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GRK 1482 Jahrbuch 2011-2014

Publications [1] Sydora B, McFarlane S, Doyle J and Fedorak R. Neonatal exposure to fecal antigens reduces intestinal inflamma- tion, Inflammatory Bowel Disease. 2011, 17: 899–906. [2] Wieten L, Berlo S, Brink C, van Kooten P, Singh M, van der Zee R, Glant T, Broere F and van Eden W. IL-10 Is Criti- cally Involved in Mycobacterial HSP70 Induced Sup- pression of Proteoglycan-Induced Arthritis, PLoS ONE. 2009, Volume 4, Issue 1, e4186. [3] Shkoda A, Ruiz P, Daniel H, Kim S, Rogler G, Sartor B and Haller D. Interleukin-10 Blocked Endoplasmic Reticulum Stress in Intestinal Epithelial Cells: Impact on Chronic Inflammation, Gastroenterology. 2007; 132: 190–207. [4] Muir W, Bryden W and Husband A, Evaluation of the Efficacy of Intraperitoneal Immunization in Reducing Salmonella typhimurium Infection in Chickens, Poultry Science, 1998. 77:1874–1883. ASSOCIATED FELLOWS GRK Progress Report 2011-2014 | Page 67 Aim The aim of the project is to identify dominant gut microbial trig- gers of experimental ileitis. We hypothesized that challenge of mice with bacterial HSP or caecal bacterial lysates via the intraperitoneal route leads to changes in (I) immune responses in association with the seve- rity of ileitis, and (II) the diversity and composition of the intes- tinal microbiota. Methods and Results We use the Tnf∆ARE/wt mouse as a model for CD-like ileitis be- cause experiments with antibiotics showed that inflammation is dependent upon intestinal bacteria in this model. Tnf∆ARE/wt mice show higher stability of Tnf-α mRNA which results in the development of ileal inflammation by the age of 8 weeks. We challenged mice (n = 4 to 7) on day 4, 11, 18 and 28 after birth by intraperitoneal injection of either a recombinant strep- tococcal HSP70, caecal bacterial lysates (CBL) from 13-week- old Tnf∆ARE/wt mice (ARE-CBL) or appropriate control solutions (Figure 1). Mice were sacrificed and samples were collected at the age of 8 weeks. The diversity and composition of luminal and mucosa-associated intestinal microbiota were analyzed by sequencing 16S rRNA gene amplicons (V4 region) using the Illumina MiSeq System. The severity of inflammation is as- sessed by scoring of paraffin-embedded distal ileal and proxi- mal colonic tissues sections as described previously (Katakura et al., 2005). Additionally, we cultured distal ileal and proximal colonic tissue to examine modifications in the release of immu- noglobulins and inflammatory cytokines by ELISA. Preliminary results showed that the challenge of Tnf∆ARE/wt mice with HSP70 did not result in significant differences in body weight development, spleen and caecum weight, T-cell popula- tions and histological scores in the distal ileum. However, beta- diversity analysis of caecal microbiota indicated substantial effects on bacterial diversity (Figure 2). The analysis of plasma samples revealed marked anti-HSP70-IgG and -IgM responses in the treatment group. The challenge of Tnf∆ARE/wt mice with ARE-CBL led to a significant reduction in spleen weights in the treatment group when compared with sham-injected mice (Figure 3). Outlook Completion of the histological scoring of distal ileum and pro- ximal colon is underway. High-throughput sequencing of 16S rRNA genes was used for the analysis of microbial diversity and composition. These data need to be evaluated in order to identify differences in the composition of luminal and mucosa- associated intestinal microbiota associated with treatments. Figure 1: Experimental design of the i.p. challenge with recombinant strep- tococcal HSP70 or caecal bacterial lysate Figure 2: Beta-diversity analysis of caecal microbiota of Tnf∆ARE/wt mice challenged with recombi- nant streptococcal HSP70 (blue = Tnfwt/wt HSP70; red = Tnf∆ARE/wt HSP70; green = Tnf∆ARE/wt control). Figure 3: Spleen weights of mice chal- lenged with ARE-CBL or control Supervisors Prof. Dr. Dirk Haller I TUM | Nutrition and Immunology Dr. Thomas Clavel I TUM | Nutrition and Immunology Start of project: May 2012 Academic background: Studies of Nutrition Science at Technische Universität München