Figure 4

The tracing of cells originally expressing Shh in tooth primordia visualized by X-gal staining. Left – whole mount staining of the lower jaw, right - corresponding histological sections. The Shh expressing cells and their descendants in the R2 rudiment and M1 germ are compared during supernumerary tooth formation in Spry4-/- mutants and control mice. The tamoxifen dose was administrated into pregnant female mice LacZ (A, B) and Spry4-/-LacZ (C, D) at E13.5, because Shh is expressed in R2 rudiment in this time. In control embryos, merging of two labelled descendant cell populations in areas of original R2 (dR2, red arrow) and M1 (dM1, yellow arrow) at E15.5 was clearly detectable (A) leading to a single area of blue cells in M1 germ at E16.5 (B). In contrast, Spry4-/-LacZ embryos showed two isolated areas of blue cells indicating descendant cell populations of original R2 (dR2) and M1 (dM1) at E15.5 (C) as well as at E16.5 (D). The area between descendant original R2 and M1 showed no Shh expression (green arrow). The anterior area of blue cells was located in the center of developing supernumerary tooth primordium (S, black arrowhead) originating in the R2 rudiment. Black arrows indicate the areas of tooth germs shown on histology without Shh expression. The scale bar indicates 2 mm on whole mount, or 100 μm on slices.