MAST CELL CYSTEINE PROTEASES IN LUNG INFLAMMATION

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Mast cell cysteine, metalloprotease, and serine proteases may play a significant role in the pathogenesis of inflammatory lung diseases. The long-term objective of this proposal is to identify the cysteine proteases present in mast cells, and to characterize their roles in mast cell dependent inflammation. We hypothesize that tissue mast cells are a major source of dipeptidyl peptidase I (DPPI) and a newly identified 60-kDa cysteine protease and that these proteases participate in inflammation by hydrolyzing extracellular proteins. Specific aims are : 1. To explore structure-function relationships of DPPI; 2. To identify cells expressing DPPI in normal and inflamed lung tissues; 3. To characterize substrate specificity of DPPI, and 4. To characterize a nove1 60 kDa cysteine protease found in mast cells. To achieve these goals: 1. Proteolytic processing of proDPPI will be studied in recombinantly expressed DPPI. 2. DPPI's expressed with mutated propeptides will be assayed for their dipeptidyl peptidase and endoprotease activity, 3. Expression of DPPI by specific cell types in normal and inflamed dog lungs will be determined by immunohistochemistry and in situ hybridization. 4. DPPI will be studied for its ability to hydrolyze extracellular peptides and proteins. 5. Mast cell functional characteristics and proteolytic activities will be studied in DPPI knockout mice. 6. A novel mast cell cysteine protease will be purified and characterized. Dr. Wolters has completed training in internal medicine and pulmonary and critical care medicine. He has demonstrated a firm commitment to a career in academic medicine and an interest in studying the pathophysiology of lung diseases. The research training plan includes: an intensive laboratory experience, didactic coursework, weekly seminars, and journal clubs. The research training will be overseen by an advisory committee including the sponsor (Dr. Caughey) and experts on cysteine proteases (Dr. McKerrow), immunohistopathology (Dr. McDonald), and pulmonary diseases (Dr. Matthay), who will guide the candidates development into an independent investigator in pulmonary research.