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Antibody-mediated targeting of the urokinase-type plasminogen activator proteolytic function neutralizes fibrinolysis in vivo

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DOI

  1. Did evolution create a flexible ligand-binding cavity in the urokinase receptor through deletion of a plesiotypic disulfide bond?

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  2. Site-specific O-glycosylation of members of the low-density lipoprotein receptor superfamily enhances ligand interactions

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  1. Immune regulation by fibroblasts in tissue injury depends on uPARAP-mediated uptake of collectins

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  2. uPARAP/Endo180 receptor is a gatekeeper of VEGFR-2/VEGFR-3 heterodimerisation during pathological lymphangiogenesis

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  3. Defective TAFI activation in hemophilia A mice is a major contributor to joint bleeding

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  4. Antibody-Mediated Neutralization of uPA Proteolytic Function Reduces Disease Progression in Mouse Arthritis Models

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Urokinase-type plasminogen activator (uPA) plays a central role in tissue remodeling processes. Most of our understanding of the role of uPA in vivo is derived from studies using gene-targeted uPA-deficient mice. To enable in vivo studies on the specific interference with uPA functionality in mouse models, we have now developed murine monoclonal antibodies (mAbs) directed against murine uPA by immunization of uPA-deficient mice with the recombinant protein. Guided by enzyme-linked immunosorbent assay, Western blotting, surface plasmon resonance, and enzyme kinetic analyses, we have selected two highly potent and inhibitory anti-uPA mAbs (mU1 and mU3). Both mAbs recognize epitopes located on the B-chain of uPA that encompasses the catalytic site. In enzyme activity assays in vitro, mU1 blocked uPA-catalyzed plasminogen activation as well as plasmin-mediated pro-uPA activation, whereas mU3 only was directed against the first of these reactions. We additionally provide evidence that mU1, but not mU3, successfully targets uPA-dependent processes in vivo. Hence, systemic administration of mU1 (i) rescued mice treated with a uPA-activable anthrax protoxin and (ii) impaired uPA-mediated hepatic fibrinolysis in tissue-type plasminogen activator (tPA)-deficient mice, resulting in a phenotype mimicking that of uPA;tPA double deficient mice. Importantly, this is the first report demonstrating specific antagonist-directed targeting of mouse uPA at the enzyme activity level in a normal physiological process in vivo.
Original languageEnglish
JournalThe journal of biological chemistry
Volume283
Issue number47
Pages (from-to)32506-15
Number of pages10
ISSN0021-9258
DOIs
Publication statusPublished - 21 Nov 2008

    Research areas

  • Animals, Antibodies, Antibodies, Monoclonal, Enzyme-Linked Immunosorbent Assay, Female, Fibrinolysis, Kinetics, Liver, Mice, Mice, Inbred C57BL, Models, Biological, Protein Binding, Recombinant Proteins, Surface Plasmon Resonance, Urokinase-Type Plasminogen Activator

ID: 39988595