QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online 24 November 2005
doi: 10.1242/dev.02175

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Zhang, X. Articles by Ornitz, D. M. Search for Related Content PubMed PubMed Citation Articles by Zhang, X. Articles by Ornitz, D. M.

Reciprocal epithelial-mesenchymal FGF signaling is required for cecal development

¹ Department of Molecular Biology and Pharmacology, Washington University School of Medicine, 660 South Euclid Avenue, St Louis, MO 63110, USA.
² Department of Pathology and Immunology, Washington University School of Medicine, 660 South Euclid Avenue, St Louis, MO 63110, USA.
³ The Center for Genome Sciences, Washington University School of Medicine, 660 South Euclid Avenue, St Louis, MO 63110, USA.

View larger version (112K): [in a new window]   Fig. 1. Fgf9, FGFR1 and FGFR2 expression in the embryonic cecum. (A,B) Whole-mount in situ hybridization was performed on isolated E11.5 (A) and E13.5 (B) wild-type GI tracts to detect Fgf9 expression. Insets show small intestine and colon. Fgf9 expression was uniformly expressed in the epithelium of the cecum, small intestine and colon. (C) Frozen section of an E13.5 whole-mount in situ hybridization showing Fgf9 expression in the cecal epithelium (arrows). (D-F) Immunohistochemical analysis of E12.5 cecal sections stained with an anti-FGFR1 (E) or anti-FGFR2 (F) polyclonal antibody. Control (D) staining, in which the primary antibody was absent, shows no signal. D',E' and F' are higher magnification views of the boxes in D,E and F, respectively. FGFR1 (E') and FGFR2 (F') are expressed in both cecal epithelium (black arrows) and mesenchyme (blue arrows). Co, colon; I, Ileum; SI, small intestine; Ce, cecum.

View larger version (108K): [in a new window]   Fig. 2. The development of the cecum is abnormal in both Fgf9-/- and Fgf10-/- embryos. (A-F) Morphologic comparison of the development of the cecum in WT, Fgf9-/- and Fgf10-/- embryos at E12.5, E14.5 and E18.5. (G-I) Hematoxylin and Eosin (H&E) stained sections of E14.5 WT, Fgf9-/- and Fgf10-/- cecum. In WT embryos (A-G), the cecum is located at the ileo-colonic junction. (A-C,H) Fgf9-/- embryos show a normal appearing epithelial tube throughout the small intestine and colon, but no identifiable cecal bud. (D-F,I) Fgf10-/- embryos develop a mesenchymal bud at the ileo-colonic junction, but no epithelial budding or expansion into cecal mesenchyme was observed. (F) By E18.5, the cecal mesenchymal bud appeared to degenerate in Fgf10-/- tissue (arrow). Dashed lines indicate the border between the epithelium and mesenchyme in the cecal region. Arrows indicate the position of the cecal tip or the position of the ileo-colonic junction in the absence of a cecum. M, mesenchyme.

View larger version (122K): [in a new window]   Fig. 3. Reduced cell proliferation rates in Fgf9-/- cecal mesenchyme and epithelium. (A,B) H&E stained paraffin sections from E12.5 wild-type (A) and Fgf9-/- (B) cecum. The amount of mesenchyme (M) in Fgf9-/- cecal tissue is reduced compared with in wild-type tissue. The insets show a low-power view of the entire cecal section; the boxed region indicates the enlarged area shown. (C,D) BrdU labeling of adjacent cecal sections demonstrating reduced cell proliferation in both the mesenchyme and epithelium (arrow) in Fgf9-/- (D) tissue compared with in wild-type tissue (C). Dashed line indicates the border of mesenchyme and epithelium; the solid line indicates the apical cap of the cecal epithelium. (E) Quantification of percentage of BrdU-positive nuclei showing significantly decreased cell proliferation in both cecal mesenchyme (*P<0.001) and epithelium (**P<0.01) in Fgf9-/- tissue.

View larger version (71K): [in a new window]   Fig. 4. FGF9 signaling is required for Fgf10 expression. Whole-mount in situ hybridization was performed on E11.5 and E12.5 embryonic GI tracts from wild-type (WT), Fgf9-/- and Fgf10-/- embryos. Fgf10 was expressed in the cecum mesenchyme in WT tissue (A,C), but was completely absent in Fgf9-/- tissue (B,D). By contrast, Fgf9 was expressed in cecum epithelium from WT and Fgf10-/- tissue (E,F). Co, colon; I, terminal ileum.

View larger version (135K): [in a new window]   Fig. 5. Effect of FGF9 and FGF10 on growth of cecal explants. E12.5 embryonic wild-type cecums were removed and embedded in Matrigel. BSA-, FGF9- or FGF10-soaked heparin beads were placed one bead diameter from the cecal explant. (A-C) During the first 12 hours, the cecal epithelium and mesenchyme did not significantly extend toward either the FGF9- or FGF10-soaked bead compared with the BSA-soaked control bead. (D-F) After 84 hours, the cecal epithelium had expanded and extended toward the FGF10 bead (E,E',H) compared to the BSA bead (D,D',G); however, no mesenchymal growth was observed (red solid line in D',E',G,H). By contrast, cecal mesenchyme showed significant growth accompanied by epithelial expansion when placed adjacent to an FGF9 bead (F,F',I). Note the significantly thickened mesenchymal regions in F' and I (line). D',E' and F' show higher magnification brightfield views of the explants in D, E and F, respectively. (G-I) H&E stained frozen sections showing the histology of cultured cecal explants at 84 hours. Red dashed lines indicate the border of mesenchyme and epithelium. The solid lines in D'-F',G-I indicate the thickness of the cecal mesenchymal layer.

View larger version (62K): [in a new window]   Fig. 6. Distal small intestinal epithelium extends toward an FGF10, but not an FGF9, source. BSA-, FGF9- or FGF10-soaked beads were placed in Matrigel two-bead diameters from a wild-type distal intestinal epithelial explant in which mesenchymal tissue had been removed. (A-D) Intestinal epithelium did not grow when exposed to a BSA-soaked control bead over an 84-hour period. (E-H) In response to an FGF9 bead, intestinal epithelium showed a modest dilation but no directional extension towards the bead. (I-L) In response to an FGF10 bead, intestinal epithelium showed a significant dilation and extension towards the bead.

View larger version (64K): [in a new window]   Fig. 7. Loss of Bmp4 but not Shh expression in the developing cecum of Fgf9-/- mice. Whole-mount in situ hybridization was performed on E12.5 and E13.5 embryonic GI tracts from wild-type (WT), Fgf9-/- and Fgf10-/- embryos. (A-D) At E12.5 and E13.5, Bmp4 was expressed strongly in the cecal mesenchyme in WT tissue (A,B) and Fgf10-/- tissue (C,D). (E,F) At these same stages, Bmp4 expression was absent in Fgf9-/- cecum. (G,H) At E12.5, Shh was expressed in cecal epithelium in WT tissue, and its expression was not significantly affected by the absence of expression of either Fgf9 or Fgf10.

View larger version (24K): [in a new window]   Fig. 8. Model of reciprocal FGF signaling in cecum development. During early cecal development, epithelial expression of FGF9 signals to mesenchymal FGFRs (FGFR1c and FGFR2c) to stimulate local mesenchymal proliferation and gene expression (Fgf10, Bmp4). Mesenchymal FGF10 signals to the epithelial FGFR2b to stimulate epithelial proliferation and extension of the epithelial bud into the expanded cecal mesenchyme. FGF9 is only able to induce mesenchymal proliferation and Fgf10 expression in the permissive junctional mesenchyme.