title: The assessment of alpha 1 proteinase inhibitor form and function in lung lavage fluid from healthy subjects.
abstract: The form and function of alpha 1 proteinase inhibitor in lung lavage fluid from healthy smoking and non smoking individuals has been accurately assessed using critically appraised techniques. The present study demonstrated that it is possible to accurately assess alpha 1 PI function in unconcentrated lavage fluid but that sample collection, storage and subsequent processing may all affect the results. Absolute levels of alpha 1 PI were elevated in subjects who smoke and a substantial quantity of inactive protein was found in both smokers and non smokers. The proportion of inactive alpha 1 PI was similar for both groups, which by inference implies that normal smoking subjects do not have decreased protection by this inhibitor at the bronchoalveolar level. Physicochemical analysis of the alpha 1 PI in these normal subjects showed that it was different from alpha 1 PI previously reported from patients with established disease and this may have important implications regarding the pathogenesis of their condition. Western immunoblotting of bronchoalveolar lavage fluid (BALF) showed that all of the alpha 1 PI was present in the native molecular mass form (54,000 Da). Pre-incubation of samples with methionine sulphoxide peptide reductase restored alpha 1 PI function only by approximately 10% suggesting the presence of little reversibly oxidised alpha 1 PI in either group. Anion exchange HPLC of BALF revealed the presence of two alpha 1 PI species, one of which co-eluted with native, oxidised or proteolyzed forms and the other which was more cationic and did not inhibit porcine pancreatic elastase. Finally, thirteen out of sixteen BALF samples inhibited more neutrophil elastase than could be accounted for by the amounts of functional alpha 1 PI present, suggesting that the presence of other inhibitors is a feature of normal lavage fluids.
authors.affiliation: Lung Immunobiochemical Research Laboratory, General Hospital, Birmingham, United Kingdom.
authors.forename: S C
authors.initials: SC
authors.lastname: Afford
authors.forename: D
authors.initials: D
authors.lastname: Burnett
authors.forename: E J
authors.initials: EJ
authors.lastname: Campbell
authors.forename: J D
authors.initials: JD
authors.lastname: Cury
authors.forename: R A
authors.initials: RA
authors.lastname: Stockley
mesh_terms: D000818:Animals; D001798:Blood Proteins / Q000302:isolation & purification* / Q000494:pharmacology / Q000502:physiology; D015153:Blotting, Western; D006801:Humans; D008168:Lung / Q000502:physiology*; D009504:Neutrophils / Q000201:enzymology; D010179:Pancreas / Q000201:enzymology; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors / Q000097:blood; D011480:Protease Inhibitors / Q000302:isolation & purification*; D012016:Reference Values; D012907:Smoking / Q000503:physiopathology; D013552:Swine; D007507:Therapeutic Irrigation; D000515:alpha 1-Antitrypsin
chemical_list: D001798:Blood Proteins; D011480:Protease Inhibitors; D000515:alpha 1-Antitrypsin; D010196:Pancreatic Elastase