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GRK 1482 Jahrbuch 2011-2014

Publications [1] Shkoda A, Ruiz PA, Daniel H, Kim SC, Rogler G, Sartor RB, Haller D. Interleukin-10 blocked endoplasmic reticulum stress in intestinal epithelial cells: impact on chronic in flammation. Gastroenterology 2007, 132:190–207. [2] Rath E, Berger E, Messlik A, Nunes T, Liu B, Kim SC, Hoo genraad N, Sans M, Sartor RB, Haller D. Induction of ds RNA-activated protein kinase links mitochondrial un- folded protein response to the pathogenesis of intestinal inflammation. Gut 2011, 61:1269-78. [3] Ron D and Walter P. Signal integration in the endoplas- mic reticulum unfolded protein response. Nat Rev Mol CellBiol 2007, 8:519-529. [4]Shen J and Prywes R. ER stress signaling by regulated proteolysis of ATF6. Methods 2005, 35:382–389. [5]Grivennikov S. Inflammation and colorectal cancer: colitis-associated neoplasia. Semin Immunopatho 2013, 35:229–244. Outlook The pathology of colorectal cancer is not yet well understood and needs to be investigated. However, there is a lack of good mouse models mimicking the human disease. The nATF6 mouse shows a spontaneous formation of large intestinal tu- mors and could serve as a model for human pathology. Further, it points to a direct link between ER UPR activation, intestinal inflammation and the establishment of intestinal tumors. Thus, we want to evaluate our model regarding links to human dis- ease and investigate the cross-link between ER UPR, intestinal inflammation, and tumorigenesis. Further, we will study the influence of the intestinal microbiota and immune relevant mechanisms to this pathologic phenotype. ASSOCIATED FELLOWS GRK Progress Report 2011-2014 | Page 71 Aim The nATF6 mouse is a promising model to investigate the effects of an epithelial specific activation of the ER UPR on intestinal homeostasis. With this model system we aim to get a better understanding of the role of ER UPR activation under pathological conditions. Especially, it may serve as a tool to find cause and effect relationships between ER UPR and intestinal inflammation. Using both the heterozygous and the homozygous mouse, which both show ER UPR activation but differ in their pheno- typical outcome, we may be able to focus on dose-dependent differences. Due to the spontaneous development of intestinal symptoms and the tumorigenic phenotype we are confident to get mecha- nistic insights into the role of ER UPR in intestinal inflammation and colorectal cancer development. Methods and Results First analyses of the heterozygous Rosa-Stop flox-nATF6 Villin Cre mice (nATF6 flox/wt Cre+) show an activation of ER UPR response on protein and RNA expression levels using Western Blot and qPCR in small and large IECs isolated by density gra- dient centrifugation. Despite this activation, no spontaneous phenotype could be detected. However, using the DSS model to induce acute intestinal inflammation, heterozygous nATF6 mice showed higher severity of disease regarding weight loss and histopathology. In line with the results on the heterozygous mice, homozygous nATF6 flox/flox Cre+ mice have an upregulated ER UPR sig- naling as detected by Western Blot. In contrast to the heterozy- gous mice, a spontaneous intestinal phenotype, characterized by the development of diarrhea and the formation of colonic and caecal tumors, establishes in these mice. This is asso- ciated with a reduced lifespan as investigated in a long-term observation experiment. Using Ki67 immunofluorescence analyses increased prolifer- ation in tumor regions could be detected. Further, H&E and Alcian blue stainings for mucus and goblet cells show goblet cell depletion, immune cell infiltration, and dilated crypts filled with cellular detritus. The involvement of immune cell infiltration and apoptosis was further substantiated using immunofluores- cence stainings Figure: Alcian Blue stainings of colonic swiss rolls. In the nATF6 flox/flox Cre- control mouse section (A) a regular distribution of goblet cells can be seen whereas in the nATF6 flox/flox Cre+ mouse (B) less goblet cells but dilated crypts filled with cellular detritus are noticeable. Supervisors Prof. Dr. Dirk Haller I TUM I Nutrition and Immunology Start of project: January 2012 Academic background: Studies of Molecular Biotechnology at Technische Universtität München