Gene expression analysis in intestinal epithelial cells of germ-free versus wildtype mice
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE13908 | GEO | 2009/09/04
SECONDARY ACCESSION(S): PRJNA110323
REPOSITORIES: GEO
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Project description:Gene expression was analyzed in intestinal epithelial cells of germ-free and wildtype mice. Experiment Overall Design: Ileums and proximal colons of 4 C57/Bl6 control and 4 C57/BL6 germ-free mice were collected and embedded in OCT. Tissue sectioned (10 mikroM) and laser capture microscopy was performed using a Zeiss/Palm laser capture system. Total RNA was extracted from about 16000 cells of the catapulted samples using RNA microprep kit (Stratagene). These 16 000 cells derive from all 4 animals (pooled approx. 4 000 cells from each mouse). We retrieved between 50 and 150 ng total RNA. Fourty ng total RNA was expanded by two rounds of in vitro cRNA amplification and affymetrix genechip Moe403 were probed at the Swegene Resource Center Lund, Sweden. Array data analysis was performed using dCHIP2004 software.
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Project description:The development and severity of inflammatory bowel diseases (IBD) and other chronic inflammatory conditions can be influenced by host genetic and environmental factors, including signals derived from commensal bacteria. However, the mechanisms that integrate these diverse cues remain undefined. Here we demonstrate that intestinal epithelial cells (IECs) isolated from IBD patients exhibit decreased expression of the epigenome-modifying enzyme histone deacetylase 3 (HDAC3). Further, genome-wide analyses of murine IECs that lack HDAC3 (HDAC3ΔIEC) revealed that HDAC3 deficiency resulted in dysregulated gene expression coupled with alterations in histone acetylation. Critically, conventionally-housed HDAC3ΔIEC mice demonstrated loss of Paneth cells, impaired IEC function and alterations in the composition of intestinal commensal bacteria. In addition, HDAC3ΔIEC mice exhibited significantly increased susceptibility to intestinal damage and inflammation, indicating that epithelial expression of HDAC3 plays a central role in maintaining intestinal homeostasis. Strikingly, rederivation of HDAC3ΔIEC mice into germ-free conditions revealed that dysregulated IEC gene expression, Paneth cell homeostasis, and intestinal barrier function were largely restored in the absence of commensal bacteria. Collectively, these data indicate that the HDAC3 is a critical factor that integrates commensal bacteria-derived signals to calibrate epithelial cell responses required to establish normal host-commensal relationships and maintain intestinal homeostasis. In this study, we performed gene expression profiling to examine how the transcriptional profiles in primary live, EpCAM+ IECs from the large intestine differed between germ-free control HDAC3FF mice (3 biological replicates) and germ-free IEC-intrinsic knockout HDAC3ΔIEC mice (3 biological replicates).
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Project description:We compare transcriptomic profiles of intestinal epithelial cells obtained from the small intestine of germ-free and conventionally-caged mice. Intestinal epithelial cells were harvested from the intestine of conventional or germ-free C57Bl6J mice. Directional polyA RNA-seq was performed on RNA fom cells using standard Illumina protocols. Microbiota induce decreased expression of the Clec2e gene.
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