Crohn's disease (CD) is a debilitating condition of unknown etiology that is poorly responsive to currently available treatments. A working hypothesis suggests that CD may represent a dysregulated immune response to antigens derived from normal intestinal bacteria in a genetically predisposed host. Our studies will take advantage of a new strain of mice referred to as SAMP1/Yit/FC that develops enteritis spontaneously without genetic or immunologic manipulations. Preliminary studies indicate that these mice develop disease only when colonized with normal murine intestinal flora and not when derived under germfree conditions. The central hypothesis of this proposal is that in SAMP1/Yit/FC mice, normal intestinal bacteria are required for the development of ileitis through the induction of antigen-specific immune responses in the gut, and that proper manipulation of the bacterial flora may result in disease amelioration. The overall objective of this proposal is to investigate, in a mechanistic fashion, the role of the bacterial flora in experimental CD. In order to achieve our goals we will: 1) Determine the effects of antibiotic/probiotic administration on the severity of ileitis. The effects of antibiotic administration, as well as that of well-characterized probiotics will be investigated. In addition, the ability of antibiotic/probiotic treatment to maintain remission following initial treatment with anti-TNF or prednisone will be studied; 2) Characterize the composition of the bacterial flora as well as its functional effects on T cell activation. State-of-the-art techniques, including 16S rRNA PCR will be used to characterize the bacterial flora colonizing the ileum versus the colon of SAMP1IYitJFC mice. In addition, a series of in vitro T-cell activation studies as well as adoptive transfer experiments following stimulation with indigenous flora will be used to attempt to define the pathogenesis of ileitis in these mice; 3) Determine the effects of germ-free conditions and the role of specific bacterial species on chronic intestinal inflammation. SAMP1/Yit/FC mice will be re-derived under specific germ-free conditions and specific bacterial species will be re-introduced to determine their ability to induce chronic ileitis in germ-free mice. In addition, adoptive transfer experiments will be performed to define whether the primary abnormality of SAMP1/Yit/FC mice occurs in the T cells or is caused by specific bacterial antigens.
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