There are five specific aims: 1) We will attempt to detect and localize oxidized alpha 1-proteinase inhibitor in lung biopsies obtained from smoking and sham-smoking baboons. For this purpose, a specific monoclonal antibody that recognizes only the oxidized inhibitor will be used as the primary staining reagent, followed by established secondary-antibody immunohistochemical methods (peroxidase-antiperoxidase, avidin-biotin). This objective is important to human health because oxidation of alpha 1- proteinase inhibitor in the lungs by cigarette smoking has been implicated as a major pathogenetic factor in pulmonary emphysema, but remains unproven. 2) We will explore the role of a newly discovered mast cell elastase in the pathogenesis of smokers' emphysema in three ways: (a) by monitoring release of the enzyme (using ELISA-techniques) from purified human lung mast cells incubated with aqueous solutions of tobacco smoke and smoke fractions; (b) by measuring levels of histamine (a marker of mast cell secretory response) in bronchoalveolar lavage fluids of human smokers (before and after smoking) and of rats following acute exposure to inhaled cigarette smoke; and (c) by attempting to induce experimental emphysema in rats and dogs using repeated intrapulmonary instillations of mast cell secretogogues. 3) We will attempt to develop new inhibitors of human neutrophil elastase (a protease suspected on being a pathogenetic factor in pulmonary emphysema) based on our recent characterization of a specific neutrophil elastase inhibitor in pneumococci. 4) We will test an oxidation-resistant, genetically-engineered, mutant (Met 358Val) alpha 1-proteinase inhibitor for its resistance to inactivation in vivo in an established animal model of acute cigarette smoke exposure. 5) Finally, we will measure levels o f the neutrophil-monocyte marker antigen (L1) in blood and airspace secretions of septic and traumatized patients at risk for adult respiratory distress syndrome (ARDS) and in those with frank ARDS. Levels of the L1 protein may prove to have predictive value of ARDS in high-risk subjects.
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