title: Protease-cleaved iron-transferrin augments oxidant-mediated endothelial cell injury via hydroxyl radical formation.
abstract: Previous work has shown that the Pseudomonas-derived protease, pseudomonas elastase (PAE), can modify transferrin to form iron complexes capable of catalyzing the formation of hydroxyl radical (.OH) from neutrophil (PMN)-derived superoxide (.O2-) and hydrogen peroxide (H2O2). As the lung is a major site of Pseudomonas infection, the ability of these iron chelates to augment oxidant-mediated pulmonary artery endothelial cell injury via release of 51Cr from prelabeled cells was examined. Diferrictransferrin previously cleaved with PAE significantly enhanced porcine pulmonary artery endothelial cell monolayer injury from 2.3-6.3 to 15.8-17.0% of maximum, resulting from exposure to H2O2, products of the xanthine/xanthine oxidase reaction, or PMA-stimulated PMNs. Iron associated with transferrin appeared to be responsible for cell injury. Spin trapping and the formation of thiobarbituric acid-reactive 2-deoxyribose oxidation products demonstrated the production of .OH in this system. The addition of catalase, dimethyl thiourea, and the hydrophobic spin trap, alpha-phenyl-n-terbutyl-nitrone, offered significant protection from injury (27.8-58.2%). Since sites of Pseudomonas infection contain other proteases, the ability of porcine pancreatic elastase and trypsin to substitute for PAE was examined. Results were similar to those observed with PAE. We conclude .OH formation resulting from protease alteration of transferrin may serve as a mechanism of tissue injury at sites of bacterial infection and other processes characterized by increased proteolytic activity.
authors.affiliation: Research Service, Veterans Affairs Medical Center, Iowa City, Iowa 52246, USA.
authors.forename: R A
authors.initials: RA
authors.lastname: Miller
authors.forename: B E
authors.initials: BE
authors.lastname: Britigan
mesh_terms: D000818:Animals; D001426:Bacterial Proteins; D002478:Cells, Cultured; D004578:Electron Spin Resonance Spectroscopy; D004727:Endothelium / Q000187:drug effects*; D017665:Hydroxyl Radical / Q000633:toxicity*; D066298:In Vitro Techniques; D007501:Iron / Q000737:chemistry*; D008666:Metalloendopeptidases / Q000378:metabolism*; D009504:Neutrophils / Q000378:metabolism; D016877:Oxidants; D010084:Oxidation-Reduction; D011550:Pseudomonas aeruginosa / Q000201:enzymology; D011651:Pulmonary Artery / Q000187:drug effects*; D013552:Swine; D014168:Transferrin / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D016877:Oxidants; D014168:Transferrin; D017665:Hydroxyl Radical; D007501:Iron; D008666:Metalloendopeptidases; C043859:pseudolysin, Pseudomonas aeruginosa