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Dental Epithelial Histomorphogenesis in vitro

¹ UMR INSERM 595 and
³ UMR 7104 CNRS-ULP INSERM U596, Faculté de Médecine, Université Louis Pasteur, 11 rue Humann, 67085 Strasbourg Cedex, France;
² Molecular Laboratory for Gene Therapy, Faculty of Stomatology, Capital University of Medical Sciences, Beijing, P.R. China;

View larger version (175K): [in this window] [in a new window]   Figure 1. In vitro development of re-associations between dental mesenchyme and dissociated epithelial cells from first lower molars at ED14. The re-associations were cultured for 12 hrs (A,F), 24 hrs (B,G), 2 days (C,H), 3 days (D,I,P,Q,R), 6 days (E,J), and 14 days (N,O,S,T). To visualize cell proliferation, we cultured ED14 first lower molars for 18 hrs in the presence of BrdU (K), and cultured re-associations for 54 hrs in vitro and 18 hrs in the presence of BrdU (P). Apoptosis was visualized after cells were stained for ssDNA on ED14 molars (L, arrow), and re-association was cultured for 3 days (Q, arrow). In situ hybridization for Shh was performed on first lower molars at ED14 (M), and re-associations were cultured for 3 days (R). After 14 days in culture, odontoblasts were functional and ameloblasts were polarized (N,O,S). Six cusps were observed, as can be seen on the histological horizontal section (O) or after 3D reconstruction of the mesenchyme (T). AM, ameloblast; BM, basement membrane; D, dentin; DE, dental epithelium; DM, dental mesenchyme; DP, dental papilla; IDE, inner dental epithelium; OD, odontoblast; ODE, outer dental epithelium; PEK, primary enamel knot; SEK, secondary enamel knot; SI, stratum intermedium; SR, stellate reticulum. Bar = 40 µm.

View larger version (102K): [in this window] [in a new window]   Figure 2. In vitro development of re-associations between dental mesenchyme and dissociated epithelial cells from first lower molars at ED13. The re-associations were cultured for 12 hrs (A), 24 hrs (B), 2 days (C), 4 days (D), 6 days (E), and 14 days (F). DE, dental epithelium; DM, dental mesenchyme; DP, dental papilla. Bar = 40 µm.

View larger version (25K): [in this window] [in a new window]   Figure 3. Scheme for the dissociation-re-association experiments. (1) About 60 tooth germs at the cap stage were dissociated by trypsin treatment, so that the dental mesenchyme could be separated from the dental epithelia. (2) The epithelia were further dissociated into single cells. (3) After filtration and centrifugation, the pellet containing mixed epithelial cells from 4 sources (SR, IDE, ODE, PEK) was fragmented and re-associated with dental mesenchyme. (4) These re-associations were cultured for 3 days to achieve the cap stage and illustrate the restoration of epithelial histogenesis.

Journal of Dental Research, Vol. 84, No. 6, 521-525 (2005)
DOI: 10.1177/154405910508400607


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