Whole-Tooth Regeneration by Allogeneic Cell Reassociation in Pig Jawbone

Although mouse models of epithelial–mesenchymal interaction-based whole-tooth regeneration have been generated, whether this strategy can be applied to humans remains unknown and preclinical investigations using a large animal model are required. In this study, we describe the development of feasible methods for whole-tooth regeneration in a swine model. We characterized the odontogenesis of dissociated single cells from early-stage tooth germs from miniature pigs (minipigs) in vitro by tracking the morphogenesis of reassociated cell pellets in an organ culture system. Despite loss of positional information, reassociated individual dental epithelial and mesenchymal cells retained their odontogenic potential and continued to recapitulate typical dental histogenesis and morphogenesis. Based on the developmental biology of natural teeth, we allotransplanted reassociated tooth germs into the jawbones of minipigs and manipulated whole-tooth regeneration by mimicking the process of tooth development. To overcome the challenge of alloimmune rejection, bone marrow mesenchymal stem cells were systemically infused, along with local aspirin administration, during allotransplantation. Radiographic data confirmed the survival and growth of the explants in minipig jawbones. Histological analysis revealed that the explants formed well-developed tooth structures and contained normally arranged dental components, including organized dentin, cementum, periodontal membrane, dental pulp, vasculature, and nervous tissue, closely resembling those of natural teeth. In conclusion, this study achieved orthotopic whole-tooth regeneration and development in a large mammal using a cell reassociation approach, supporting the potential for regeneration of human teeth in situ. The methods developed in this study to manipulate pig tooth germ cells in vitro and in vivo provide a reference for studying whole-tooth regeneration and tooth development in large animals. Of importance, compared with conventional ectopic tooth regeneration, conducted in the omentum, subcutaneous tissues, or kidney capsule (among other locations) with low with immune reactivity in rodent models, this study achieved orthotopic regeneration and development of whole teeth in a large mammal, representing a large stride toward the realization of tooth regenerative therapy for humans with missing teeth.