Innovative dental stem cell-based research approaches: the future of dentistry

Over the past decade, the dental field has benefited from recent findings in stem cell biology and tissue engineering that led to the elaboration of novel ideas and concepts for the regeneration of dental tissues or entire new teeth. In particular, stem cell-based regenerative approaches are extremely promising since they aim at the full restoration of lost or damaged tissues, ensuring thus their functionality. These therapeutic approaches are already applied with success in clinics for the regeneration of other organs and consist of manipulation of stem cells and their administration to patients. Stem cells have the potential to self-renew and to give rise to a variety of cell types that ensure tissue repair and regeneration throughout life. During the last decades, several adult stem cell populations have been isolated from dental and periodontal tissues, characterized, and tested for their potential applications in regenerative dentistry. Here we briefly present the various stem cell-based treatment approaches and strategies that could be translated in dental practice and revolutionize dentistr

Roles of innervation in developing and regenerating orofacial tissues

The head is innervated by 12 cranial nerves (I-XII) that regulate its sensory and motor functions. Cranial nerves are composed of sensory, motor, or mixed neuronal populations. Sensory neurons perceive generally somatic sensations such as pressure, pain, and temperature. These neurons are also involved in smell, vision, taste, and hearing. Motor neurons ensure the motility of all muscles and glands. Innervation plays an essential role in the development of the various orofacial structures during embryogenesis. Hypoplastic cranial nerves often lead to abnormal development of their target organs and tissues. For example, Möbius syndrome is a congenital disease characterized by defective innervation (i.e., abducens (VI) and facial (VII) nerves), deafness, tooth anomalies, and cleft palate. Hence, it is obvious that the peripheral nervous system is needed for both development and function of orofacial structures. Nerves have a limited capacity to regenerate. However, neural stem cells, which could be used as sources for neural tissue maintenance and repair, have been found in adult neuronal tissues. Similarly, various adult stem cell populations have been isolated from almost all organs of the human body. Stem cells are tightly regulated by their microenvironment, the stem cell niche. Deregulation of adult stem cell behavior results in the development of pathologies such as tumor formation or early tissue senescence. It is thus essential to understand the factors that regulate the functions and maintenance of stem cells. Yet, the potential importance of innervation in the regulation of stem cells and/or their niches in most organs and tissues is largely unexplored. This review focuses on the potential role of innervation in the development and homeostasis of orofacial structures and discusses its possible association with stem cell populations during tissue repair

Extracellular matrix remodelling in dental pulp tissue of carious human teeth through the prism of single-cell RNA sequencing

Carious lesions are bacteria-caused destructions of the mineralised dental tissues, marked by the simultaneous activation of immune responses and regenerative events within the soft dental pulp tissue. While major molecular players in tooth decay have been uncovered during the past years, a detailed map of the molecular and cellular landscape of the diseased pulp is still missing. In this study we used single-cell RNA sequencing analysis, supplemented with immunostaining, to generate a comprehensive single-cell atlas of the pulp of carious human teeth. Our data demonstrated modifications in the various cell clusters within the pulp of carious teeth, such as immune cells, mesenchymal stem cells (MSC) and fibroblasts, when compared to the pulp of healthy human teeth. Active immune response in the carious pulp tissue is accompanied by specific changes in the fibroblast and MSC clusters. These changes include the upregulation of genes encoding extracellular matrix (ECM) components, including COL1A1 and Fibronectin (FN1), and the enrichment of the fibroblast cluster with myofibroblasts. The incremental changes in the ECM composition of carious pulp tissues were further confirmed by immunostaining analyses. Assessment of the Fibronectin fibres under mechanical strain conditions showed a significant tension reduction in carious pulp tissues, compared to the healthy ones. The present data demonstrate molecular, cellular and biomechanical alterations in the pulp of human carious teeth, indicative of extensive ECM remodelling, reminiscent of fibrosis observed in other organs. This comprehensive atlas of carious human teeth can facilitate future studies of dental pathologies and enable comparative analyses across diseased organs

Notch Signaling Pathway in Tooth Shape Variations throughout Evolution

Evolutionary changes in vertebrates are linked to genetic alterations that often affect tooth crown shape, which is a criterion of speciation events. The Notch pathway is highly conserved between species and controls morphogenetic processes in most developing organs, including teeth. Epithelial loss of the Notch-ligand Jagged1 in developing mouse molars affects the location, size and interconnections of their cusps that lead to minor tooth crown shape modifications convergent to those observed along Muridae evolution. RNA sequencing analysis revealed that these alterations are due to the modulation of more than 2000 genes and that Notch signaling is a hub for significant morphogenetic networks, such as Wnts and Fibroblast Growth Factors. The modeling of these tooth crown changes in mutant mice, via a three-dimensional metamorphosis approach, allowed prediction of how Jagged1-associated mutations in humans could affect the morphology of their teeth. These results shed new light on Notch/Jagged1-mediated signaling as one of the crucial components for dental variations in evolution

Early determination of the periodontal domain by the wnt-antagonist Frzb/Sfrp3

Odontogenesis results from the continuous and reciprocal interaction between cells of the oral epithelium and cranial neural crest-derived mesenchyme. The canonical Wnt signaling pathway plays a fundamental role in mediating these interactions from the earliest stages of tooth development. Here we analyze by in situ hybridization the expression patterns of the extracellular Wnt antagonist Frzb/Sfrp3. Although Frzb is expressed in dental mesenchymal cells from the earliest stages of odontogenesis, its expression is absent from a tiny population of mesenchymal cells immediately adjacent to the invaginating dental epithelium. Cell proliferation studies using BrdU showed that the Frzb expressing and Frzb non-expressing cell populations display different proliferative behavior during the initial stages of odontogenesis. DiI-mediated cell-fate tracing studies demonstrated that the Frzb expressing cells contribute to the formation of the dental follicle, the future periodontium. In contrast, the Frzb non-expressing cells give rise to the dental pulp. The present results indicate that Frzb is discriminating the presumptive periodontal territory from the rest of the dental mesenchyme from the very beginning of odontogenesis, where it might act as a barrier for the diffusion of Wnt molecules, thus regulating the activation of Wnt-dependent transcription within dental tissues