Dental pulp tissue engineering in a tooth-slice scaffold device

Immature necrotic teeth might benefit from the regeneration of dental pulp tissue which may allow for the completion of root formation. Objective: To characterize a tissue engineering based approach for the dental pulp that involves the use of dental pulp stem cells seeded in a tooth slice/scaffold device. Methods: Freshly extracted human molars were cut into 1 mm thick discs. The pulp tissue was removed and a biodegradable scaffold made with Poly-L-Lactic-Acid (PLLA) was cast in the root canal space of the tooth slice. Tooth slice/scaffold devices were treated with 10% EDTA and immediately seeded with 5x105 human dental pulp stem cells (DPSC, gift from S. Shi), 5x105 human dental pulp fibroblasts (DPF), or 2.5x105 DPSC + 2.5x105 DPF. DPSC were stained with CFDA SE® and DPF with SNARF®1 before seeding in the scaffold. In addition, tooth slices containing DPSC and/or DPF were implanted subcutaneously in the dorsum of severe combined immunodeficient (SCID) mice. Mice were euthanized after 21 days and samples analyzed by H&E staining and immunohistochemistry for Factor VIII to evaluate dental pulp vascularization. Results: After 5 days in culture, we observed that seeding of DPSC and DPF allowed for higher cell density than seeding of tooth slice/scaffolds with each cell type separately (p≤0.05). Confocal microscopy revealed some cellular processes extending into dentinal tubules. Histological evaluation of tooth slices implanted in vivo revealed the generation of a tissue with cellularity and architecture similar to a dental pulp. Conclusion: The approach presented here allowed for the engineering of a tissue with morphological features that resemble those of a normal dental pulp tissue. Supported by a grant from the Delta Dental Foundation and funds from the Department of Cariology, Restorative Sciences and Endodontics, University of Michigan School of Dentistry.