surfaces. J.Biological Chemistry, 266, 7353–7358. Osterud, B., Rapaport, S.I., 1977. Activation of factor IX by the reaction product of tissue factor and factor VII: additional pathway for initiating blood coagulation. Proc. Natl. Acad. Sci., 74, 5260–5264. Proctor, R.R., Rapaport, S.I., 1961. The partial thromboplastin time with kaolin: A simple screening test for first stage clotting factor deficiencies. Amer. J. Clin. Path., 36, 212–219. Ratnoff, O.D., Davie, E.W., 1964. Waterfall sequence for intrinsic blood clotting. Science, 145, 1310–1312. Roberts, H.R., 2003. Oscar Ratnoff: His contributions to the golder era of coagulation research. British J. Haematology, 122, 180–192. Shariat-Madar, Z., Mahdi, F., Schmaier, A.H., 2002. Identification and characterization of prolylcarboxypeptidase as an endothelial cell prekallikrein activator. J. Biological Chemistry, 277, 17962–17969. Silverberg, M., Dunn, J.T., Garen, L., Kaplan, A.P., 1980. Autoactivation of human Hageman factor. Demonstration utilizing a synthetic substrate. J. Biological Chemistry, 255, 7281–7286. Wiggins, R.C., Cochrane, C.C., 1979. The autoactivation of rabbit Hageman factor. J. Experimental Medicine, 150, 1122–1133.
Physiologic hemostasis and thrombosis occur in the intravascular compartment. In general, the endothelium functions as a constitutive anticoagulant surface. Circulating platelets form a locus, which, upon activation, coagulation factors assemble and physiologic hemostasis occurs. Activation of platelets results in a hemostatic plug by the adherence of platelets by von Willebrand factor to collagen or glycoprotein Ib-V-IXa on the vessel wall. The close contact of platelets on the vessel wall results in an activation mechanism with inside-out signaling and activation of various platelet receptors. On or about the activating platelet surface, assemblies of coagulation proteins occur leading to their activation and the formation of thrombin. Thrombin is the main clotting enzyme by proteolyzing fibrinogen. It is also a potent platelet activator that proteolyzes its platelet receptors, protease activated receptor 1 and protease activated receptor 4. Activation of platelets induces their aggregation forming a hemostatic plug and the cessation of bleeding (Fig. 1). For the last 40 years, the coagulation system, which is a subset of the hemostatic system, has been known as a series of zymogens that get sequentially activated to serine proteases in a waterfall cascade. This notion, however, is starting to be replaced by a revised hypothesis that the coagulation system is a group of zymogens that when activated become serine proteases in protein assemblies associated with cell membranes. Physiologic hemostasis consists of three protein assemblies. The initiating enzyme of physiologic hemostasis is activated factor VII (factor VIIa) in complex with the protein tissue factor. Tissue factor is ubiquitous in all cells and cryptically present on cell membranes. When expressed, it complexes with factor VI I. The mechanism(s) by which factor VII becomes activated is not completely known, but some factor VIIa appears to be constantly available. The tissue factor-factor VIIa complex has the ability to activate factor IX to factor IXa (intrinsic tenase) or activate factor X by factor Xa (extrinsic tenase). The second critical protein assembly is called tenase. It consists of activated factor IX (factor IXa) in the presence of thrombin activated factor VIII:C (factor VIIIa) that converts zymogen factor X (FX) to activated factor Xa (FXa). The assembly of tenase increases the rate of factor X activation 1.4 X 108 over the rate of factor X activation by factor IXa alone. The third critical protein assembly in physiologic hemostasis is prothrombinase. It consists of the assembly of activated factor X in the presence of thrombin activated factor V converting zymogen prothrombin (Factor II) to thrombin (Factor IIa). The rate of factor Xa activation of factor II alone is increased by 1.7 X 108 in prothrombinase. Once thrombin or factor IIa is formed, the enzyme proteolyzes the substrate fibrinogen to liberate fibrinopeptides A and B and to allow the residual Aalpha, Bbeta, and gamma chains of fibrinogen to associate end-to-end and side-to-side to form soluble cross-linked fibrin into an insoluble fibrin clot. Thrombin-activated factor XIII cross-links fibrin. Thus, physiologic hemostasis can be characterized as a group of zymogen, cofactors, and substrates. There are three groups of proteins that comprise the coagulation proteins: the phospholipid-bound zymogens, the surface-bound zymogens, and the hemostatic cofactors and substrates. The phospholipid-bound zymogens are the so-called vitamin K dependent proteins, factors II (72 kDa), VII (50 kDa), IX (56 kDa), and X (56 kDa). These proteins that have similar structures require vitamin K for an essential carboxylation reaction that takes place on each of these proteins’ glutamic acid residues on their amino terminal ends, 1
2
Coagulation Factors
Fig. 1. Overview of Hemostasis. When a vessel is injured, platelets adhere to the site of injury by von Willebrand factor (VWF). The close contact of platelets leads to their activation. There are multiple platelet receptors, ADP, alpha adrenergic, collagen, platelet activating factor (PAF) , and thrombin. Release of platelet fibrinogen allows for it to bind to the heterodimeric complex of platelet glycoprotein IIb and IIIa (alpha2Bbeta3 integrins). The platelet aggregate forms an initial hemostatic plug. Simultaneous with platelet activation, an assembly of coagulation factor occurs and becomes activated on or about the platelet surface. Physiologic hemostasis is intiated by tissue factor that probably activates factor IX to ; factor IXa, which then activates factor X to factor Xa. Factor Xa activates prothrombin (II) to thrombin which proteolyses fibrinogen (Fb) to form a fibrin clot. Thrombin also activates platelets.
making them alpha-carboxyglutamic acid. This carboxylation reaction allows these proteins to bind to lipid and cell membranes, where they are activated. Without this carboxylation reaction, these proteins do not function properly in the coagulation system. The vitamin K antagonist warfarin inhibits two enzymes essential for vitamin K metabolism for this carboxylation reaction. As a result, all the phospholipid-bound zymogens are abnormal proteins that function as inhibitors to physiologic hemostatic reactions. The importance of the phospholipid-bound, vitamin K-dependent proteins in the coagulation system was emphasized by the creation of each gene deletion in mice. The knockout mouse for factors VII, X, and II are lethal hemorrhagic states. The factor IX knockout, like the clinical human disorder of factor IX deficiency, hemophilia B, is a severe bleeding disorder. It, along with factor VIII deficiency, is probably the most severe bleeding disorder that allows for full gestation and delivery. With the exception of factor IX, the phospholipid-based hemostatic factors VII, X, and II are best measured by the thromboplastin- or tissue factor-induced prothrombin time. The second group of proteins that comprise the so-called coagulation proteins are the surface-bound zymogens of the plasma kallikrein/kinin system. These proteins are factor
Coagulation Factors
XII (Hageman factor), prekallikrein deficiency (Fletcher factor), and factor X I. Deficiencies of factor XII (FXII) (80 kDa) and prekallikrein (PK) (88 kDa) are not associated with bleeding. Factor XI deficiency is associated with a mild bleeding state that becomes manifest after surgery or trauma. Factor XII and prekallikrein along with factor XI (160 kDa) were considered to be part of the physiologic coagulation system because both proteins contribute to the shortening of the coagulation time when blood is collected into glass tubes. This fact is the basis for the surface-activated coagulation assays, the activated partial thromboplastin time (APTT) Proctor and Rapaport (1961) and activated clotting time (ACT). These proteins contribute to these assays because factor XII has the unique property to autoactivate when exposed to an artificial, negatively charged surface like a glass tube Wiggins and Cochrane (1979), Silverberg et al (1980). When factor XII autoactivates, it then activates prekallikrein to plasma kallikrein that then amplifies the activation of more zymogen factor XII to factor XIIa. The activated factor XII then activates factor XI to factor XIa, initiating the cascade of serine proteases that lead to thrombin formation. These reactions are accelerated by their cofactor, high molecular weight kininogen (see below). The fact that this occurs in glass tubes was the basis that allowed for Ratnoff and Davies to propose the coagulation cascade hypothesis forty years ago Ratnoff and Davie (1964), Macfarlane (1964), Roberts (2003) (Fig. 2). The third group of proteins in physiologic hemostasis are the cofactors and substrates for the enzymes of the system. Tissue factor is a 47 kDa protein that is found on the membrane of cells and serves as a critical cofactor and, perhaps, cell-surface receptor for factor VIIa’s activities. The absence of tissue factor is incompatible with life. Factor VIII (antihemophilic factor) (330 kDa) is an essential cofactor when activated by thrombin for factor IXa in tenase. Its deficiency is associated with the most common, severe bleeding disorder, Hemophilia A. Factor V (330 kDa) is an essential cofactor for factor Xa’s activation of prothrombin. Its deficiency is associated with severe hemorrhage in gestation or at birth and is virtually incompatible with life. Both factors VIII and V are structurally similar to each other and ceruloplasm. High molecular weight kininogen (120 kDa) (HK) is a cofactor and substrate of the surface-activated serine proteases factor XIIa, plasma kallikrein, and factor XIa. It is also the parent protein of the biologically active peptide bradykinin. A deficiency of HK gives a long surface-activated coagulation test (APTT,
Fig. 2. The Coagulation Cascade. There are two mechanisms for factor X (X) activation. On a negatively charged surface, factor XII (XII) autoactivates to factor XIIa (XIIa). This activation event is accelerated by prekallikrein (PK) and high molecular weight kininogen (HK). Factor XIIa than activates factor XI (XI) to factor XIa (XIa). Factor XIa activates factor IX (IX) to factor IXa (IXa). Factor IX can also be activated independent of factor XIa by tissue factor (TF) and factor VIIa (VIIa) complex. Factor IXa in the presence of activated factor VIII (VIIIa) activates factor X to factor Xa (Xa). The tissue factor and factor VIIa complex can also activate factor X directly, but the presence of an inhibitor, tissue factor pathway inhibitor, not shown in the figure, probably prevents this pathway under physiologic circumstances. Formed factor Xa in the presence of activated factor V (Va) converts prothrombin (II) to thrombin (IIa). Thrombin then proteolyses fibrinogen to form a fibrin clot.
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Coagulation Factors
ACT) like factor XII without a bleeding state. Thus, it is important to make the distinction between those proteins that contribute to physiologic hemostasis and those proteins (XII, PK, HK) that only contribute to the clinical assays used to assess hemostatic function. The recognition that factor XII, prekallikrein, and high molecular weight kininogen deficiency are not associated with bleeding suggested that another hypothesis for physiologic hemostasis needed to be developed. The first evidence that led to the development of an alternative hypothesis for physiologic hemostasis was the recognition that tissue factor-factor VIIa could directly activate factor IX Osterud and Rapaport (1977). This pathway led to bypass of factor XIa activation of factor I X. Up until recently, see below, factor XI was believed to be activated only by factor XIIa. The second evidence that was essential to develop an alternative hypothesis for physiologic hemostasis with the observation by Gailini and Broze (1991) and Naito and Fujikawa (1991) that thrombin can activate factor XI directly and independent of the proteins of the plasma kallikrein/kinin system. These bypass mechanisms offered physiologically cogent mechanisms for thrombin formation that are independent of factor XII and prekallikrein activation in the presence of high molecular weight kininogen. A revised mechanism for physiologic hemostasis is presented in Fig. 3. In addition to the prothrombotic, thrombin forming hemostatic system, two regulatory systems are present as part of the coagulation system. These two systems include the clot lysing, fibrinolytic system and the proteins that comprise the anticoagulation system. The fibrinolytic system consists of the zymogen plasminogen being activated to the clot lysing enzyme, plasmin, by tissue plasminogen activator (tPA) or single chain urokinase (ScuPA, prourokinase) when activated to two chain urokinase (tcuPA, urokinase). Formed plasmin degrades cross-linked, insoluble and soluble fibrin and soluble fibrinogen into fibrin(ogen) degradation products or lysed clot (Fig. 4).
Fig. 3. Physiologic Hemostatic Mechanism. A current hypothesis for physiologic hemostasis is the initiation of activation by the tissue factor-VIIa complex (TF-VIIa). Since there is sufficient tissue factor pathway inhibitor (TFPI), TF-VIIa activation of factor X to factor Xa (Xa) probably does not occur. Rather TF-VIIa activates factor IX to factor IXa (IXa). Factor IXa in the presence of activated factor VIII (VIIIa) activates factor Xa to Xa. Factor Xa in the presence of activated factor V (Va) activates prothrombin to thrombin (IIa). Thrombin proteolyzes fibrinogen to form fibrin. If more thrombin is needed, it can activate factor XI (XI) to factor XIa (XIa). The XIa then activates factor IX to IXa to amplify the system. Increased IIa can prevent fibrinolysis by activating carboxypeptidase U [thrombin activable fibrinolysis inhibitor TAFIa)] to reduce fibrinolysis of formed fibrin by inactivating plasmin. This mechanism also provides for an alternative means for factor XI activation. Factor XI can be activated by the plasma kallikrein/kinin system, but since deficiencies of factor XII (XII), prekallikrein (PK), and high molecular weight kininogen (HK) are not associated with bleeding defects, this pathway of activation is not physiologic. Physiologic activation of the plasma kallikrein/kinin system probably arises by prekallikrein activation on endothelial cells bound to HK by the enzyme, prolylcarboxypeptidase (PRCP) Shariat-Madar et al (2002). Formed kallikrein (HK-K) bound to HK then activates XII to factor XIIa (XIIa). Factor XIIa can activate PK to kallikrein to amplify activation of this system. Factor XIIa can also activate factor XI to XIa. This latter mechanism may be important in pathophysiologic circumstances such as sepsis.
Coagulation Factors
Fig. 4. The Fibrinolytic System. Zymogen plasminogen is activated to plasmin by tissue plasminogen activator (tPA), single chain urokinase plasminogen activator (ScuPA), or two chain urokinase plasminogen activator (TcuPA). Plasmin then proteolyses fibrin or fibrinogen to degrade these proteins and form fibrin or fibrinogen degradation products (FDP).
The plasma anticoagulation system has two major components. The major in vivo anticoagulation system consists of the protein C-protein S system. Protein C is a 62 kDa vitamin K–dependent zymogen, which binds to an endothelial cell membrane protein called thrombomodulin. Thrombin activates protein C bound to thrombomodulin to make activated protein C. Activated protein C binds to protein S, a 69 kDa vitamin K–dependent protein that allows activated protein C to degrade factors Va and VIIIa to make them inactive to serve as cofactors for the generation of thrombin or factor Xa in prothrombinase or tenase, respectively. Activated protein C also binds to the endothelial cell protein C receptor (EPCR) to proteolyze protease activated receptor 1 on endothelial cells to stimulate the liberation of tPA that activates plasminogen (Fig. 5). The second anticoagulation system in vivo of the coagulation system is the group of SERPINS, serine protease inhibitors. The major SERPIN in plasma of the hemostatic system is antithrombin, a 58 kDa protein that inhibits thrombin and factor Xa as well as factors XIIa, XIa, IXa, VIIa, and plasma kallikrein. The importance of antithrombin is that its gene knockout in mice is incompatible with normal gestation. Another SERPIN, C1 inhibitor is a major regulator of factor XIIa, plasma kallikrein, and factor XIa in plasma.
Fig. 5. The Protein C Anticoagulation System. Protein C is binds to thrombomodulin and is activated by thrombin to activated protein C (APC). APC binds to the endothelial cell protein C receptor (EPCR) to activate protease activated receptor 1 (PAR1) on endothelial cells. This mechanism results in tissue plasminogen activator release from endothelial cells and induction of fibrinolysis. APC in the presence of its cofactor, protein S (PS), inactivates activated forms of factor VIIIa (VIIIa) and factor Va (Va) into inactive forms (VIIIi and Vi, respectively). Inactivation of VIIIa and Va prevents the cofactor activity of these proteins for factor X (X) and factor II (II) activation to factor Xa (Xa) and thrombin (IIa), respectively, to generate more clot. Reduction of thrombin formation and stimulation of fibrinolysis is the combined mechanisms by which the protein C system functions as the major anticoagulant system.
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Coagulation Factors
Plasminogen activator inhibitor-1 (PAI-1) is the major regulator of tPA and tcuP A. PAI-1 is also a good inhibitor of plasma kallikrein and factor XIa. Heparin cofactor II and protease nexin I are two SERPINS that exclusively serve as thrombin inhibitors. Protein Z inhibitor is a SERPIN that in the presence of protein Z, a vitamin K-dependent protein, is an inhibitor of factor Xa. Alpha-2-antiplasmin is the major SERPIN that regulates plasmin. Tissue factor pathway inhibitor (TFPI) is a Kunitz-type serine protease inhibitor of the factor VIIa-tissue factor complex bound to factor X/Xa. Last, the amyloid betaprotein precursor inhibits coagulation factors XIa, IXa, VIIa, and X. The exact physiologic role of this latter Kunitz-type serine protease inhibitor is not known, but it may serve as a cerebral anticoagulant.
Journal Citations Gailani, D., Broze, G.J., 1991. Factor XI activation in a revised model of blood coagulation. Science, 253, 909–912. Macfarlane, R.G., 1964. An enzyme cascade in the blood clotting mechanism, and its function as a biochemical amplifier. Nature, 202, 498–499. Naito, K., Fujikawa, K., 1991. Activation of human blood coagulation factor XI independent of factor XI I. Factor XI is activated by thrombin and factor XIa in the presence of negatively charged surfaces. J.Biological Chemistry, 266, 7353–7358. Osterud, B., Rapaport, S.I., 1977. Activation of factor IX by the reaction product of tissue factor and factor VII: additional pathway for initiating blood coagulation. Proc. Natl. Acad. Sci., 74, 5260–5264. Proctor, R.R., Rapaport, S.I., 1961. The partial thromboplastin time with kaolin: A simple screening test for first stage clotting factor deficiencies. Amer. J. Clin. Path., 36, 212–219. Ratnoff, O.D., Davie, E.W., 1964. Waterfall sequence for intrinsic blood clotting. Science, 145, 1310–1312. Roberts, H.R., 2003. Oscar Ratnoff: His contributions to the golder era of coagulation research. British J. Haematology, 122, 180–192. Shariat-Madar, Z., Mahdi, F., Schmaier, A.H., 2002. Identification and characterization of prolylcarboxypeptidase as an endothelial cell prekallikrein activator. J. Biological Chemistry, 277, 17962–17969. Silverberg, M., Dunn, J.T., Garen, L., Kaplan, A.P., 1980. Autoactivation of human Hageman factor. Demonstration utilizing a synthetic substrate. J. Biological Chemistry, 255, 7281–7286. Wiggins, R.C., Cochrane, C.C., 1979. The autoactivation of rabbit Hageman factor. J. Experimental Medicine, 150, 1122–1133.