Although the precise etiology is unknown, inflammatory bowel disease (IBD) is thought to occur as a result of a dysregulated mucosal immune response to environmental factors in a genetically predisposed individual. IL-18 is a cytokine that plays an important role in the pathogenesis of several chronic Th1-mediated disorders, including Crohn's disease (CD). In fact, a dramatic shift in IL-18 expression occurs during CD from intestinal epithelial cells (IEC) to mucosal immune cells (i.e. macrophages and dendritic cells), as the severity of disease increases. In addition, recent evidence supports the role of IL-18 as a protective factor during the acute phase of mucosal immune responses, when IEC are the primary source of IL-18. This novel function for IL-18 contrasts with the pathogenic role IL-18 is believed to play in more chronic phases of Th1-mediated inflammation. A genetic basis for differences in IL-18 regulation and expression observed in IBD may exist since recent studies have reported the association of specific single nucleotide polymorphisms (SNPs) in the IL-18 promoter region and several autoimmune diseases. Therefore, using genetic and molecular biology techniques as well as experimental models intestinal inflammation, the present study is designed to investigate the specific genetic factors that regulate IL-18 synthesis and to determine the precise function of epithelial and immune cell-derived IL-18 in the acute versus chronic phases of IBD. The central hypothesis of the present proposal is that IL-18 plays a key role in regulating normal innate immune responses in the gut mucosa and dysregulation of IL-18 may result in chronic intestinal inflammation characteristic of IBD. The following three specific aims are proposed to test this hypothesis: 1) Determine the relationship between polymorphisms in the IL-18 promoter region and IBD. The role of IL-18 in IBD susceptibility will be defined using an association approach by screening for recently described IL-18 promoter polymorphisms and performing case association studies of these genetic markers in well-characterized IBD and control populations. IBD multiplex families will also be used to test linkage disequilibrium between these marker loci and putative disease susceptibility loci in order to further characterize the transmission pattern of allelic variants. 2) Define the mechanism(s) of polymorphic IL-18 gene regulation in different mucosal cell populations. The functional relevance of IL-18 promoter polymorphisms will be achieved by creation of reporter constructs, site-directed mutagenesis experiments, and in vitro transfection assays in epithelial and macrophage cell lines. In addition, differential transcription factor binding and subsequent transcriptional regulation will be assessed in order to determine how the IL-18 gene is differentially regulated by IL-18 promoter polymorphisms, and if transcriptional control varies among different intestinal cell types. 3) Evaluate the specific role of IL-18 derived from different gut mucosal cell populations in an in vivo setting using experimental models of intestinal inflammation. The extent and severity of disease will be assessed in mice genetically- and immunologically-manipulated to produce IL-18 in either hematopoietic- (i.e. immune cells) or non-hematopoietic-cells (i.e. epithelial cells) following the induction of acute or chronic intestinal inflammation. These experiments will mechanistically address, in an in vivo setting, if IL-18 derived from specific mucosal cell populations and expressed during the acute versus chronic phases of disease, are involved in the pathogenesis of IBD. The ultimate goal of the present research proposal is to define the precise role of IL-18 in CD in order to develop specific treatment modalities aimed at modifying the natural course of this devastating disease.
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