Fig. 5. Accumulation of ß-catenin in Foxf mutant intestinal epithelium. ß-Catenin immunostaining (A-D,F-K) and TUNEL assay (E; red nuclei indicate apoptotic cells) of E18.5 small intestine of wild type (A,F,H,J), Foxf1^-/+; Foxf2^-/+ (B-E) and Foxf2^-/- (G,I,K). (A) In normal E18.5 small intestine, ß-catenin is most abundant in the basal epithelial cells of the intervillus pits and decreases along the villus axis (non-specific staining in the gut lumen is due to the stickiness of the meconium). (B-D) In Foxf mutants, ß-catenin concentration stays high all the way to the villus tip. Foxf1^-/+; Foxf2^-/+ differs from Foxf2^-/- in having a mosaic pattern with sharp boundaries between regions of the epithelium with high and low ß-catenin (C,D; arrowheads in B). (E) Patches of TUNEL-positive (apoptotic) cells match the low ß-catenin mosaic pattern. (F,G) Higher magnifications show nuclear localization of ß-catenin at base of villi in wild type (F), but ubiquitous nuclear ß-catenin in Foxf mutants (G). (H) Only membrane-associated ß-catenin is found only in cells from upper half of wild-type villus; nuclei (white arrowhead) are negative. (I-K) Both nuclear (white arrowheads) and membrane-associated ß-catenin are found in cells from the upper half of Foxf2^-/- villus (I; circular organelle in adjacent cell is a goblet cell mucus vesicle) and from the base of wild-type (J) and Foxf2^-/- villus (K). Epithelial depolarization is beginning in K, in spite of the seemingly intact contact with the mesenchyme: the membrane associated ß-catenin has begun to expand from the lateral and into the apical surface (black arrowhead) and nuclei are centrally located in the cells. (L-Q). Normal or reduced amounts of the major glycosaminoglycan proteoglycans in Foxf mutant intestine. Immunostaining with antisera against nidogen (Entactin; L,O), perlecan (M,P) and syndecan 1 (N,Q) on cryosections of E18.5 small intestine from wild type (L-N) and Foxf2^-/- (O-Q). Nidogen and perlecan are constituents of the mesenchymal ECM and present in normal amounts in Foxf mutants. Syndecan 1 is membrane anchored, present on the basolateral face of epithelial cells and reduced in Foxf mutant intestine.