Fig. 3.
Contribution of dHAND lineage cells to arch-derived structures of the jaw of dHAND-Cre;R26R embryos. Transverse (A–C) and frontal (D–I) sections through the heads of dHAND-Cre;R26R (A, D, G), dHAND-lacZ (B, E, H), and wild type (C, F, I) embryos. Frozen sections from dHAND-Cre;R26R and dHAND-lacZ embryos were stained for β-gal activity and counterstained with nuclear fast red (E13.5) or eosin (E18.5). Paraffin sections from wild type embryos were probed with a 35S-labeled riboprobe against dHAND. After developing, sections were counterstained with Harris hematoxylin and photographed under darkfield conditions. To aid in the visualization of specific structures, yellow lines have been added to some of the darkfield panels. (A–C) Transverse sections through the lower jaw of E13.5 embryos. (A) Labeled cells in dHAND-Cre;R26R embryos are observed in structures derived from the first pharyngeal arch, including mandibular bone (md), Meckel’s cartilage (mc), tongue (t), and surrounding mesenchyme/connective tissue. Abundant labeled cells are also observed in the mesenchyme of the developing submandibular glands (smg), but not in the glandular epithelium (see inset, A′). The mesenchyme of the upper jaw (future maxilla; *) and oral cavity epithelium do not contain labeled cells. (B) Labeled cells in dHAND-lacZ transgenic embryos are observed in a similar pattern to those in dHAND-Cre;R26R embryos, though labeled cells in Meckel’s cartilage are mixed with unlabeled cells. (C) Endogenous dHAND expression is observed in the lamina propria of the tongue and in the mesenchyme surrounding the proximal ducts of the submandibular gland. Very little expression is observed in the mandible or Meckel’s cartilage. (D–I) Frontal sections through the mandible of E18.5 embryos. (D) β-gal-stained cells in dHAND-Cre;R26R embryos are present in the connective tissue of the tongue (t), with highest staining in the lamina propria (black arrow). Almost all cells in the mandible, Meckel’s cartilage (yellow arrow), and surrounding mesenchyme are also labeled. (E) Labeled cells in dHAND-lacZ embryos are again observed in a similar pattern to those in dHAND-Cre;R26R embryos. However, a mixture of labeled and unlabeled cells composes the mandibular bone and Meckel’s cartilage. (F) Faint dHAND expression is observed in the mandibular bone. Very little expression is observed in Meckel’s cartilage. The asterisks in (D–F) mark the enamel organ of the left incisor. (G) In a more proximal section to that shown in (D–F), labeled cells in dHAND-Cre;R26R embryos are observed in Meckel’s cartilage and in the articular (ap) and coronoid (cp) processes of the mandible and surrounding perichondrium. However, fewer labeled cells are observed near the junction with the ossified mandible, an area undergoing endochondral ossification and composed of hypertrophic chondrocytes (see inset, G′). (H) Labeled cells are mixed with unlabeled cells in Meckel’s cartilage, but are not observed in the anterior and coronoid processes of the mandible. The perichondrium surrounding the process is composed almost solely of labeled cells. (I) Endogenous dHAND expression is not observed in the mandibular processes or in Meckel’s cartilage, though expression is observed in the sympathetic ganglion (yellow arrow). Due to the angle of section, this structure is not observed in the dHAND-Cre-R26R or dHAND-lacZ embryo sections, though scattered labeled cells are present in the sympathetic ganglion of dHAND-Cre-R26R embryos (see Fig. 4). mo, molar.