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GRK 1482 Jahrbuch 2011-2014

Publications [1] Lakatos PL. Recent trends in the epidemiology of in- flammatory bowel disease: up or down? World J Gastro- enterol. 2006, 12: 6102-6108. [2] Seril DN, Liao J, Yang CS and Yang GY. Systemic iron supplementation replenishes iron stores without enhan- cing colon cancerogenesis in murine models of ulcera- tive colitis: Comparison with iron-enriched diet. Dig Dis Sci. 2005, 50: 696-707. [3] Werner T, Wagner SJ, Martínez I, Walter J, Chang JS, Clavel T, Kisling S, Schuemann K, Haller D. Depletion of luminal iron alters the gut microbiota and prevents Crohn‘s disease-like ileitis. Gut. 2011, 60: 325-33. [4] Bain CC, Scott CL, Uronen-Hansson H, Gudjonsson S, Jansson O, Grip O, Guilliams M, Malissen B, Agace WW and Mowat A. Resident and pro-inflammatory macro- phages in the colon represent alternative context- dependentfatesofthesameLy6Chimonocyteprecursors. Mucosal Immunol. 2012, doi: 0.1038/mi.2012.89. PhD FELLOWS GRK Progress Report 2011-2014 | Page 37 Aim We hypothesize that increased concentrations of luminal iron can enhance already existing intestinal inflammation in mouse models of colitis via interplay of changes in microbiota and im- mune responses in the colon due of iron overload. The aims of our study are: 1) to confirm differences in inflammatory response due to iron modulated diet in T cell transfer colitis as well as in Stat3LysMCre colitis model 2) to investigate if the proinflammatory effect of iron-rich diet is transferrable by fecal transplantation 3) try to identify and characterize immune cell populations as a possible mediators of immune responses caused by modulations of iron diet in our animal models of coli- tis with main focus on intestinal innate immune cells (macro- phages and DCs) and their interaction with T cells. Methods and Results In order to confirm the possible protective role of iron depleted diet in our colitis models, Rag-/- and Stat3LysMcre mice are going to be held on iron deficient vs. iron enriched diet in form of iron sulphate or hemin. Within 3-4 months Stat3LysMCre mice will spontaneously develop ileocolitis because of the granulo- cyte/macrophage defect in Stat3 signaling. In the T cell transfer colitis model Rag-/- mice are injected i.p. with CD4+CD62L+ naïve T cells which leads to the development of colitis. Our pre- liminary results show a protective effect of iron-deficient diet in Rag-/- mice after induction of colitis when compared with the mice that were kept on iron supplemented diet or control diet. However, mice kept on diet containing hemin showed delayed weight loss although they showed signs of severe diarrhea and inflammation confirmed by histology. In order to investigate whether enhanced inflammation caused by luminal iron over- load is exclusively a consequence of iron induced changes in microbiota composition, we will perform 16S RNA sequencing of fecal content from mice treated with different diets In addi- tion, microbiota transplantation from healthy Rag-/- mice kept on iron-supplemented diets into Rag-/- mice will be performed and the efect on colitis induction will be analyzed. To investiga- te the role of luminal iron on immune responses in our colitis models, the innate immune cells (DCs and macrophages) will be isolated from colon tissue and characterized phenotypically and functionally. For this purpose, we established an optimized LPL/IEL preparation protocol and a FACS staining panel suitable for macrophage and DC analysis. Supervisors PD Dr. Anne Krug | TUM | Klinikum rechts der Isar | Internal Medicine II Dr. Wolfgang Reindl | TUM | Klinikum rechts der Isar | Internal Medicine II Prof. Dr. Dirk Haller | TUM | Nutrition and Immunology Start of project: November 2011 Academic background: Studies of Molecular Biotechnology at the University of Zagreb, Croatia and of Bioindustrial Techniques, University of Orleans, France Outlook Initial experiments have confirmed a protective role of iron de- pleted diet in our T cell transfer model of colitis. For our future experiments we plan to investigate the mechanism by which luminal iron changes immune responses by dissecting direct and indirect effects of luminal iron on intestinal immune cells focusing on macrophages and DCs. Project outlook Different subpopulations of DCs and macrophages in healthy vs. infla- med gut will be analyzed. CX3CR1gfp+/+ Rag-/- mice are currently being generated which allow better discrimination of these subpopulations. Using ex vivo assays we plan to assess the ability of these cells isola- ted from mice held on different iron-supplemented diets to process and present antigen, to support Treg generation or to induce Th1/Th17 responses. Furthermore, mRNA expression levels for main iron metabolic proteins will be analyzed for different DC/macrophage subpopulations

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