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Control vs CF Small Intestine

ABSTRACT: Total RNA was prepared from the entire small intestines of 40 day old Control and CFTR null mice (2 males and 1 female of each genotype), congenic on the black6 background, using TRIzol reagent. Mice were fed Peptamen from age 10 days to prevent intestinal obstruction. Keywords: parallel sample

ORGANISM(S): Mus musculus

PROVIDER: GSE765 | GEO | 2004/01/26

SECONDARY ACCESSION(S): PRJNA87719

REPOSITORIES: GEO

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Project description:Paneth cells of intestinal crypts contribute to host defense by producing antimicrobial peptides that are packaged as granules for secretion into the crypt lumen. Here, we provide evidence using light and electron microscopy that postsecretory Paneth cell granules undergo limited dissolution and accumulate within the intestinal crypts of cystic fibrosis (CF) mice. On the basis of this finding, we evaluated bacterial colonization and expression of two major constituents of Paneth cells, i.e., {alpha}-defensins (cryptdins) and lysozyme, in CF murine intestine. Paneth cell granules accumulated in intestinal crypt lumens in both untreated CF mice with impending intestinal obstruction and in CF mice treated with an osmotic laxative that prevented overt clinical symptoms and mucus accretion. Ultrastructure studies indicated little change in granule morphology within mucus casts, whereas granules in laxative-treated mice appear to undergo limited dissolution. Protein extracts from CF intestine had increased levels of processed cryptdins compared with those from wild-type (WT) littermates. Nonetheless, colonization with aerobic bacteria species was not diminished in the CF intestine and oral challenge with a cryptdin-sensitive enteric pathogen, Salmonella typhimurium, resulted in greater colonization of CF compared with WT intestine. Modest downregulation of cryptdin and lysozyme mRNA in CF intestine was shown by microarray analysis, real-time quantitative PCR, and Northern blot analysis. Based on these findings, we conclude that antimicrobial peptide activity in CF mouse intestine is compromised by inadequate dissolution of Paneth cell granules within the crypt lumens. Total RNA was extracted from pooled small intestines of three WT and three CF mice using Tri-Reagent (Molecular Research Center, Cincinnati, OH), and poly(A) RNA was purified by using the MicroPoly(A) mRNA purification kit (Ambion, Austin, TX). The WT and CF poly(A) RNA samples were sent to IncyteGenomics (St. Louis, MO) where they were labeled with cyanine 3 (Cy3) and Cy5, respectively, and hybridized with the UniGEM1.31 array representing 9,570 known genes and expressed sequence tags.

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Control vs CF Small Intestine

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