Three dimensional dental epithelial-mesenchymal constructs of predetermined size and shape for tooth regeneration.

While it is known that precise dental epithelial-mesenchymal (DE-DM) cell interactions provide critical functions in tooth development, reliable methods to establish proper DE-DM cell interactions for tooth regeneration have yet to be established. To address this challenge, and to generate bioengineered teeth of predetermined size and shape, in this study, we characterize three dimensional (3D) pre-fabricated DE-DM cell constructs. Human dental pulp cell seeded Collagen gel layers were co-cultured with porcine DE cells suspended in Growth Factor Reduced (GFR) Matrigel. The resulting 3D DE-DM cell layers were cultured in vitro, or implanted and grown subcutaneously in vivo in nude rats. Molecular, histological and immunohistochemical (IHC) analyses of harvested implants revealed organized DE-DM cell interactions, the induced expression of dental tissue-specific markers Amelogenin (AM) and Dentin Sialophosphoprotein (DSPP), and basement membrane markers Laminin 5 and collagen IV, and irregular mineralized tissue formation after 4 weeks. We anticipate that these studies will facilitate the eventual establishment of reliable methods to elaborate dental tissues, and full sized teeth of specified sized and shape.