In inflammatory airway diseases such as acute and chronic infection, asthma (late phase), cystic fibrosis, and inhalant exposure to ozone there is an influx of neutrophils into the airways. These cells have been implicated in the pathophysiology of these diseases. The role of adhesion molecules in the recruitment of neutrophils into inflamed tissue has focused mainly on interactions between leukocytes and endothelial cells. Much less is known about adhesion between neutrophils and airway epithelial cells during or after migration of neutrophils across the epithelium into the airway. To date, intercellular adhesion molecule-1 (ICAM-1) is the only identified molecule on airway epithelium that mediates adhesion with neutrophils. Data from the P.I.'s lab strongly suggest that infection with respiratory viruses or ozone exposure induces non-ICAM-1 adhesion molecules that also interact with leukocyte CD18 integrins. The long term goals of this research are 1) to identify and characterize the specific adhesion molecules involved in these interactions, especially novel adhesion molecules on airway epithelial cells and 2) to determine the role of these molecules in the pathophysiology of airway inflammation and their potential as therapeutic targets for anti-inflammatory therapy.
The specific aims of this pilot project are: (1) to clone and characterize novel airway epithelial cell adhesion molecules using a cDNA library from ozone-exposed airway epithelial cells expressed in 293 cells. Trasnsfected 293 cells will be enriched either for antibody binding using our new monoclonal antibodies that recognize and block a non-ICAM-1 epithelial cell adhesion molecule, or using a solid-phase integrin binding assay that will enrich for binding to a natural ligand of the sought-after molecule(s). cDNAs rescued from cells enriched for the desired function will cloned and sequenced. (2) to characterize cloned cDNAs encoding for adhesion molecules of interest in pure expression systems by transfection into COS or HeLa cells in order to a) confirm the adhesive function of products encoded by cloned cDNAs of interest and b) compare the proinflammatory effects of neutrophil-epithelial cell adhesion via ICAM-1 vs. novel cloned adhesion molecules, including stimulated release of selected cellular products and prolongation of leukocyte survival (delayed apoptosis). Success in cloning and characterizing novel airway epithelial adhesion molecules will not only provide important new information about inflammatory airway pathophysiology, but also will yield new potential targets for anti-inflammatory therapy in the airways.
