Components of the innate immune system comprise the first line of defense against the encroachment of microorganisms at mucosal surfaces. Deficiencies in this system are believed to play a role in diseases such as Cystic Fibrosis, which is characterized by chronic bacterial infection. Defensin peptides are abundant components of the innate immune system, particularly in the human upper airway. Respiratory mucosal epithelium produces human beta-defensin precursors, known as hBD-1 and hBD-2, which undergo amino-terminal processing to attain full antimicrobial activity. However, the mechanism by which this processing occurs is not known. Previous work in mice has demonstrated that the matrix metalloproteinase MMP-7 co-localizes with and cleaves the pro segment from intestinal alpha-defensin precursors. In humans, MMP-7 is expressed in a variety of glandular epithelia throughout the human body, making it an ideal candidate for the enzyme that processes hBD-1 and -2. In addition, preliminary data showed that MMP-7, like hBD-2, is prominently upregulated in epithelial cells exposed to bacteria, further supporting the idea that MMP-7 participates in host defense. This proposal will test the hypothesis that MMP-7 and the hBDs are co-regulated; furthermore, that MMP-7 mediates processing of beta-defensin precursors in airway tissue, thus producing more efficacious defense molecules at the mucosal surface. To test this hypothesis, the following specific aims are proposed: (1) assess co-expression of MMP-7 and beta-defensins produced by conducting airway epithelial cells both constitutively and following bacterial infection. This will address the likelihood that these molecules could be functionally interactive; (2) analyze the beta-defensin isoforms produced from airway cells and tissue in the presence of MMP-7. Determine if MMP-7 is required for beta-defensin processing in an in vivo-like context. This will be accomplished by using metalloproteinase inhibitors as well as genetically defined cells; and (3) quantitatively assess the antimicrobial properties of beta-defensin isoforms produced by MMP-7 cleavage. The proposed experiments should provide insight into the role of MMP-7 in mucosal epithelium and how this metalloenzyme contributes to mechanisms of host defense.
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