Abstract
1012
The intestinal epithelium is a continuously differentiating tissue in which the high rate of cellular self-renewal may contribute to the susceptibility of intestinal epithelial cells to malignant transformation. To address the physiological role of Notch signaling in intestinal homeostasis and cell lineage commitment, the gene encoding Pofut1, a fucosyltransferase required for the activity of all Notch receptors, was deleted in the mouse intestinal epithelium, through Villin-Cre-mediated recombination. Canonical Notch signaling inhibition was confirmed by concomittant down- and up-regulation of Hes1 and Math1 mRNA levels, respectively. The body weight of Pofut1F/F:VillinCre mice was dramatically reduced compared to control littermates. Macroscopically, an increased layer of mucus secretion was observed on the intestinal epithelial surface of Pofut1F/F:VillinCre mice. Removal of Pofut1 led to a massive increase in commitment to the secretory cell lineage characterized by increased numbers of goblet cells, Paneth cells and entero-endocrine cells, as revealed by alcian blue, lysosyme and chromogranin A/B immunostaining, respectively. Consistent with this, the levels of mRNA encoding gut hormones (CCK, GIP, Glucagon, Secretin, Somatostatin), Paneth cell markers (MMP7) and mucus-secreting cell markers (MUC2, TFF3, Spink4, Spdef, FIZZ2) were enhanced, as determined by Affymetrix GeneChip® arrays. The specific allocations of Paneth and entero-endocrine cells along the crypt-villus axis were not modified in the small intestine and no changes in expression of the ephrin/Eph pathway components were observed. However, goblet cells accumulated in the crypts of both the small and large intestine. Interestingly, determination of cell renewal capacity in the intestinal mucosa, through immunostaining of the PCNA and Ki67 antigens, revealed that the transit amplifying compartment was maintained in the upper crypts of the intestinal mucosa whereas very rare proliferating cells were detected in the colonic epithelium. This was associated with abnormal tissue architecture in the colon, the epithelium being composed entirely of differentiated, mostly mucus- secreting cells. Consistent microarray gene expression data revealed that absorptive cell markers (L-FABP, AFP, IAP, DPP4, ApoB, CD36) were repressed in intestinal epithelium lacking Pofut1, with a more drastic effect in the colonic epithelium. Therefore, we conclude that canonical Notch signaling maintains intestinal homeostasis by promoting crypt progenitor cell proliferation and controlling the balance between secretory and absorptive cell types.
98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA
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