title: Relative roles of elastase and reactive oxygen species in the degradation of human glomerular basement membrane by intact human neutrophils.
abstract: Glomerular basement membrane (GBM) damage and proteinuria occurring during the early phase of acute glomerulonephritis are often neutrophil (PMN) dependent. The present study sought to identify the potential roles of PMN derived elastase and reactive oxygen species (ROS) in the pathogenesis of glomerular basement damage in an homologous in vitro model of anti-GBM nephritis using intact PMN. Human PMN (5 x 10(6)), incubated with human GBM (0.5 mg) pretreated with human anti-GBM IgG, degraded 10.3 +/- 1.1% of the GBM type IV collagen in six hours (8 micrograms/hr), and underwent a two-hour respiratory burst. The same number of sonically disrupted PMN solubulized 22.4 +/- 5.1% of GBM under the same incubation conditions. The inclusion of the elastase inhibitors alpha 1 proteinase inhibitor (alpha 1Pi), and a smaller highly-specific synthetic compound (L658,758), reduced degradation by PMN homogenates by 84.8% and 85.7%, respectively, whereas they were only able to inhibit intact PMN mediated degradation by a maximum of 49.2% and 50.9%, respectively. The inclusion of EDTA (10 mM), an inhibitor of metalloproteinases, reduced GBM degradation by APMA activated and disrupted PMN by only 7.5%. Incubation of PMN with diphenylene iodonium (DPI) abolished PMN reactive oxygen species generation by > 95% but preserved elastase release. This compound did not directly affect GBM degradation. It did, however, abolish the inhibitory effect of ROS on alpha 1Pi activity and thus indirectly reduced GBM damage by up to 20%.
authors.affiliation: Institute of Nephrology, Royal Infirmary, Cardiff, Wales, United Kingdom.
authors.forename: K L
authors.initials: KL
authors.lastname: Donovan
authors.forename: M
authors.initials: M
authors.lastname: Davies
authors.forename: G A
authors.initials: GA
authors.lastname: Coles
authors.forename: J D
authors.initials: JD
authors.lastname: Williams
mesh_terms: D001485:Basement Membrane / Q000378:metabolism*; D002395:Catecholamines / Q000494:pharmacology; D002511:Cephalosporins; D003094:Collagen / Q000378:metabolism; D005921:Glomerulonephritis / Q000209:etiology; D006801:Humans; D048288:Imidazolines; D007074:Immunoglobulin G / Q000378:metabolism; D007678:Kidney Glomerulus / Q000378:metabolism*; D009504:Neutrophils / Q000187:drug effects / Q000502:physiology*; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors / Q000502:physiology*; D011759:Pyrrolidines / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D016897:Respiratory Burst; D000515:alpha 1-Antitrypsin / Q000494:pharmacology
chemical_list: D002395:Catecholamines; D002511:Cephalosporins; D048288:Imidazolines; D007074:Immunoglobulin G; D011759:Pyrrolidines; D017382:Reactive Oxygen Species; D000515:alpha 1-Antitrypsin; C065332:L 658758; D003094:Collagen; C011775:(3,4-dihydroxyphenylamino)-2-imidazoline; D010196:Pancreatic Elastase