Lufaxin, a novel factor Xa inhibitor from the salivary gland of the sand fly Lutzomyia longipalpis blocks protease-activated receptor 2 activation and inhibits inflammation and thrombosis in vivo
Autor: | Michalis Kotsyfakis, Teresa C.F. Assumpção, Dana C. Gilmore, Angélica Dutra-Oliveira, José M. C. Ribeiro, Jesus G. Valenzuela, Clarissa Teixeira, Ivo M.B. Francischetti, Robson Q. Monteiro, Nicolas Collin, Anderson Sá-Nunes, Daniella M. Mizurini |
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Jazyk: | angličtina |
Rok vydání: | 2012 |
Předmět: |
Time Factors
medicine.drug_mechanism_of_action Anti-Inflammatory Agents Ferric Compounds Salivary Glands Mice Thromboplastin Cloning Molecular Protease-activated receptor 2 Salivary gland medicine.diagnostic_test Recombinant Proteins medicine.anatomical_structure Coagulation Biochemistry Factor Xa Insect Proteins Female Partial Thromboplastin Time Cardiology and Cardiovascular Medicine Protein Binding Partial thromboplastin time Molecular Sequence Data Factor Xa Inhibitor Calorimetry Biology Article Chlorides Fibrinolytic Agents Prothrombinase Cell Line Tumor medicine Animals Humans Receptor PAR-2 Amino Acid Sequence Blood Coagulation Inflammation Dose-Response Relationship Drug Thrombosis Surface Plasmon Resonance Rats Mice Inbred C57BL Molecular Weight Disease Models Animal HEK293 Cells Prothrombin Time Anti-Inflammatory Agents/chemistry Anti-Inflammatory Agents/isolation & purification Blood Coagulation/drug effects Factor Xa/antagonists & inhibitors Factor Xa/metabolism Fibrinolytic Agents/chemistry Fibrinolytic Agents/isolation & purification Inflammation/blood Inflammation/metabolism Insect Proteins/chemistry Insect Proteins/isolation & purification Psychodidae/chemistry Receptor PAR-2/antagonists & inhibitors Receptor PAR-2/metabolism Recombinant Proteins/antagonists & inhibitors Recombinant Proteins/metabolism Salivary Glands/chemistry Thromboplastin/antagonists & inhibitors Thromboplastin/metabolism Thrombosis/blood Thrombosis/chemically induced Psychodidae Fibrinolytic agent Factor Xa Inhibitors |
Zdroj: | Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 32, no. 9, pp. 2185-2198 |
Popis: | Objective— Blood-sucking arthropods’ salivary glands contain a remarkable diversity of antihemostatics. The aim of the present study was to identify the unique salivary anticoagulant of the sand fly Lutzomyia longipalpis , which remained elusive for decades. Methods and Results— Several L. longipalpis salivary proteins were expressed in human embryonic kidney 293 cells and screened for inhibition of blood coagulation. A novel 32.4-kDa molecule, named Lufaxin, was identified as a slow, tight, noncompetitive, and reversible inhibitor of factor Xa (FXa). Notably, Lufaxin’s primary sequence does not share similarity to any physiological or salivary inhibitors of coagulation reported to date. Lufaxin is specific for FXa and does not interact with FX, Dansyl-Glu-Gly-Arg-FXa, or 15 other enzymes. In addition, Lufaxin blocks prothrombinase and increases both prothrombin time and activated partial thromboplastin time. Surface plasmon resonance experiments revealed that FXa binds Lufaxin with an equilibrium constant ≈3 nM, and isothermal titration calorimetry determined a stoichiometry of 1:1. Lufaxin also prevents protease-activated receptor 2 activation by FXa in the MDA-MB-231 cell line and abrogates edema formation triggered by injection of FXa in the paw of mice. Moreover, Lufaxin prevents FeCl 3 -induced carotid artery thrombus formation and prolongs activated partial thromboplastin time ex vivo, implying that it works as an anticoagulant in vivo. Finally, salivary gland of sand flies was found to inhibit FXa and to interact with the enzyme. Conclusion— Lufaxin belongs to a novel family of slow-tight FXa inhibitors, which display antithrombotic and anti-inflammatory activities. It is a useful tool to understand FXa structural features and its role in prohemostatic and proinflammatory events. |
Databáze: | OpenAIRE |
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