Defensins and cytokines in inflammatory bowel disease
Sammanfattning: Ulcerative colitis (UC) and Crohn’s disease (CD) constitute the two major inflammatory bowel diseases in man. Both are serious chronic illnesses of the intestine with severe debilitating effects. The etiology of the diseases is unknown, but involvement of both adaptive and innate immune reactions seems to be major factors in the pathogenesis. In this thesis the roles of key molecules of the adaptive immunity, i.e. interleukin-2 (IL-2), and innate immunity, i.e. ?-defensins, were studied both in human inflammation of the large intestine and in mouse colitis models.?-defensins are small endogenous peptides with antimicrobial activity. Previous studies showed that expression of human ?-defensin-2 (hBD-2), hBD-3, and hBD-4 is induced in colonic epithelial cells of UC patients. Here we demonstrate that cells expressing these three ?-defensins are present also in the colonic lamina propria of UC patients and less frequently in CD patients, and controls. These cells were identified as mature plasma cells by the highly specific CD138 marker, by their prominent IgA or IgG expression, and by their ultra structural characteristics. Immunoelectron microscopy analysis of the hBD-2 peptide demonstrated synthesis and transport for secretion. Defensin producing plasma cells were 2-3 times more abundant in UC colon than in control and CD colon. Additionally, hBD-2 mRNA expression was demonstrated in 3 out of 4 well-characterized plasma cell lines.Defensin expression was studied in large intestinal mucosa before and after onset of colitis in two colitis models - the IL-2 KO mouse and the dextran sulphate sodium (DSS) induced colitis mouse. Mouse ?-defensin-3 (mBD-3) and mBD-4 mRNA was expressed in colonic epithelial cells of homozygous IL-2 KO (IL-2-/-) mice with established colitis (15 weeks old) and at significantly higher levels than in apparently healthy wild type mice, heterozygous (IL-2+/-) mice, and 5 weeks old IL-2-/- mice. Similarly mBD-3 was expressed in epithelial cells of DSS treated mice with chronic colitis but not in DSS treated mice with acute inflammation. Cells expressing mBD-3 mRNA were seen also in colonic lamina propria of diseased animals. Thus, expression of ?-defensins in the colonic epithelium seems to be a consequence of the chronic inflammation.IL-2+/- mice have reduced levels of IL-2 in the intestinal mucosa but are clinically healthy. IL-2+/- mice showed markedly reduced susceptibility to DSS induced colitis. This was associated with a significantly reduced infiltration of both CD4+ and CD8+ T cells in the colonic mucosa and lower expression levels of the cytokines IL-2, IL-4, and IL-10 in colonic T cells compared to DSS treated wild type mice. These results suggest that reduced level of IL-2 leads to attenuated activation and function of colonic T cells in turn causing a milder colitis in response to DSS challenge.Interestingly, IL-2+/- mice had a reduced frequency of regulatory T cells (CD4+CD25+) in both small and large intestine compared to wild type mice. As the small intestine of IL-2-/- and IL-2+/- mice appear normal, small intestinal T cells have never been critically analyzed in these mice. The cytokine profile of small intestinal T cells in IL-2-/- and IL-2+/- mice was changed compared to wild type control mice (IL-2+/+) with significantly elevated expression levels of IL-10 and IL-4. DSS treatment of IL-2+/- mice caused a marked reduction in cytokine expression levels in small intestinal T-cells. These results suggest that lack of IL-2 and even the partial decrease seen in IL-2+/- mice influence T cell function locally in the intestinal mucosa and cause a skewed cytokine milieu also in the small intestine despite its normal histology.In this thesis we demonstrate the pivotal importance of different T-cell subsets in intestinal inflammation. The upregulation of intestinal antimicrobial peptides seems to be a consequence of chronic inflammation in an effort to minimize intestinal damage.
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