American Society for Investigative Pathology, June 2011, Vol 3, No. 2

Hot off the Press

Adipose Tissue-Derived Stem Cell in Vitro Differentiation in a Three-Dimensional Dental Bud Structure
Federico Ferro, Renza Spelat, Giuseppe Falini, Annarita Gallelli, Federica D'Aurizio, Elisa Puppato, Maura Pandolfi, Daniela Cesselli, Carlo Alberto Beltrami, Francesco Saverio Ambesi-Impiombato, Francesco Curcio

The American Journal of Pathology, Volume 178, Issue 5 , Pages 2299-2310, May 2011 DOI: 10.1016/j.ajpath.2011.01.055

A synopsis written by co-author and ASIP trainee Dr. Renza Spelat at the University of Udine, Udine, Italy Department of Pathology and Experimental and Clinical Medicine

Permanent tooth loss in humans is an irreversible condition. On the contrary, many animals regenerate teeth a few times during their life span (i.e. fish and reptiles), and others (i.e. rodents) have continuously growing teeth. However, it has been reported that rarely a few human subjects had a third generation of teeth. Also some people reported supra-numeral teeth eruption, evidencing the possibility to maintain the capacity to regenerate teeth. Moreover it was recently reported that teeth have been missing from birds for at least 60 million years. Although latent, the early signaling pathways involved in odontogenesis remain inducible also in avian species. Productive interactions between different tissues are the basic language needed for organs and tissue development. Previous experiments reported that single cell suspension, dissociated from the third molar tooth germs and seeded into biodegradable scaffolds, formed tooth structures containing dentin, odontoblast, a well-defined pulp chamber, putative Hertwig’s root sheath epithelia, and enamel organ. It was also demonstrated that stem cells, including Embryonic Stem cells (ES), neural stem cells, and Bone Marrow Stem cells (BMSCs), could be induced to reprogram into odontogenic fate to support tooth formation. These cells, when aggregated and recombined with the embryonic dental epithelium and oral-derived mesenchymal cells, which possess odontogenic potential, could respond to the inductive signal and initiate odontogenesis resulting in tooth formation. These studies support the idea that the odontogenic process can be initiated in stem cells from non-dental origins when proper odontogenic signals are provided, the availability of dental derived mesenchymal and epithelial cells being the limiting factor, which makes this approach interesting but not practicable for future dental regenerative therapies. In addition these results show that instead of forming one tooth with similar size and shape with respect to the physiological organ, small tooth crowns were formed throughout the implant. Also in these structures, tooth root development was only rudimentary, and consequently teeth did not erupt.

Our study was designed in the attempt to overcome the previous limitations and shortcomings, by providing a single cell type isolated from human adult adipose tissue, which may differentiate into a dental bud-like structure. We isolated a single population of Adipose tissue-derived Stem Cells (ASCs) that were aggregated and induced to transdifferentiate in vitro, obtaining a specific three-dimensional organization and a phenotype resembling a dental-bud. This three-dimensional simil-dental bud structure expressed ameloblastic and odontoblastic specific markers and moreover underwent mineralization and prismatic hydroxyapatite nano-crystals production, typically present in enamel, about 200nm length and dentin, about 50nm length. In addition this result was obtained without using any kind of structural matrix or scaffold to guide the process. The major achievement of our experimental approach is that ASC aggregates seem to reproduce in vitro the epithelial-mesenchymal interactions occurring in vivo, and show a three-dimensional structure comprising an intermediate layer similar to a basement membrane. Epithelial-mesenchymal interactions are the hallmark of tooth development and are involved in many other organ morphogenetic processes; these complicated, sequential, reciprocal interactions are mediated by the spatiotemporal expression of about 300 tooth-related genes and by the secretion of about 100 growth and transcription factors that are reiteratively used in regulatory loops. In conclusion our approach, recreating an in vitro simplified model of dental development, provides a convenient tool that may be used to better understand the reciprocal exchange of signals between epithelial and mesenchymal layers and possibly could be applied in vivo, to provide a new dental regenerative therapy.