Organotypic 3D Cellular Models Mimicking the Epithelio-Ectomesenchymal Bilayer During Odontogenesis

Odontogenesis, the intricate process of tooth development, involves complex interactions between oral ectoderm epithelial cells and ectomesenchymal cells derived from the cephalic neural crest, regulated by major signaling pathways. Dental developmental anomalies provide valuable insights for the clinical diagnosis of rare diseases. More than 30% of patients with rare diseases who undergo molecular analysis suffer from diagnostic errancy. In the search for up-to-date technologies and methods to study the pathophysiology of new candidate genetic variants, causing tooth mineralized tissue anomalies, we have developed an original model of tooth organoids with human or mouse cell lines of ameloblast-like cells and odontoblasts derived from the pulp. This <i>in vitro</i> 3D cellular model reproducing the two main compartments of the bell stage of tooth development between ameloblasts and odontoblasts, specific to enamel and dentin morphogenesis, respectively, mimics the epithelial-mesenchymal interactions during the dental bell stage of tooth morphogenesis and will facilitate the study of enamel and dentin genetic anomalies, allowing the functional validation of newly identified mutations (variants of uncertain significance or new candidate genes). Impact Statement We have developed and characterized innovative 3D bilayer organotypic cellular models with ameloblast-like cells and odontoblast-like cells (both murine and human) reproducing some of the interactions during the bell stage of odontogenesis. This is a first model of such type, considering human lineages and not only primary dental cells, with high reproducibility, and including recently developed polymeric poly-(lactic-coglycolic acid) microscaffolds for increasing cell survival and favoring cell differentiation. This work opens new perspectives to the study of tooth morphogenesis <i>in vitro</i> and associated genetic disorders, either for fundamental studies or for the development of new diagnostic tools of structure for dental rare diseases, such as amelogenesis imperfecta or dentinogenesis imperfecta.