Our current knowledge on tooth development derives primarily from studies in mice. Very little is known about gene expression and function during human odontogenesis. Sonic Hedgehog (SHH) signaling has been demonstrated to play crucial roles in the development of multiple organs in mice, including the tooth. However, if SHH signaling molecules are expressed and function in the developing human embryonic tooth remain unknown.We conducted microarray assay to reveal the expression profile of SHH signaling pathway molecules. We then used in situ hybridization to validate and reveal spatial and temporal expression patterns of a number of selected molecules, including SHH, PTC1, SMO, GLI1, GLI2, and GLI3, in the developing human embryonic tooth germs, and compared them with that in mice. We found that all these genes exhibit similar but slightly distinct expression patterns in the human and mouse tooth germ at the cap and bell stages.Our results demonstrate the operation of active SHH signaling in the developing human tooth and suggest a conserved function of SHH signaling pathway during human odontogenesis.

Children often receive inferior alveolar nerve blocks (IANBs) when their third molars are just beginning to develop. The location of the third-molar follicle is close to where the needle penetrates during an IANB. The authors examined the possible association between IANBs and missing third molars.The authors examined 439 potential sites of third-molar development for evidence of third-molar follicles on panoramic radiographs of randomly selected children 7 years and older. The authors conducted a statistical comparison of the incidence of missing third-molar follicles in a control group of children who had no history of receiving IANBs with children in a test group who had a definitive history of receiving IANBs by means of generalized estimating equations.The authors found a statistically significant greater incidence of missing third-molar follicles in mandibular quadrants that had a definitive history of receiving IANBs compared with mandibular quadrants that had no history of receiving IANB.IANBs administered to young children when the third-molar tooth bud is immature may stop third-molar development. Owing to the significant clinical implications, further research is needed to verify these results.Dentists inadvertently may be stopping the development of third molars when administering IANBs to children.

To characterize the subtypes of ameloblastoma by differentiation markers.Expression of 9 major acidic epithelial keratins was immunohistochemically examined in 28 ameloblastomas.Keratin 15 (K15) expression patterns corresponded to histological variants: follicular, plexiform and acanthomatous. Tumor nests comprising K15-expressing basal cells mimicked oral epithelium or dental lamina, and tumor nests comprising K15-negative basal cells mimicked outer enamel epithelium. Keratin 19 (K19) was consistently expressed in solid/multicystic ameloblastoma and unicystic ameloblastoma, while peripheral ameloblastoma and desmoplastic ameloblastoma contained K19-negative cells.The 4 current subtypes had unvaried expression patterns within each group. However, they could be divided into 2 groups by K19 expression pattern: solid/multicystic and unicystic versus extraosseous/peripheral and desmoplastic. K15 expression pattern represented various types of differentiation for tumor nests mimicking tooth germ and oral epithelium. The results clarify the homogeneity and heterogeneity of ameloblastoma cell lineage and differentiation.

Bone-marrow-derived mesenchymal stem cells (MSCs) demonstrate neuro-protective effects in several disease models. By producing growth-factors, cytokines and chemokines, they promote survival of neurons in damaged brain areas. Alternative MSC sources, such as human tooth germ stem cells (hTGSCs), have been investigated for their neuro-protective properties. They ameliorate effects of neuro-toxic agents by paracrine mechanisms, however these secreted bio-active molecules are not yet characterized. Therefore, the current study aimed to provide a detailed analysis of the secretome of hTGSCs. Brain cells were exposed to various toxic materials, including Alzheimer's β-amyloid peptide (β-AP) and 6-hydroxy-dopamine (6-OHDA). When co-cultured with hTGSCs, the activity of a number of anti-oxidant enzymes (catalase, glutathione-s-transferase, glutathione-peroxidase, superoxide-dismutase) was increased and neuronal death/apoptosis was subsequently reduced. The composition of the secreted bio-active materials is influenced by various pre-existing factors such as oxygen and glucose deprivation and the age of cells (passage number). This report reveals for the first time that the neuro-protective secretome of hTGSCs and the micro-environment of cells have a mutual and dynamic impact on one another.

Stem cells from the dental apical papilla (SCAPs) can be induced to differentiate along both osteoblast and odontoblast lineages. However, little knowledge is available concerning their differentiation efficiency in osteogenic media containing additional KH2 PO4 .Stem cells from the dental apical papilla were isolated from apical papillae of immature third molars and treated with two kinds of mineralization-inducing media, MM1 and MM2, differing in KH2 PO4 concentration. Proliferation and osteo/odontogenic differentiation capacity of MM1/MM2-treated SCAPs were investigated and compared both in vitro and in vivo.Cell counting and flow cytometry demonstrated that MM2 containing 1.8 mm additional KH2 PO4 significantly enhanced proliferative potential of SCAPs, compared to MM1. Osteo/odontogenic capacity of SCAPs was much better in MM2 medium than in MM1, as indicated by elevated alkaline phosphatase activity, increased calcium deposition and upregulated expression of osteo/odontoblast-specific genes/proteins (for example, runt-related transcription factor 2, osterix, osteocalcin, dentin sialoprotein, and dentin sialophosphoprotein). In vivo transplantation findings proved that SCAPs in MM2 group generated more mineralized tissues, and presented higher expression of osteo/odontoblast-specific proteins (osteocalcin and dentin sialoprotein) than those in the MM1 group.Mineralization-inducing media supplemented with 1.8 mm additional KH2 PO4 significantly enhanced cell proliferation and improved differentiation capacity of SCAPs along osteo/odontogenic cell lineages, compared to counterparts lacking additional KH2 PO4 .

Peripheral odontoma is rare, and only two cases of congenital peripheral odontoma have been reported. Congenital oral fibroma is also rare. We describe a unique case of congenital peripheral developing odontoma accompanied by congenital teratomatous fibroma in an infant. Both tumors were difficult to detect on radiography. Two small masses were seen in the median anterior portion of the palatal mucosa of a 9-month-old boy. The masses had been present since birth and were surgically removed at age 28 months, when one of the masses had grown to a diameter of 8 mm. Histopathologic examination showed a fibrous lesion and a tooth germ-like rounded lesion composed of dental papilla, enamel organ, dentin, and cementum. Although congenital odontoma is rare, it should be considered when selecting appropriate treatment, as early radiographic detection is difficult.

A symphyseal mandibular facial cleft (Tessier 30) is a rare congenital abnormality. Nonunion of bone in the symphysis of the mandible, median cleft of the lower lip and tongue adherence in the midline to the floor of the mouth are the principle features of the anomaly. A satisfactory result can be achieved with early surgical correction of both hard and soft tissue defects in the age of 6 months. More than 3 years follow-up shows total bony fusion of the symphysis and normal lower lip function. There were only left deciduous central incisor and the ipsilateral tooth bud of the permanent central incisor missing.

The aim of this study was to investigate the behavior of rat incisor tissues during the inhibition of tooth eruption. Twenty Sprague-Dawley rats were used in this study, and incisor eruption was inhibited by a screw pin. Animals were sacrificed 1, 3, 7 and 14 days after the start of the experiment. Cross-sections at the mesial point of the mandibular first molar and sagittal sections of the mandibular tooth germ area were examined using immunohistochemical and immunofluorescence methods. For morphometric analysis, numbers of TRAP-positive cells were calculated against the total number of cells. In cross-sections from the experimental group, dentin was thickened and pulp tissue was constricted day by day. On days 1, 3 and 7, nestin-positive cells were observed in all odontoblast cell bodies and processes, while on day 14 fewer nestin-positive cells were seen than in the control group. On day 14, the mesial area of the periodontal ligament was constricted and the number of TRAP-positive cells in the mesial area was significantly higher than in the control group. In sagittal sections, enamel formation was found to be increased on days 7 and 14. Furthermore, in the enamel matrix amelogenin was expressed more strongly than in the control group. PCNA-positive cells were significantly increased in cells of the tooth germ compared with the control group. These results suggest that inhibition of tooth eruption accelerates the apical elongation with resorption of the mesial area of the alveolar bone and stimulates cell proliferation with thickened enamel towards the apical end.

Small ubiquitin-related modifier (SUMO) conjugation (SUMOylation) is a post-translational modification involved in various cellular processes including the regulation of transcription factors. In this study, to analyze the involvement of SUMOylation in odontoblast differentiation, we examined the immunohistochemical localization of SUMO-1, SUMO-2/3, and Osterix during rat tooth development. At the bud and cap stages, localization of SUMOs and Osterix was hardly detected in the dental mesenchyme. At the bell stage, odontoblasts just beginning dentin matrix secretion and preodontoblasts near these odontoblasts showed intense immunoreactivity for these molecules. However, after the root-formation stage, these immunoreactivities in the odontoblasts decreased in intensity. Next, to examine whether the SUMOylation participates in dentin regeneration, we evaluated the distribution of SUMOs and Osterix in the dental pulp after cavity preparation. In the coronal pulp chamber of an untreated rat molar, odontoblasts and pulp cells showed no immunoreactivity. At 4 days after cavity preparation, positive cells for SUMOs and Osterix appeared on the surface of the dentin beneath the cavity. Odontoblast-like cells forming reparative dentin were immunopositive for SUMOs and Osterix at 1 week, whereas these immunoreactivities disappeared after 8 weeks. Additionally, we further analyzed the capacity of SUMO-1 to bind Osterix by performing an immunoprecipitation assay using C2C12 cells, and showed that Osterix could undergo SUMOylation. These results suggest that SUMOylation might regulate the transcriptional activity of Osterix in odontoblast lineage cells, and thus play important roles in odontoblast differentiation and regeneration.

The tooth root is an important part of the tooth that works together with the surrounding periodontium to maintain the tooth in the alveolar socket. The root develops after crown morphogenesis. While the molecular and cellular mechanisms of early tooth development and crown morphogenesis have been extensively studied, little is known about the molecular mechanisms controlling tooth root formation. Here, we show that β-catenin is strongly expressed in odontoblast-lineage cells and is required for root formation. Tissue-specific inactivation of β-catenin in developing odontoblasts produced molars lacking roots and aberrantly thin incisors. At the beginning of root formation in the mutant molars, the cervical loop epithelium extended apically to form Hertwig's epithelial root sheath (HERS), but root odontoblast differentiation was disrupted and followed by the loss of some HERS inner layer cells. However, the outer layer of the HERS extended without the root, and the mutant molars finally erupted. The periodontal tissues extensively invaded the dental pulp. These results indicate that there is a cell-autonomous requirement for Wnt/β-catenin signaling in the dental mesenchyme for root formation.