title: An in vitro model for polymorphonuclear-leukocyte-induced injury to an extracellular matrix. Relative contribution of oxidants and elastase to fibronectin release from amnionic membranes.
abstract: Alteration of the extracellular matrix by inflammatory cells is believed to be important in both lung injury and the subsequent restoration of lung architecture. Here we describe the results of the interaction between an acellular human amnionic membrane model and stimulated human polymorphonuclear neutrophils (PMN) in vitro. Polymorphonuclear neutrophil suspensions were placed on one surface of the amnion, and either the chemotactic peptide FMLP or the cell membrane activator phorbol myristate acetate (PMA) was placed on the opposite side of the amnion. Stroma and basement membrane sides of the amnion were separately exposed to the PMN. The PMN suspension was removed and centrifuged, and the supernatant was assayed for superoxide anion (O2-.) and for elastase activity. Injury to the acellular amnion was evaluated by transmission electron microscopy and by measurement of fibronectin (FN) released from the membrane matrix. Although both stimulants cause a concentration-dependent release of O2-., only PMA stimulated elastase release. These effects were similar when either the stroma or the basement membrane side was exposed to PMN. PMA-stimulated cells and supernatants from PMA-stimulated cells caused solubilization of membrane at different incubation times. Electron microscopy confirmed the disruption of the basement membrane of the amnion by PMA-stimulated PMN. Oxidant scavengers (SOD and catalase) did not prevent matrix degradation, and elastase inhibition by a specific chloromethylketone inhibitor diminished FN release on both sides of the amnion by activated PMN supernatants, but only on the basement membrane side by intact PMN. We conclude that in this model, elastase rather than oxygen radicals solubilizes FN from the matrix.
mesh_terms: D000650:Amnion / Q000276:immunology* / Q000378:metabolism; D001485:Basement Membrane / Q000187:drug effects / Q000378:metabolism; D002374:Catalase / Q000378:metabolism; D005109:Extracellular Matrix / Q000276:immunology*; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008854:Microscopy, Electron; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000276:immunology*; D010196:Pancreatic Elastase / Q000378:metabolism; D013268:Stimulation, Chemical; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D005353:Fibronectins; D013481:Superoxides; D009240:N-Formylmethionine Leucyl-Phenylalanine; D002374:Catalase; D013482:Superoxide Dismutase; D010196:Pancreatic Elastase; D013755:Tetradecanoylphorbol Acetate