title: Activation of Mac-1 (CD11b/CD18)-bound factor X by released cathepsin G defines an alternative pathway of leucocyte initiation of coagulation.
abstract: Leucocyte initiation of coagulation preserves the haemostatic balance and may aberrantly contribute to vascular injury. In addition to the extrinsic activation mediated by tissue factor: factor VIIa, monocytes express an alternative procoagulant response after binding of the zymogen factor X to the integrin Mac-1 (CD11b/CD18). Here, factor X-activating activity was found in purified monocyte granules, and coincided with size-chromatographed fractions containing cathepsin G. In contrast, elastase-containing granule fractions did not activate factor X. In the presence of Ca2+ ions, purified cathepsin G, but not elastase, cleaved factor X to a approximately 54 kDa catalytically active derivative, structurally indistinguishable from the procoagulant product generated on monocytes after binding to Mac-1. Factor X activation by purified cathepsin G involved limited proteolysis of a novel Leu177-Leu178 peptide bond in the zymogen's activation peptide. Cathepsin G activation of factor X was completely inhibited by alpha 1 antichymotrypsin, or soybean trypsin inhibitor, or by a neutralizing antiserum to cathepsin G, while eglin, or an anti-elastase antibody, were ineffective. Affinity chromatography on active-site-dependent inhibitors Glu-Gly-Arg-chloromethyl ketone or benzamidine completely abolished factor Xa activity generated by cathepsin G. Cathepsin G was not constitutively detected on the monocyte surface by flow cytometry. However, inflammatory stimuli, including formyl peptide or phorbol ester, or Mac-1 engagement with its ligands fibrinogen, factor X or serum-opsonized zymosan, triggered monocyte degranulation and cathepsin G activation of factor X. These findings demonstrate that monocytes can alternatively initiate coagulation in a sequential three-step cascade, including (i) binding of factor X to Mac-1, (ii) discharge of azurophil granules, and (iii) limited proteolytic activation of membranebound factor X by cathepsin G. By rapidly forming thrombin and factor Xa in a protected membrane microenvironment, this pathway may contribute a "priming' signal for clotting, anticoagulation and vascular cell signal transduction, in vivo.
authors.affiliation: Boyer Center for Molecular Medicine, Department of Pathology, Yale University School of Medicine, New Haven, CT 06536, USA.
authors.forename: J
authors.initials: J
authors.lastname: Plescia
authors.forename: D C
authors.initials: DC
authors.lastname: Altieri
mesh_terms: D001777:Blood Coagulation; D056649:Cathepsin G; D002403:Cathepsins / Q000378:metabolism*; D002458:Cell Fractionation; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003594:Cytoplasmic Granules / Q000378:metabolism*; D010450:Endopeptidases / Q000378:metabolism; D005170:Factor X / Q000378:metabolism*; D015951:Factor Xa / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D007962:Leukocytes / Q000187:drug effects / Q000502:physiology*; D016177:Macrophage-1 Antigen / Q000502:physiology*; D009000:Monocytes; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D010196:Pancreatic Elastase / Q000378:metabolism; D012697:Serine Endopeptidases; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D016177:Macrophage-1 Antigen; D009240:N-Formylmethionine Leucyl-Phenylalanine; D005170:Factor X; D002403:Cathepsins; D010450:Endopeptidases; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G; D010196:Pancreatic Elastase; D015951:Factor Xa; D013755:Tetradecanoylphorbol Acetate