Allergen-induced IgE and Th2-associated inflammation have been implicated in the pathogenesis in allergic asthma. The inflammation is characterized by various types of cellular infiltrates, including eosinophils and T cells. A subset of CD4+ T cells (Th2) which has been distinguished functionally by its pattern of cytokine secretion, is thought to play a key role. Th2 cells are thought to promote pulmonary allergic responses through their secretions of cytokines, IL-4 and IL-5, which promote IgE production and mast cell development, and eosinophilia, respectively. Recent studies have provided evidence for a suppressive role of interferon gamma (IFN) in the development of Th2 and cytokine-mediated allergic responses. These studies demonstrate the importance of cytokines and the regulation of Th2 responses, and suggest potential therapeutic approaches using Th2-inhibitory cytokines. The objective of this proposal is to determine the role of IFN-gamma in the regulation of Ag-induced airway inflammation and bronchial hyperreactivity by gene transfer of IFN-gamma directly into lung mucosal cells. The investigators hypothesize that delivery of this gene to the lung mucosal cells will decrease local eosinophilic inflammation and limit tissue destruction and loss of pulmonary function. The model for these studies will evaluate hyperresponsiveness to well characterized Ags, Conalbumin and Ovalbumin, in inbred strains of mice. In support of this approach are findings that the potent IFN-gamma inducer, IL-12, is able to reverse Ag-induced inflammation and airway hyperresponsiveness. Thus, this model is ideal for the design of gene transfer experiments to examine the therapeutic efficacy of IFN-gamma using in vivo gene delivery. The investigators propose to develop, characterize, and test the efficacy of two independent gene delivery systems. These include the expression of IFN-gamma delivered to lung mucosa through liposome and adenoviral mediated gene transfer.
Specific aim 1 proposes to develop and characterize both liposome and second generation recombinant adenoviral vector systems for delivery of IFN-gamma expression which will be tested in vitro in a human bronchial epithelial cell line. The second specific aim proposes to optimize the route and dose of gene delivery by both liposome- and adenovirus-mediated gene transfer using methods of detecting IFN-gamma gene expression which include RTPCR, ELISA, immunocytochemistry and in situ hybridization. The third specific aim attempts to address the in vivo efficacy of transduced IFN-gamma genes in the suppression of Ag-induced airway inflammation and bronchial hyperresponsiveness. The development of this model will provide an experimental basis for exploring the potential efficacy of gene therapy using cytokines for allergic diseases in humans.
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