The long term objective of this proposal is to test the hypothesis that the introduction of highly charged anionic polyelectrolytes into lung airway fluid as a result of inflammation and cell death contributes to the pathology of cystic fibrosis. Specifically the work will focus on the polyelectrolyte properties of F-actin and DNA as they relate to the abilities of these polymers to 1) increase the elasticity of CF sputum; 2) inactivate cationic antimicrobial proteins and peptides in the lung epithelium, and 3) interfere with gene therapy strategies that employ DNA-cationic lipid complexes for gene delivery into the epithelium. The experimental work will employ light scattering, fluorescence and polarization microscopy, rheology, and bacterial growth and lysis assays to study purified systems in which F-actin and DNA are caused to form bundles by lysozyme, defensins, histories and other cationic peptides released by epithelial cells and leukocytes. These studies will be expanded into determining how similar effects occur in sputum. Theories developed for polyelectrolyte condensation will be exploited to design soluble, polyvalent anions that independently or in combination with DNA- or actin-directed proteins such as DNAse and gelsolin can dissociate filament bundles and liberate antimicrobial function. The long term outcome of this work would be to determine if strategies based on depolymerizing DNA and cytoskeletal fibers or on dissociating bundles of these filaments by manipulating the ionic environment of airway fluid have potential to alleviate the pathology of CF.
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