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Stem-cell-based Tissue Engineering of Murine Teeth

Department of Craniofacial Development, Floor 28, GKT Dental Institute, King’s College, Guy’s Hospital, London Bridge, London SE1 9RT, UK;

View larger version (47K): [in a new window]   Figure 1. Sections of heterotypic recombinations between ES cell aggregations and embryonic oral epithelium. (A,D) Lightfield photomicrograph showing epithelial bud formation in recombinant explants. (B,C) Sections adjacent to (A) showing radioactive in situ hybridization for Lhx7 (B) and Msx1 (C). (E) Section adjacent to (D) showing radioactive in situ hybridization for Pax9 (E). Tooth germ epithelium is outlined in red. Scale bar: 100 µm.

View larger version (71K): [in a new window]   Figure 2. Sections of heterotypic recombination between neural stem cell aggregations and embryonic oral epithelium (A-D), between bone-marrow-derived cells and embryonic oral epithelium (E-H), and between NIH3T3 cell aggregations and embryonic oral epithelium (I-L). (A) Lightfield photomicrograph showing localization of epithelium in a recombinant explant. (B-D) Sections adjacent to (A) showing radioactive in situ hybridization for Lhx7 (B), Msx1 (C), and Pax9 (D). (E-H) Adjacent sections of a recombination between bone-marrow-derived cells and embryonic oral epithelium. (E) Example of GFP expression in embryonic oral epithelium in a recombination. (F-H) Expression of Lhx7 (F), Msx1 (G), and Pax9 (H) in bone-marrow-derived cells adjacent to the embryonic oral epithelium. (I) Lightfield photomicrograph showing localization of epithelium in recombinant explants. (J-L) Sections adjacent to (I) showing no expression of Lhx7 (J), Msx1 (K), and Pax9 (L). Tooth germ epithelium is outlined in red. Scale bar: 100 µm.

View larger version (94K): [in a new window]   Figure 3. Recombinant explant between bone-marrow-derived cells and oral epithelium following 12 days of development in a renal capsule. All the tissues visible are donor-derived, since the host kidney makes no cellular contribution to the tissue. Where epithelium in the recombinations was from GFP mice, in situ hybridization of sections of these tissues confirmed that all mesenchyme-derived cells were of wildtype origin (not shown). BO, bone; Am, ameloblasts; DP, dental pulp; OD, odontoblasts, E, enamel; D, dentin. Scale bar: 80 µm.

View larger version (146K): [in a new window]   Figure 4. Upper jaw teeth in adult mouse mouth 26 days after transplantation of E14.5 molar explants. Molar tooth primordia with surrounding tissue dissected from E14.5 C56/B6 mice were cultured in vitro for 24 hrs. The mouse dentition consists of one incisor separated from three molars by a toothless region (diastema) in each quadrant of the mouth. A small incision was made in the soft tissue of the maxilla in the diastema of adult (over 20 wks) male mice. The explants (approximately 2 mm) were placed in the incision and fixed with surgical glue (Vetbond, 3M, St. Paul, MN, USA). The transplanted explants were left for 26 days, during which animals were fed a soft diet. After fixation and decalcification, wax serial frontal sections were cut and stained (H&E). A, incisors. B, first molar. C, second molar. D, third molar. E, ectopic tooth in diastema region (between A and B). Arrows show ectopic bone. F, High magnification of boxed area in E, showing periodontal-ligament-like tissue. d = dentin. pd = pre-dentin. Scale bars: 1.2 mm (A-D); 1.0 mm (E); 50 µm (F).