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Frontiers in Bioscience (Landmark... Jun 2011Knowledge about species compatibility is crucial for proper interpretation of data from in vivo experiments with human proteins in pharmacological models and of data... (Comparative Study)
Comparative Study Review
Knowledge about species compatibility is crucial for proper interpretation of data from in vivo experiments with human proteins in pharmacological models and of data from cross-species in vitro experiments. Information about the cross-species compatibility of tissue factor (TF) and coagulation factor (F) VII (FVII) has accumulated since the early history of coagulation research. Many observations were connected to the introduction and development of the prothrombin time (PT) assay where fibrin clot formation was observed when tissue extracts of different origins were added to recalcified human or non-human plasmas. Studies on cross-species TF-FVIIa compatibility entered into a new area with the cloning and recombinant expression of TF and FVII from a number of species as well as with the possibility of specific amino acid substitution. TF and/or FVIIa from cattle, dog, rabbit, mouse, rat and zebrafish have been purified and characterized in varying detail. In addition to adding knowledge about the species-specific TF-FVIIa interactions, cross-species studies often reveal information which adds to the general view of the structural and functional properties of the human TF-FVIIa complex. This review briefly outlines the features of human TF and FVIIa, their intermolecular interactions, and the biological effects of TF-FVIIa complex formation and compares this information to findings obtained in studies addressing TF or FVIIa of non-human origin. By examples we point to difficulties which may arise from the transcendence across species borders and how some cross-species data have advanced our understanding of the structure and function of the human TF-FVIIa complex.
Topics: Amino Acid Sequence; Animals; Blood Coagulation; Cattle; Dogs; Factor VIIa; Humans; Mice; Models, Molecular; Molecular Sequence Data; Protein Interaction Domains and Motifs; Rabbits; Rats; Sequence Homology, Amino Acid; Species Specificity; Thromboplastin; Zebrafish
PubMed: 21622229
DOI: 10.2741/3906 -
Biological Chemistry Jul 2013Tissue factor (TF), an initiator of blood coagulation in vivo, is expressed in a variety of cells. Sufficient natural TF has been isolated to clone and express... (Comparative Study)
Comparative Study Review
Tissue factor (TF), an initiator of blood coagulation in vivo, is expressed in a variety of cells. Sufficient natural TF has been isolated to clone and express recombinant proteins ranging from full-length TF to its extracellular domain. Because of the limited availability of natural TF, recombinant proteins have been used as surrogates. Despite the differences in their post-translational modifications, it has been accepted that membrane-anchored recombinant TFs are quite similar to the natural TF. Recent studies, however, have shown that post-translational modifications play an important role in TF-triggered thrombin generation.
Topics: Blood Coagulation; Factor VIIa; Humans; Protein Processing, Post-Translational; Recombinant Proteins; Thrombin; Thromboplastin
PubMed: 23412875
DOI: 10.1515/hsz-2012-0350 -
Journal of Thrombosis and Haemostasis :... Aug 2019Blood coagulation factor Va serves an indispensable role in hemostasis as cofactor for the serine protease factor Xa. In the presence of an anionic phospholipid membrane... (Review)
Review
Blood coagulation factor Va serves an indispensable role in hemostasis as cofactor for the serine protease factor Xa. In the presence of an anionic phospholipid membrane and calcium ions, factors Va and Xa assemble into the prothrombinase complex. Following formation of the ternary complex with the macromolecular zymogen substrate prothrombin, the latter is rapidly converted into thrombin, the key regulatory enzyme of coagulation. Over the years, multiple binding sites have been identified in factor Va that play a role in the interaction of the cofactor with factor Xa, prothrombin, or the anionic phospholipid membrane surface. In this review, an overview of the currently available information on these interactive sites in factor Va is provided, and data from biochemical approaches and 3D structural protein complex models are discussed. The structural models have been generated in recent years and provide novel insights into the molecular requirements for assembly of both the prothrombinase and the ternary prothrombinase-prothrombin complexes. Integrated knowledge of functionally important regions in factor Va will allow for a better understanding of factor Va cofactor activity.
Topics: Binding Sites; Blood Coagulation; Cell Membrane; Factor Va; Factor Xa; Humans; Models, Molecular; Phospholipids; Protein Binding; Protein Interaction Domains and Motifs; Prothrombin; Structure-Activity Relationship; Thromboplastin
PubMed: 31102425
DOI: 10.1111/jth.14487 -
The Journal of Biological Chemistry Apr 2013Trypsin-like proteases are synthesized as inactive zymogens and convert to the mature form upon activation by specific enzymes, often assisted by cofactors. Central to...
Trypsin-like proteases are synthesized as inactive zymogens and convert to the mature form upon activation by specific enzymes, often assisted by cofactors. Central to this paradigm is that the zymogen does not convert spontaneously to the mature enzyme, which in turn does not feed back to activate its zymogen form. In the blood, the zymogens prothrombin and prethrombin-2 require the prothrombinase complex to be converted to the mature protease thrombin, which is unable to activate prothrombin or prethrombin-2. Here, we show that replacement of key residues within the activation domain causes these zymogens to spontaneously convert to thrombin. The conversion is started by the zymogen itself, which is capable of binding ligands at the active site, and is abrogated by inactivation of the catalytic residue Ser-195. The product of autoactivation is functionally and structurally equivalent to wild-type thrombin. Zymogen autoactivation is explained by conformational selection, a basic property of the trypsin fold uncovered by structural and rapid kinetics studies. Both the zymogen and protease undergo a pre-existing equilibrium between active and inactive forms. The equilibrium regulates catalytic activity in the protease and has the potential to unleash activity in the zymogen to produce autoactivation. A new strategy emerges for the facile production of enzymes through zymogen autoactivation that is broadly applicable to trypsin-like proteases of biotechnological and clinical interest.
Topics: Amino Acid Substitution; Enzyme Activation; Humans; Mutation, Missense; Prothrombin; Thromboplastin
PubMed: 23467412
DOI: 10.1074/jbc.M113.451542 -
Annual Review of Physiology 2011Venous thromboembolism (VTE) is a leading cause of morbidity and mortality worldwide. However, the mechanisms by which clots are formed in the deep veins have not been... (Review)
Review
Venous thromboembolism (VTE) is a leading cause of morbidity and mortality worldwide. However, the mechanisms by which clots are formed in the deep veins have not been determined. Tissue factor (TF) is the primary initiator of the coagulation cascade and is essential for hemostasis. Under pathological conditions, TF is released into the circulation on small-membrane vesicles termed microparticles (MPs). Recent studies suggest that elevated levels of MP TF may trigger thrombosis. This review provides an overview of the role of TF in VTE.
Topics: Animals; Biomarkers; Blood Coagulation; Cell-Derived Microparticles; Disease Models, Animal; Humans; Mice; Neoplasms; Thromboplastin; Venous Thrombosis; Wounds and Injuries
PubMed: 20690821
DOI: 10.1146/annurev-physiol-042210-121137 -
Biorheology 2015Microfluidic devices create precisely controlled reactive blood flows and typically involve: (i) validated anticoagulation/pharmacology protocols, (ii) defined reactive... (Review)
Review
Microfluidic devices create precisely controlled reactive blood flows and typically involve: (i) validated anticoagulation/pharmacology protocols, (ii) defined reactive surfaces, (iii) defined flow-transport regimes, and (iv) optical imaging. An 8-channel device can be run at constant flow rate or constant pressure drop for blood perfusion over a patterned collagen, collagen/kaolin, or collagen/tissue factor (TF) to measure platelet, thrombin, and fibrin dynamics during clot growth. A membrane-flow device delivers a constant flux of platelet agonists or coagulation enzymes into flowing blood. A trifurcated device sheaths a central blood flow on both sides with buffer, an ideal approach for on-chip recalcification of citrated blood or drug delivery. A side-view device allows clotting on a porous collagen/TF plug at constant pressure differential across the developing clot. The core-shell architecture of clots made in mouse models can be replicated in this device using human blood. For pathological flows, a stenosis device achieves shear rates of >100,000 s(-1) to drive plasma von Willebrand factor (VWF) to form thick long fibers on collagen. Similarly, a micropost-impingement device creates extreme elongational and shear flows for VWF fiber formation without collagen. Overall, microfluidics are ideal for studies of clotting, bleeding, fibrin polymerization/fibrinolysis, cell/clot mechanics, adhesion, mechanobiology, and reaction-transport dynamics.
Topics: Animals; Blood Circulation; Collagen; Hemodynamics; Humans; Microfluidic Analytical Techniques; Miniaturization; Thromboplastin; von Willebrand Factor
PubMed: 26600269
DOI: 10.3233/BIR-15065 -
Circulation Feb 2006Tissue factor (TF), formerly known as thromboplastin, is the key initiator of the coagulation cascade; it binds factor VIIa resulting in activation of factor IX and... (Review)
Review
Tissue factor (TF), formerly known as thromboplastin, is the key initiator of the coagulation cascade; it binds factor VIIa resulting in activation of factor IX and factor X, ultimately leading to fibrin formation. TF expression and activity can be induced in endothelial cells, vascular smooth muscle cells, and monocytes by various stimuli such as cytokines, growth factors, and biogenic amines. These mediators act through diverse signal transduction mechanisms including MAP kinases, PI3-kinase, and protein kinase C. Cellular TF is present in three pools as surface, encrypted, and intracellular protein. TF can also be detected in the bloodstream, referred to as circulating or blood-borne TF. Elevated levels of TF are observed in patients with cardiovascular risk factors such as hypertension, diabetes, dyslipidemia, and smoking as well as in those with acute coronary syndromes. TF may indeed be involved in the pathogenesis of atherosclerosis by promoting thrombus formation; in addition, it can induce migration and proliferation of vascular smooth muscle cells. As a consequence, therapeutic strategies have been developed to specifically interfere with the action of TF such as antibodies against TF, site-inactivated factor VIIa, or recombinant TF pathway inhibitor. Inhibition of TF action appears to be an attractive target for the treatment of cardiovascular diseases.
Topics: Animals; Blood Coagulation; Cardiovascular Diseases; Endothelium, Vascular; Gene Expression Regulation; Humans; Thromboplastin
PubMed: 16461845
DOI: 10.1161/CIRCULATIONAHA.105.567297 -
International Journal of Molecular... Jan 2023Apolipoprotein CIII (ApoCIII) represents a key regulator of plasma lipid metabolism and a recognized risk factor for atherosclerosis and cardiovascular diseases. Beyond...
Apolipoprotein CIII (ApoCIII) represents a key regulator of plasma lipid metabolism and a recognized risk factor for atherosclerosis and cardiovascular diseases. Beyond the regulation of lipoprotein trafficking, ApoCIII is also involved in endothelial dysfunction and monocyte recruitment related to atherothrombosis. With tissue factor (TF) being the primary initiator of the blood coagulation cascade, we hypothesized that ApoCIII-treated monocytes could express it. Hence, human CD14-monocytes and autologous neutrophils were incubated with ApoCIII and sera from human subjects containing previously measured ApoCIII amounts. By RT-qPCR and ELISA, CD14-monocytes, but not neutrophils, were found to show increased mRNA expression and production of TNFα, IL-1β and IL-6 as well as TF mRNA once exposed to ultra-purified ApoCIII. By flow cytometry, CD14-monocytes were found to rapidly express TF on their cell surface membrane when incubated with either ApoCIII or sera with known concentrations of ApoCIII. Finally, preincubation with specific ApoCIII-neutralizing antibodies significantly reduced the ability of most sera with known concentrations of ApoCIII to upregulate TF protein, other than partially inhibiting cytokine release, in CD14-monocytes. In sum, herein we demonstrate that ApoCIII activates CD14-monocytes to express TF. The data identify a potential mechanism which links circulating apolipoproteins with inflammation and atherothrombosis-related processes underlying cardiovascular risk.
Topics: Humans; Apolipoprotein C-III; Apolipoproteins; Monocytes; RNA, Messenger; Thromboplastin
PubMed: 36768547
DOI: 10.3390/ijms24032223 -
Journal of Atherosclerosis and... 2005Hemostatic factors play a crucial role in generating thrombotic plugs at sites of vascular damage (atherothrombosis). However, whether hemostatic factors contribute... (Review)
Review
Hemostatic factors play a crucial role in generating thrombotic plugs at sites of vascular damage (atherothrombosis). However, whether hemostatic factors contribute directly or indirectly to the pathogenesis of atherosclerosis remains uncertain. Autopsy studies have revealed that intimal thickening represents the first stage of atherosclerosis and that lipid-rich plaque arises from such lesions. Several factors contribute to the start of intimal thickening. Platelets release several growth factors and bioactive agents that play a central role in development of not only thrombus but also of intimal thickening. We have been investigating which coagulation factors simultaneously, or subsequently with platelet aggregation, participate in thrombus formation. Tissue factor (TF) is an essential initiator of blood coagulation that is expressed in various stages of atherosclerotic lesions in humans and other animals. Factors including thrombin and fibrin, which are downstream of the coagulation cascade activated by TF, also contribute to atherosclerosis. TF is involved in cell migration, embryogenesis and angiogenesis. Thus TF, in addition to factors downstream of the coagulation cascade and the protease-activated receptor 2 activation system, would be a multifactorial regulator of atherogenesis.
Topics: Arteriosclerosis; Blood Coagulation; Fibrin; Humans; Receptor, PAR-2; Thrombin; Thromboplastin
PubMed: 15725689
DOI: 10.5551/jat.12.1 -
Thrombosis and Haemostasis May 2016Osteosarcoma is the most common primary malignant bone tumour. Patients often develop lung metastasis and have a poor prognosis despite extensive chemotherapy and...
Osteosarcoma is the most common primary malignant bone tumour. Patients often develop lung metastasis and have a poor prognosis despite extensive chemotherapy and surgical resections. Tissue Factor is associated with poor clinical outcome in a wide range of cancer types, and promotes angiogenesis and metastasis. The role of Tissue Factor in OS tumourigenesis is unknown. Fifty-three osteosarcoma pre-treatment biopsies and four osteosarcoma cell lines were evaluated for Tissue Factor expression, and a possible association with clinical parameters was investigated. Tissue Factor function was inhibited in an osteosarcoma cell line (143B) by shRNA knockdown or specific antibodies, and pro-tumourigenic gene expression, proliferation, matrigel invasion and transwell migration was examined. 143B cells were implanted in mice in the presence of Tissue Factor-blocking antibodies, and tumour volume, micro-vessel density and metastases in the lung were evaluated. Tissue Factor was highly expressed in 73.6 % of osteosarcoma biopsies, and expression associated significantly with disease-free survival. Tissue Factor was expressed in all four investigated cell lines. Tissue Factor was knocked down in 143B cells, which led to reduced expression of IL-8, CXCL-1, SNAIL and MMP2, but not MMP9. Tissue Factor knockdown or inhibition with antibodies reduced matrigel invasion. Tissue Factor antibodies limited 143B tumour growth in vivo, and resulted in decreased intra-tumoural micro-vessel density. Furthermore, lung metastasis from the primary tumour was significantly reduced. Thus, Tissue Factor expression in osteosarcoma reduces metastasis-free survival in patients, and increases pro-tumourigenic behaviour both in vitro and in vivo.
Topics: Adolescent; Animals; Bone Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Child; Disease-Free Survival; Female; Gene Expression; Gene Knockdown Techniques; Humans; Male; Mice, Inbred NOD; Mice, SCID; Neoplasm Invasiveness; Osteosarcoma; RNA, Messenger; RNA, Neoplasm; Thromboplastin; Young Adult
PubMed: 26763081
DOI: 10.1160/TH15-07-0541