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Fig. 3. Analysis of skeletal elements in chimeric embryos. Contribution of Smo^n/n (A-H) or Smo^n/+ (J) cells to the bone collar of long bones (A-F,J) and to the mandible (G,H) in chimeric embryos at E17.5. Smo^n/n and Smo^n/+ cells are stained blue as a result of the expression of ß-galactosidase and the sections are counterstained with Nuclear Fast Red (A,B,G,H), H&E (C,J), or Methyl Green (D). Mineralized tissues are stained black by the von Kossa method (A,B,G,H,J). (A,B) The bone collar forms on one side of the tibia (A, arrowheads) or the fibula (B, arrowheads), but fails to develop from the perichondrium on the opposite side which is composed of mutant cells. (C) The Smo^n/n perichondrial cells differentiate into chondrocytes (arrows). (D) Safranin-O stains red the cartilaginous matrix deposited by chondrocytes differentiated from the Smo^n/n perichondrial cells. (E,F) Dark-field images of in situ hybridization using ³⁵S-labeled riboprobes for Col1(II) (E) and Col 1(X) (F) on sections similar to that in D. (G,H) Smo^n/n cells contribute to the osteoblast population of the mandible. Asterisk in G denotes Meckel's cartilage. The boxed region in G is shown at a higher magnification in H. (J) In chimeras composed of Smo^n/+ and wild-type cells, the Smo^n/+ cells contribute to the bone collar (black arrowheads).

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