The Assessment of α1Proteinase 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 (54000 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 proteolysed forms and the other which was more cationic and did not inhibit porcine pancreatic elastase. Finally, thirteen cut 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.