Intestine Specific Deletion of N-WASP Leads to Alteration of Gut Homeostasis in Mice : 394


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Intestine Specific Deletion of N-WASP Leads to Alteration of Gut Homeostasis in Mice : 394
Titre de la conférence
Digestive Disease Week (DDW)
Garber J.J., Nguyen D.D., Maillard M.H., Mizoguchi E., Bhan A.K., Snapper S.B.
Chicago, Illinois, May 30-June 4, 2009
Statut éditorial
Date de publication
Background: Wiskott-Aldrich Syndrome protein (WASP) is a cytoplasmic protein in hematopoietic cells that regulates actin assembly via the Arp2/3 complex. Deficiencies in WASP are associated with IBD in humans and spontaneous colitis in mice, and recent genomewide linkage studies provide evidence of novel susceptibility factors for UC in the region of the Arp2/3 locus. We sought to determine the role of the ubiquitously expressed NWASP protein, a key regulator of Arp2/3 activity and cytoskeletal dynamics, in the intestinal epithelium. Methods: To generate mice with gut-restricted deletion of N-WASP (intestine NWASP KO, iNWKO), mice expressing Cre recombinase under the control of the villin promoter (tgv-cre) were mated to mice homozygous for a floxed N-WASP allele (N-WASPL2L/L2L). In all experiments, iNWKO mice (N-WASPL2L/L2Ltgv-cre) were compared with NWASPL2L/L2L lacking the villin-Cre transgene. Intestinal epithelial cells were isolated by EDTA dissociation and centrifugation, and tissue was examined with H&E, Alcian blue, immunofluorescence and electron microscopy (EM). To examine roliferation and migration of N-WASP deficient enterocytes, mice were injected IP with BrdU and sacrificed at 2 and 24 hours. To assess N-WASP deletion in inflammation, 6-week old mice (5 KO,5 WT) were given 3.5% DSS in drinking water for 5 days, and sacrificed on day 12 for histology. Results: iNWKO mice were viable and fertile, but failed to appropriately gain weight; adults weighed on average 30% less than their WT (N-WASPL2L/L2L) littermate controls (n=25,p=0.007). There was no spontaneous development of enterocolitis. N-WASP deletion was confirmed
by PCR, and the absence of protein confirmed by Western blot on intestinal epithelial lysates. Numbers of Paneth, goblet and BrdU-incorporating crypt progenitor cells were similar. Notably, epithelial cells in iNWKO mice contained uncondensed nuclei and there was markedly increased enterocyte migration at 24 hours. EM revealed disorganized and clustered
colonic microvilli and the absence of a terminal web. The distribution of E-cadherin was similar between iNWKO mice and WT controls. iNWKO mice given DSS exhibited more weight loss, and a trend toward higher clinical scores of disease activity. Histology was similar in both DSS groups, with the exception of one iNWKO mouse that required euthanasia on day 9; this mouse demonstrated severe architectural changes, ulceration, and neutrophilic infiltrate. Conclusion: Deletion of intestinal N-WASP leads to a phenotype of wasting, increased intestinal cell turnover and microvillus structural abnormalities, linking the actin cytoskeleton to the maintenance of gut homeostasis.
Création de la notice
08/01/2010 15:26
Dernière modification de la notice
18/05/2018 15:17
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