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David Gailani - One of the best experts on this subject based on the ideXlab platform.
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endothelial pai 1 plasminogen activator inhibitor 1 blocks the Intrinsic Pathway of coagulation inducing the clearance and degradation of fxia activated factor xi
Arteriosclerosis Thrombosis and Vascular Biology, 2019Co-Authors: Cristina Puy, David Gailani, Anh T P Ngo, Jiaqing Pang, Ravi S Keshari, Matthew W Hagen, Monica T Hinds, Andras Gruber, Florea Lupu, Owen J T MccartyAbstract:Objective- Activation of coagulation FXI (factor XI) by FXIIa (activated factor XII) is a prothrombotic process. The endothelium is known to play an antithrombotic role by limiting thrombin generation and platelet activation. It is unknown whether the antithrombotic role of the endothelium includes sequestration of FXIa (activated factor XI) activity. This study aims to determine the role of endothelial cells (ECs) in the regulation of the Intrinsic Pathway of coagulation. Approach and Results- Using a chromogenic assay, we observed that human umbilical veins ECs selectively blocked FXIa yet supported kallikrein and FXIIa activity. Western blotting and mass spectrometry analyses revealed that FXIa formed a complex with endothelial PAI-1 (plasminogen activator inhibitor-1). Blocking endothelial PAI-1 increased the cleavage of a chromogenic substrate by FXIa and the capacity of FXIa to promote fibrin formation in plasma. Western blot and immunofluorescence analyses showed that FXIa-PAI-1 complexes were either released into the media or trafficked to the early and late endosomes and lysosomes of ECs. When baboons were challenged with Staphylococcus aureus to induce a prothrombotic phenotype, an increase in circulating FXIa-PAI-1 complex levels was detected by ELISA within 2 to 8 hours postchallenge. Conclusions- PAI-1 forms a complex with FXIa on ECs, blocking its activity and inducing the clearance and degradation of FXIa. Circulating FXIa-PAI-1 complexes were detected in a baboon model of S. aureus sepsis. Although ECs support kallikrein and FXIIa activity, inhibition of FXIa by ECs may promote the clearance of intravascular FXIa. Visual Overview- An online visual overview is available for this article.
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The Intrinsic Pathway of Coagulation as a Target for Antithrombotic Therapy.
Hematology oncology clinics of North America, 2016Co-Authors: Allison P. Wheeler, David GailaniAbstract:Plasma coagulation in the activated partial thromboplastin time assay is initiated by sequential activation of coagulation factors XII, XI, and IX. While this series of proteolytic reactions is not an accurate model for hemostasis in vivo, there is mounting evidence that factor XI and factor XII contribute to thrombosis, and that inhibiting them can produce an antithrombotic effect with a small effect on hemostasis. This article discusses the contributions of components of the Intrinsic Pathway to thrombosis in animal models and humans, and results of early clinical trials of drugs targeting factors IX, XI, and XII.
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Intrinsic Pathway of Coagulation and Arterial Thrombosis
Arteriosclerosis thrombosis and vascular biology, 2007Co-Authors: David Gailani, Thomas RennéAbstract:Formation of a fibrin clot is mediated by a group of tightly regulated plasma proteases and cofactors. While this system is essential for minimizing blood loss from an injured blood vessel (hemostasis), it also contributes to pathologic fibrin formation and platelet activation that may occlude vessels (thrombosis). Many antithrombotic drugs target key elements of the plasma coagulation mechanism such as thrombin and factor Xa, based on the premise that plasma elements contributing to thrombosis are primarily those involved in hemostasis. Recent studies with genetically altered mice raise questions about this paradigm. Deficiencies of the Intrinsic Pathway proteases factor XII and factor XI are not associated with abnormal hemostasis in mice, but impair formation of occlusive thrombi in arterial injury models, indicating that Pathways not essential for hemostasis participate in arterial thrombosis. If factor XII or factor XI make similar contributions to thrombosis in humans, these proteases could be ideal targets for drugs to treat or prevent thromboembolic disease with minimal risk of therapy-associated bleeding.
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The Intrinsic Pathway of coagulation: a target for treating thromboembolic disease?
Journal of thrombosis and haemostasis : JTH, 2007Co-Authors: David Gailani, Thomas RennéAbstract:Summary. The classic Intrinsic Pathway of coagulation is triggered by contact activation of the plasma protease factor (F)XII, followed by sequential proteolytic activation of FX1 and FIX. While a key mechanism for initiating coagulation in some clinically useful in vitro assays, the absence of abnormal bleeding associated with congenital FXII deficiency indicates that the Intrinsic Pathway is not important for normal blood coagulation in vivo. However, recent work with mice lacking FXII or FXI suggest that these proteases make important contributions to formation of pathologic intravascular thrombi. In models of arterial injury, FXII or FXI null mice are protected from formation of platelet rich occlusive thrombi to a degree similar to that seen in FIX deficient mice (a model for the severe bleeding disorder hemophilia B) or to wild type mice treated with high dose heparin. FXII or FXI deficiency does not appear to prevent the initiation of thrombus formation in these models, but instead causes significant thrombus instability that prevents occlusion of the vessel. These findings raise the possibility that a Pathway similar or identical to the Intrinsic Pathway may operate in vivo under some circumstances. Furthermore, the disproportionate importance of FXII and FXI to occlusive thrombus formation compared to normal hemostasis makes these proteases attractive candidates for therapeutic inhibitors to treat or prevent thromboembolic disorders.
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targeting coagulation factor xii provides protection from pathological thrombosis in cerebral ischemia without interfering with hemostasis
Journal of Experimental Medicine, 2006Co-Authors: Christoph Kleinschnitz, David Gailani, Bernhard Nieswandt, Guido Stoll, Martin Bendszus, Kai Schuh, H U Pauer, Peter Burfeind, Christoph Renne, Thomas RennéAbstract:Formation of fibrin is critical for limiting blood loss at a site of blood vessel injury (hemostasis), but may also contribute to vascular thrombosis. Hereditary deficiency of factor XII (FXII), the protease that triggers the Intrinsic Pathway of coagulation in vitro, is not associated with spontaneous or excessive injury-related bleeding, indicating FXII is not required for hemostasis. We demonstrate that deficiency or inhibition of FXII protects mice from ischemic brain injury. After transient middle cerebral artery occlusion, the volume of infarcted brain in FXII-deficient and FXII inhibitor–treated mice was substantially less than in wild-type controls, without an increase in infarct-associated hemorrhage. Targeting FXII reduced fibrin formation in ischemic vessels, and reconstitution of FXII-deficient mice with human FXII restored fibrin deposition. Mice deficient in the FXII substrate factor XI were similarly protected from vessel-occluding fibrin formation, suggesting that FXII contributes to pathologic clotting through the Intrinsic Pathway. These data demonstrate that some processes involved in pathologic thrombus formation are distinct from those required for normal hemostasis. As FXII appears to be instrumental in pathologic fibrin formation but dispensable for hemostasis, FXII inhibition may offer a selective and safe strategy for preventing stroke and other thromboembolic diseases.
Pengfei Jiang - One of the best experts on this subject based on the ideXlab platform.
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the extrinsic coagulation cascade and tissue factor Pathway inhibitor in macrophages a potential therapeutic opportunity for atherosclerotic thrombosis
Thrombosis Research, 2014Co-Authors: Pengfei Jiang, Dong Xue, Yingjia Zhang, Yuan Liu, Milan Makale, Santosh Kesari, Thomas S Edgington, Cheng LiuAbstract:Abstract Objectives The coagulation protease cascade plays the central requisite role in initiation of arterial atherothrombosis. However, the relative participation of the extrinsic as compared to the Intrinsic Pathway is incompletely resolved. We have investigated in vivo the relative importance of the extrinsic and Intrinsic Pathways to define which is more essential to atherothrombosis and therefore the preferable prophylactic therapeutic target. We further addressed which type of plaque associated macrophage population is associated with the thrombotic propensity of atherosclerotic plaques. Methods Both photochemical injury and ferric chloride vascular injury models demonstrated arterial thrombosis formation in ApoE deficient mice. We found that direct interference with the extrinsic Pathway, but not the Intrinsic Pathway, markedly diminished the rate of thrombus formation and occlusion of atherosclerotic carotid arteries following experimental challenge. To explore which plaque macrophage subtype may participate in plaque thrombosis in regard to expression tissue factor Pathway inhibitor (TFPI), bone marrow derived macrophages of both M and GM phenotypes expressed tissue factor (TF), but the level of TFPI was much greater in M- type macrophages, which exhibited diminished thrombogenic activity, compared to type GM-macrophages. Results and conclusions Our works support the hypothesis that the TF-initiated and direct extrinsic Pathway provides the more significant contribution to arterial plaque thrombogenesis. Activation of the TF driven extrinsic Pathway can be influenced by differing colony-stimulating factor influenced macrophage TFPI-1 expression. These results advance our understanding of atherothrombosis and identify potential therapeutic targets associated with the extrinsic Pathway and with macrophages populating arterial atherosclerotic plaques.
Nigel Mackman - One of the best experts on this subject based on the ideXlab platform.
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Intrinsic Pathway of Coagulation and Thrombosis.
Arteriosclerosis thrombosis and vascular biology, 2019Co-Authors: Steven P. Grover, Nigel MackmanAbstract:Activation of the Intrinsic Pathway of coagulation contributes to the pathogenesis of arterial and venous thrombosis. Critical insights into the involvement of Intrinsic Pathway factors have been derived from the study of gene-specific knockout animals and targeted inhibitors. Importantly, preclinical studies have indicated that targeting components of this Pathway, including FXI (factor XI), FXII, and PKK (prekallikrein), reduces thrombosis with no significant effect on protective hemostatic Pathways. This review highlights the advances made from studying the Intrinsic Pathway using gene-specific knockout animals and inhibitors in models of arterial and venous thrombosis. Development of inhibitors of activated FXI and FXII may reduce thrombosis with minimal increases in bleeding compared with current anticoagulant drugs.
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the role of tissue factor and factor viia in hemostasis
Anesthesia & Analgesia, 2009Co-Authors: Nigel MackmanAbstract:The coagulation system has been historically divided into three Pathways: the extrinsic, Intrinsic, and common Pathways (Fig. 1).1–3 The tissue factor (TF): Factor VII/VIIa (FVII/FVIIa) complex was called the “extrinsic” Pathway because an exogenous agent (i.e., TF) was required for activation of the clotting factors in plasma. The TF:FVIIa complex is the key initiator of the coagulation protease cascade and activates both FIX to FIXa and FX to FXa. This leads to the formation of low amounts of thrombin, which activates the cofactors FV and FVIII. The prothrombin time is determined by adding exogenous TF to plasma and is used to assess the activity of the extrinsic Pathway. Components of the “Intrinsic” Pathway (FXII, FXI, FIX, and FVIII) are all present in blood.4 The tenase complex (FVIIIa:FIXa) plays a key role in amplifying the clotting cascade by activating FX to FXa. The activated partial prothrombin time uses kaolin or other negatively charged substances to activate FXII and is used to assess the activity of the Intrinsic Pathway. The prothrombinase complex (FVa:FXa) and thrombin are referred to as the common Pathway. The prothrombinase complex activates prothrombin to thrombin, which is the central protease of the clotting cascade. Thrombin cleaves fibrinogen into soluble fibrin monomers that polymerize. Thrombin also activates the transglutaminase FXIII to FXIIIa that in turn cross-links soluble fibrin monomers into a fibrin matrix. Finally, thrombin activates platelets by cleavage of protease activated receptors.5 Figure 1 Simplified version of the clotting cascade. Tissue factor (TF) and FVIIa comprise the extrinsic Pathway. FXIIa, FXIa, FIXa, and FVIIIa are members of the Intrinsic Pathway. The common Pathway is made up of FVa, FXa, and thrombin. Thrombin cleaves fibrinogen, ... The coagulation cascade is regulated by several anticoagulants. A kunitz-type serine protease inhibitor called TF Pathway inhibitor inhibits the TF:FVIIa complex by forming a quaternary complex with FXa.6 Protein C is converted to activated protein C by a thrombin-thrombomodulin complex located on the surface of endothelial cells.7 Activated protein C in association with its cofactor protein S cleaves and inactivates the cofactors FV and FVIII. The primary target of the anticoagulant protein antithrombin is thrombin, although it can also inactivate other coagulation proteases in the cascade, including FIXa, FXa, FXIa, and FXIIa.8
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role of the extrinsic Pathway of blood coagulation in hemostasis and thrombosis
Arteriosclerosis Thrombosis and Vascular Biology, 2007Co-Authors: Nigel Mackman, Rachel E TilleyAbstract:Hemostasis requires both platelets and the coagulation system. At sites of vessel injury, bleeding is minimized by the formation of a hemostatic plug consisting of platelets and fibrin. The traditional view of the regulation of blood coagulation is that the initiation phase is triggered by the extrinsic Pathway, whereas amplification requires the Intrinsic Pathway. The extrinsic Pathway consists of the transmembrane receptor tissue factor (TF) and plasma factor VII/VIIa (FVII/FVIIa), and the Intrinsic Pathway consists of plasma FXI, FIX, and FVIII. Under physiological conditions, TF is constitutively expressed by adventitial cells surrounding blood vessels and initiates clotting. In addition so-called blood-borne TF in the form of cell-derived microparticles (MPs) and TF expression within platelets suggests that TF may play a role in the amplification phase of the coagulation cascade. Under pathologic conditions, TF is expressed by monocytes, neutrophils, endothelial cells, and platelets, which results in an elevation of the levels of circulating TF-positive MPs. TF expression within the vasculature likely contributes to thrombosis in a variety of diseases. Understanding how the extrinsic Pathway of blood coagulation contributes to hemostasis and thrombosis may lead to the development of safe and effective hemostatic agents and antithrombotic drugs.
Thomas Renné - One of the best experts on this subject based on the ideXlab platform.
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novel roles for factor xii driven plasma contact activation system
Current Opinion in Hematology, 2008Co-Authors: Felicitas Muller, Thomas RennéAbstract:Purpose of reviewBlood coagulation is a tightly regulated process, involving vascular endothelium, platelets, and plasma coagulation factors. Formation of fibrin involves a series of sequential proteolytic reactions, initiated by the ‘extrinsic’ and ‘Intrinsic’ Pathway of coagulation. As hereditary
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Intrinsic Pathway of Coagulation and Arterial Thrombosis
Arteriosclerosis thrombosis and vascular biology, 2007Co-Authors: David Gailani, Thomas RennéAbstract:Formation of a fibrin clot is mediated by a group of tightly regulated plasma proteases and cofactors. While this system is essential for minimizing blood loss from an injured blood vessel (hemostasis), it also contributes to pathologic fibrin formation and platelet activation that may occlude vessels (thrombosis). Many antithrombotic drugs target key elements of the plasma coagulation mechanism such as thrombin and factor Xa, based on the premise that plasma elements contributing to thrombosis are primarily those involved in hemostasis. Recent studies with genetically altered mice raise questions about this paradigm. Deficiencies of the Intrinsic Pathway proteases factor XII and factor XI are not associated with abnormal hemostasis in mice, but impair formation of occlusive thrombi in arterial injury models, indicating that Pathways not essential for hemostasis participate in arterial thrombosis. If factor XII or factor XI make similar contributions to thrombosis in humans, these proteases could be ideal targets for drugs to treat or prevent thromboembolic disease with minimal risk of therapy-associated bleeding.
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The Intrinsic Pathway of coagulation: a target for treating thromboembolic disease?
Journal of thrombosis and haemostasis : JTH, 2007Co-Authors: David Gailani, Thomas RennéAbstract:Summary. The classic Intrinsic Pathway of coagulation is triggered by contact activation of the plasma protease factor (F)XII, followed by sequential proteolytic activation of FX1 and FIX. While a key mechanism for initiating coagulation in some clinically useful in vitro assays, the absence of abnormal bleeding associated with congenital FXII deficiency indicates that the Intrinsic Pathway is not important for normal blood coagulation in vivo. However, recent work with mice lacking FXII or FXI suggest that these proteases make important contributions to formation of pathologic intravascular thrombi. In models of arterial injury, FXII or FXI null mice are protected from formation of platelet rich occlusive thrombi to a degree similar to that seen in FIX deficient mice (a model for the severe bleeding disorder hemophilia B) or to wild type mice treated with high dose heparin. FXII or FXI deficiency does not appear to prevent the initiation of thrombus formation in these models, but instead causes significant thrombus instability that prevents occlusion of the vessel. These findings raise the possibility that a Pathway similar or identical to the Intrinsic Pathway may operate in vivo under some circumstances. Furthermore, the disproportionate importance of FXII and FXI to occlusive thrombus formation compared to normal hemostasis makes these proteases attractive candidates for therapeutic inhibitors to treat or prevent thromboembolic disorders.
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targeting coagulation factor xii provides protection from pathological thrombosis in cerebral ischemia without interfering with hemostasis
Journal of Experimental Medicine, 2006Co-Authors: Christoph Kleinschnitz, David Gailani, Bernhard Nieswandt, Guido Stoll, Martin Bendszus, Kai Schuh, H U Pauer, Peter Burfeind, Christoph Renne, Thomas RennéAbstract:Formation of fibrin is critical for limiting blood loss at a site of blood vessel injury (hemostasis), but may also contribute to vascular thrombosis. Hereditary deficiency of factor XII (FXII), the protease that triggers the Intrinsic Pathway of coagulation in vitro, is not associated with spontaneous or excessive injury-related bleeding, indicating FXII is not required for hemostasis. We demonstrate that deficiency or inhibition of FXII protects mice from ischemic brain injury. After transient middle cerebral artery occlusion, the volume of infarcted brain in FXII-deficient and FXII inhibitor–treated mice was substantially less than in wild-type controls, without an increase in infarct-associated hemorrhage. Targeting FXII reduced fibrin formation in ischemic vessels, and reconstitution of FXII-deficient mice with human FXII restored fibrin deposition. Mice deficient in the FXII substrate factor XI were similarly protected from vessel-occluding fibrin formation, suggesting that FXII contributes to pathologic clotting through the Intrinsic Pathway. These data demonstrate that some processes involved in pathologic thrombus formation are distinct from those required for normal hemostasis. As FXII appears to be instrumental in pathologic fibrin formation but dispensable for hemostasis, FXII inhibition may offer a selective and safe strategy for preventing stroke and other thromboembolic diseases.
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the Intrinsic Pathway of coagulation is essential for thrombus stability in mice
Blood Cells Molecules and Diseases, 2006Co-Authors: Thomas Renné, Bernhard Nieswandt, David GailaniAbstract:Blood coagulation is a highly regulated process involving interactions between platelets, plasma coagulation factors, and the vessel wall. During coagulation in vivo, fibrin formation is thought to be initiated when plasma factor VIIa forms a complex with the membrane protein tissue factor. Coagulation factor XII (FXII, Hageman factor) is required for some in vitro coagulation systems; however, FXII deficiency is not associated with hemorrhage, leading to the conclusion that it is not necessary for hemostasis. We generated FXII-deficient mice to study the contributions of FXII to thrombosis and hemostasis in arterial injury models and in models of acute arterial occlusion. FXII-deficient mice do not experience excessive injury-related bleeding; however, intravital fluorescence microscopy and blood flow measurements in three separate arterial beds revealed a severe defect in formation and stabilization of platelet-rich occlusive thrombi induced by different methods of injuries. Similar findings were observed for mice deficient in factor XI, a substrate of activated FXII. Infusion of human FXII into FXII null mice restored thrombus formation. These findings demonstrate that FXII-mediated fibrin formation is crucial for pathological arterial thrombosis but not for hemostasis and suggest that FXII could be an ideal target for safe anticoagulation.
Bruce A. Sullenger - One of the best experts on this subject based on the ideXlab platform.
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a kallikrein targeting rna aptamer inhibits the Intrinsic Pathway of coagulation and reduces bradykinin release
Journal of Thrombosis and Haemostasis, 2017Co-Authors: K Steen A Burrell, Juliana M. Layzer, Bruce A. SullengerAbstract:SummaryBackground Plasma kallikrein is a serine protease that plays an integral role in many biological processes including coagulation, inflammation, and fibrinolysis. The main function of kallikrein in coagulation is the amplification of activated factor XIIa (FXIIa) generation which ultimately leads to thrombin generation and fibrin clot formation. Kallikrein is generated by FXIIa-mediated cleavage of the zymogen, prekallikrein which is usually complexed with the nonenzymatic cofactor, high-molecular-weight kininogen (HK). HK also serves as a substrate for kallikrein to generate the pro-inflammatory peptide, bradykinin (BK). Interestingly, prekallikrein-deficient mice are protected from thrombotic events while retaining normal hemostatic capacity. Therefore, therapeutically targeting kallikrein may provide a safer alternative to traditional anticoagulants with anti-inflammatory benefits. Objectives We sought to isolate and characterize an RNA aptamer that binds and inhibits plasma kallikrein and to elucidate its mechanism of action. Methods and Results Using convergent Systematic Evolution of Ligands by Exponential Enrichment (SELEX), we isolated an RNA aptamer that targets kallikrein. This aptamer, Kall1-T4, specifically binds to both prekallikrein and kallikrein with similar subnanomolar binding affinities and dose-dependently prolongs fibrin clot formation in an aPTT coagulation assay. In a purified in vitro system, Kall1-T4 inhibits the reciprocal activation of prekallikrein and FXII primarily by reducing the rate of FXIIa-mediated prekallikrein activation. Additionally, Kall1-T4 significantly reduces kallikrein-mediated HK cleavage and subsequent BK release. Conclusions We have isolated a specific and potent inhibitor of prekallikrein/kallikrein activity that serves as a powerful tool for further elucidating the role of kallikrein in thrombosis and inflammation. This article is protected by copyright. All rights reserved.
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Inhibiting the Intrinsic Pathway of coagulation with a factor XII-targeting RNA aptamer.
Journal of thrombosis and haemostasis : JTH, 2013Co-Authors: R. S. Woodruff, Juliana M. Layzer, Martin L. Ogletree, Bruce A. SullengerAbstract:Summary Background Exposure of the plasma protein factor XII (FXII) to an anionic surface generates activated FXII that not only triggers the Intrinsic Pathway of blood coagulation through the activation of FXI but also mediates various vascular responses through activation of the plasma contact system. While deficiencies of FXII are not associated with excessive bleeding, thrombosis models in factor-deficient animals have suggested that this protein contributes to stable thrombus formation. Therefore, FXII has emerged as an attractive therapeutic target to treat or prevent pathological thrombosis formation without increasing the risk for hemorrhage. Objectives Using an in vitro directed evolution and chemical biology approach, we sought to isolate a nuclease-resistant RNA aptamer that binds specifically to FXII and directly inhibits FXII coagulant function. Methods and Results We describe the isolation and characterization of a high-affinity RNA aptamer targeting FXII/activated FXII (FXIIa) that dose dependently prolongs fibrin clot formation and thrombin generation in clinical coagulation assays. This aptamer functions as a potent anticoagulant by inhibiting the autoactivation of FXII, as well as inhibiting Intrinsic Pathway activation (FXI activation). However, the aptamer does not affect the FXIIa-mediated activation of the proinflammatory kallikrein-kinin system (plasma kallikrein activation). Conclusions We have generated a specific and potent FXII/FXIIa aptamer anticoagulant that offers targeted inhibition of discrete macromolecular interactions involved in the activation of the Intrinsic Pathway of blood coagulation.
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antithrombotic therapy in acute coronary syndrome how far up the coagulation cascade will we go
Current Cardiology Reports, 2010Co-Authors: Becky Woodruff, Bruce A. Sullenger, Richard C BeckerAbstract:The contribution of thrombosis to the natural history and clinical expression of advanced atherosclerotic coronary artery disease is well established. Less well understood is the biochemical and pathobiological distinction between normal hemostasis and thrombosis as the proximate cause of acute coronary syndrome. In this article, we summarize an evolving area of interest within the field of antithrombotic therapy—the contact system and the Intrinsic Pathway of coagulation, focusing our discussion on factors XI, XII, and IX to include their biochemical properties, relationship to arterial thrombosis phenotypes, and rational for future investigation of targeted pharmacotherapy.
