Increasing resistance to conventional antibiotics has focused attention on alternative anti-infectious therapies. Antimicrobial peptides are promising new agents that have a low susceptibility to microbial resistance mechanisms. Unlike conventional antibiotics, antimicrobial peptides are encoded by single genes and can be introduced into infected tissues by gene therapy approaches. The ultimate goal of this study is to introduce antimicrobial peptide genes into explanted human cells, and to use these augmented cells to enhance host defense mechanisms against infection. The immediate goal is to develop a model system, in which the human-Beta-defensin-2 (HBD-2) gene will be expressed in HBD-2 negative cells; its antimicrobial effect will be tested in vitro and in vivo. The HBD-2 gene was chosen because, unlike most known defensins, it does not require tissue-specific processing. A retrovirus carrying the HBD-2 gene has permitted successful transduction of mouse fibroblast NIH/3T3 cells to secrete functional antimicrobial peptides in vitro. Additionally, successful transduction of various other cell types to secrete HBD-2 has been accomplished. While antibacterial activity of HBD-2 in vitro has been clearly demonstrated, limited information is available regarding HBD-2 function in vivo. Using a novel approach, there are plans to test whether secreted HBD-2 is functional in vivo. The central hypothesis is that this antimicrobial gene therapy is effective in enhancing host innate immunity against infection. One short-term objective is to test whether the transduced cells will defend against infection in vivo using mouse models.
The Specific Aims i nclude: 1) to determine the temporal expression and the antimicrobial effects of HBD-2 in transplanted cells in vivo in a mouse model.
This Aim will utilize SCID mice to grow HBD-2 expressing tumors which model will allow quantitative measurement of the antimicrobial effect of HBD-2 in vivo. 2) To establish a tissue-engineered wound healing model to test antimicrobial gene therapy in mice.
This Aim will establish a model that simulates a clinical situation for testing the application of this antimicrobial gene therapy approach. Successful completion of these Aims will be the first step in establishing in vivo animal study models to explore the future applicability of antimicrobial gene therapy. The proposed approach is intended to provide potential benefit in clinical applications in three ways: 1) transplanted antimicrobial secreting cells can be used in wounded, engineered or infected tissues to prevent and/or fight against infections; 2) these cells may reduce the need for conventional antibiotics; and 3) these cells may provide long-term augmentation of the innate immune system for immunocompromised patients.
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