Continuous Tooth Replacement Research Articles

Sox2 marks epithelial competence to generate teeth in mammals and reptiles

Tooth renewal is initiated from epithelium associated with existing teeth. The development of new teeth requires dental epithelial cells that have competence for tooth formation, but specific marker genes for these cells have not been identified. Here, we analyzed expression patterns of the transcription factor Sox2 in two different modes of successional tooth formation: tooth replacement and serial addition of primary teeth. We observed specific Sox2 expression in the dental lamina that gives rise to successional teeth in mammals with one round of tooth replacement as well as in reptiles with continuous tooth replacement. Sox2 was also expressed in the dental lamina during serial addition of mammalian molars, and genetic lineage tracing indicated that Sox2(+) cells of the first molar give rise to the epithelial cell lineages of the second and third molars. Moreover, conditional deletion of Sox2 resulted in hyperplastic epithelium in the forming posterior molars. Our results indicate that the Sox2(+) dental epithelium has competence for successional tooth formation and that Sox2 regulates the progenitor state of dental epithelial cells. The findings imply that the function of Sox2 has been conserved during evolution and that tooth replacement and serial addition of primary teeth represent variations of the same developmental process. The expression patterns of Sox2 support the hypothesis that dormant capacity for continuous tooth renewal exists in mammals.

Dual BrdU-PCNA immunodetection of proliferative cells in dental and orofacial tissues of teleosts

We have developed a protocol for simultaneous detection of BrdU and PCNA on paraffin sections of teleost tissues and have tested it successfully on dental and orofacial tissues of Atlantic salmon (Salmo salar) and zebrafish (Danio rerio). By combining decalcification of the tissue, appropriate antigen retrieval steps, and high quality tissue sections, this protocol allows to study the instantaneous pattern of proliferating cells, as well as cell tracing in large specimens. The technique is therefore highly suitable to study skeletal and dental tissues, tissues that are traditionally not easily accessible to molecular and cellular techniques. Dual BrdU-PCNA immunostaining may serve as a useful tool in experimental studies on fish developmental or regenerative processes such as tissue turnover in orofacial epithelia, continuous tooth replacement, and in the identification of putative stem cells for which the cycling state of BrdU label retaining cells needs to be determined.

Replacing the first-generation dentition in pufferfish with a unique beak.

Teleost fishes comprise approximately half of all living vertebrates. The extreme range of diversity in teleosts is remarkable, especially, extensive morphological variation in their jaws and dentition. Some of the most unusual dentitions are found among members of the highly derived teleost order Tetraodontiformes, which includes triggerfishes, boxfishes, ocean sunfishes, and pufferfishes. Adult pufferfishes (Tetraodontidae) exhibit a distinctive parrot-like beaked jaw, forming a cutting edge, unlike in any other group of teleosts. Here we show that despite novelty in the structure and development of this "beak," it is initiated by formation of separate first-generation teeth that line the embryonic pufferfish jaw, with timing of development and gene expression patterns conserved from the last common ancestor of osteichthyans. Most of these first-generation larval teeth are lost in development. Continuous tooth replacement proceeds in only four parasymphyseal teeth, as sequentially stacked, multigenerational, jaw-length dentine bands, before development of the functional beak. These data suggest that dental novelties, such as the pufferfish beak, can develop later in ontogeny through modified continuous tooth addition and replacement. We conclude that even highly derived morphological structures like the pufferfish beak form via a conserved developmental bauplan capable of modification during ontogeny by subtle respecification of the developmental module.

Molecular genetics of supernumerary tooth formation

Despite advances in the knowledge of tooth morphogenesis and differentiation, relatively little is known about the aetiology and molecular mechanisms underlying supernumerary tooth formation. A small number of supernumerary teeth may be a common developmental dental anomaly, while multiple supernumerary teeth usually have a genetic component and they are sometimes thought to represent a partial third dentition in humans. Mice, which are commonly used for studying tooth development, only exhibit one dentition, with very few mouse models exhibiting supernumerary teeth similar to those in humans. Inactivation of Apc or forced activation of Wnt/β(catenin signalling results in multiple supernumerary tooth formation in both humans and in mice, but the key genes in these pathways are not very clear. Analysis of other model systems with continuous tooth replacement or secondary tooth formation, such as fish, snake, lizard, and ferret, is providing insights into the molecular and cellular mechanisms underlying succesional tooth development, and will assist in the studies on supernumerary tooth formation in humans. This information, together with the advances in stem cell biology and tissue engineering, will pave ways for the tooth regeneration and tooth bioengineering.

Reptilian tooth development

Dental patterns in vertebrates range from absence of teeth to multiple sets of teeth that are replaced throughout life. Despite this great variation, most of our understanding of tooth development is derived from studies on just a few model organisms. Here we introduce the reptile as an excellent model in which to study the molecular basis for early dental specification and, most importantly, for tooth replacement. We review recent snake studies that highlight the conserved role of Shh in marking the position of the odontogenic band. The distinctive molecular patterning of the dental lamina in the labial-lingual and oral-aboral axes is reviewed. We explain how these early signals help to specify the tooth-forming and non-tooth forming sides of the dental lamina as well as the presumptive successional lamina. Next, the simple architecture of the reptilian enamel organ is contrasted with the more complex, mammalian tooth bud and we discuss whether or not there is an enamel knot in reptilian teeth. The role of the successional lamina during tooth replacement in squamate reptiles is reviewed and we speculate on the possible formation of a vestigial, post-permanent dentition in mammals. In support of these ideas, we present data on agamid teeth in which development of a third generation is arrested. We suggest that in diphyodont mammals, similar mechanisms may be involved in reducing tooth replacement capacity. Finally, we review the location of label-retaining cells and suggest ways in which these putative dental epithelial stem cells contribute to continuous tooth replacement.

Unique and shared gene expression patterns in Atlantic salmon (Salmo salar) tooth development

To validate the use of Atlantic salmon (Salmo salar L.) as a model species in research on the mechanism of continuous tooth replacement, we have started to collect data on the molecular control underlying tooth formation in this species. This study reports expression patterns in the lower jaw dentition of a number of key regulatory genes such as bmp2, bmp4, and sox9 and structural genes such as col1alpha 1 and osteocalcin (= bgp, Bone Gla Protein) by means of in situ hybridization using salmon-specific, digoxygenin-labeled antisense riboprobes. We compare expression of these genes to that in other skeletogenic cells in the lower jaw (osteoblasts, chondroblasts, and chondrocytes). Our studies reveal both expression patterns that are in accordance to studies on mammalian tooth development and patterns that are specific to salmon, or teleosts. The epithelial expression of sox9 and a shift of the expression of bmp2 from epithelium to mesenchyme have also been observed during mammalian tooth development. Different from previous reports are the expressions of col1alpha 1 and osteocalcin. In contrast to what has been reported for zebrafish, osteocalcin is not expressed in odontoblasts, nor in the osteoblasts involved in the attachment of the teeth. At the lower jaw, osteocalcin is expressed in mature and/or resting osteoblasts only. As expected, col1alpha 1 is expressed in odontoblasts. Surprisingly, it is also strongly expressed in the inner dental epithelium, representing the first report of ameloblast involvement in collagen type I transcription. Whether the collagen is translated and secreted into the enameloid remains to be demonstrated.

Continuous tooth replacement: the possible involvement of epithelial stem cells

Epithelial stem cells have been identified in integumental structures such as hairs and continuously growing teeth of various rodents, and in the gut. Here we propose the involvement of epithelial stem cells in the continuous tooth replacement that characterizes non-mammalian vertebrates, as exemplified by the zebrafish. Arguments are based on morphological observations of tooth renewal in the zebrafish and on the similarities between molecular control of hair and tooth formation. Dissection of the molecular cascades underlying the regulation of the epithelial stem cell niche might open perspectives for new regenerative treatment strategies in clinical dentistry.