The initiation knot is a signaling center required for molar tooth development.

Signaling centers, or organizers, regulate many aspects of embryonic morphogenesis. In the mammalian molar tooth, reiterative signaling in specialized centers called enamel knots (EKs) determines tooth patterning. Preceding the primary EK, transient epithelial thickening appears, the significance of which remains debated. Using tissue confocal fluorescence imaging with laser ablation experiments, we show that this transient thickening is an earlier signaling center, the molar initiation knot (IK), that is required for the progression of tooth development. IK cell dynamics demonstrate the hallmarks of a signaling center: cell cycle exit, condensation and eventual silencing through apoptosis. IK initiation and maturation are defined by the juxtaposition of cells with high Wnt activity to Shh-expressing non-proliferating cells, the combination of which drives the growth of the tooth bud, leading to the formation of the primary EK as an independent cell cluster. Overall, the whole development of the tooth, from initiation to patterning, is driven by the iterative use of signaling centers.

Sox2 maintains epithelial cell proliferation in the successional dental lamina.

OBJECTIVES: The successional dental lamina is the distinctive structure on the lingual side of the vertebrate tooth germ. The aim of this study was to investigate the relationship among Sox2, Claudin10 and laminin5 and the role of Sox2 in successional dental lamina proliferation during vertebrate tooth development. MATERIALS AND METHODS: To understand the successional dental lamina, two types of successional tooth formation, that in geckos (with multiple rounds of tooth generation) and that in mice (with only one round of tooth generation), were analysed. RESULTS: Unique coexpression patterns of Sox2 and Claudin10 expression were compared in the successional dental lamina from the cap stage to the late bell stage in the mouse tooth germ and in juvenile gecko teeth to support continuous tooth replacement. Furthermore, Laminin5 expression was shown in the cap stage and decreased after the bell stage. Upon comparing the epithelial cell cycles and cell proliferation in successional dental lamina regions between mouse and gecko molars using BrdU and IdU staining and pulse-chase methods, distinctive patterns of continuous expression were revealed. Moreover, Sox2 overexpression with a lentiviral system resulted in hyperplastic dental epithelium in mouse molars. CONCLUSIONS: Our findings indicate that the regulation of Sox2 in dental lamina proliferation is fundamental to the successional dental lamina in both species.

Role of osteoprotegerin in the regulation of dental epithelial­mesenchymal signaling during tooth development.

Dental epithelial­mesenchymal signaling is crucial for tooth development, but the detailed mechanism is not fully understood. Using microarray analysis, it was revealed that the expression of osteoprotegerin, an important factor regulating bone remodeling, significantly increased after removal of the dental epithelium. Immunohistochemical staining revealed that osteoprotegerin expression within the dental mesenchyme was quite low during the prenatal period, but significantly increased after birth. To investigate the influence of osteoprotegerin upon tooth development, first­molar tooth germs from embryonic day 14.5 (E14.5) Chinese Kunming mice were treated with different concentrations of osteoprotegerin. It was revealed that osteoprotegerin could inhibit the expression of odontogenic markers while promoting the expression of osteogenic markers, thereby disrupting tooth morphogenesis. These findings were further supported by in vitro and in vivo cultures. Finally, quantitative reverse transcription­polymerase chain reaction and immunofluorescence studies revealed that, after osteoprotegerin treatment, the activity of the wingless/integrated (Wnt)/ß­catenin pathway increased, indicating that increased osteoprotegerin expression in prenatal tooth development could lead to uncontrolled upregulation of the Wnt/ß­catenin pathway.

Wnt/ß-catenin signaling, which is activated in odontomas, reduces Sema3A expression to regulate odontogenic epithelial cell proliferation and tooth germ development.

Odontomas, developmental anomalies of tooth germ, frequently occur in familial adenomatous polyposis patients with activated Wnt/ß-catenin signaling. However, roles of Wnt/ß-catenin signaling in odontomas or odontogenic cells are unclear. Herein, we investigated ß-catenin expression in odontomas and functions of Wnt/ß-catenin signaling in tooth germ development. ß-catenin frequently accumulated in nucleus and/or cellular cytoplasm of odontogenic epithelial cells in human odontoma specimens, immunohistochemically. Wnt/ß-catenin signaling inhibited odontogenic epithelial cell proliferation in both cell line and tooth germ development, while inducing immature epithelial bud formation. We identified Semaphorin 3A (Sema3A) as a downstream molecule of Wnt/ß-catenin signaling and showed that Wnt/ß-catenin signaling-dependent reduction of Sema3A expression resulted in suppressed odontogenic epithelial cell proliferation. Sema3A expression is required in appropriate epithelial budding morphogenesis. These results suggest that Wnt/ß-catenin signaling negatively regulates odontogenic epithelial cell proliferation and tooth germ development through decreased-Sema3A expression, and aberrant activation of Wnt/ß-catenin signaling may associate with odontoma formation.

Establishment of tooth blood supply and innervation is developmentally regulated and takes place through differential patterning processes.

Teeth are richly supported by blood vessels and peripheral nerves. The aim of this study was to describe in detail the developmental time-course and localization of blood vessels during early tooth formation and to compare that to innervation, as well as to address the putative role of vascular endothelial growth factor (VEGF), which is an essential regulator of vasculature development, in this process. The localization of blood vessels and neurites was compared using double immunofluorescence staining on sections at consecutive stages of the embryonic (E) and postnatal (PN) mandibular first molar tooth germ (E11-PN7). Cellular mRNA expression domains of VEGF and its signaling receptor VEGFR2 were studied using sectional radioactive in situ hybridization. Expression of VEGF mRNA and the encoded protein were studied by RT-PCR and western blot analysis, respectively, in the cap and early bell stage tooth germs, respectively. VEGFR2 was immunolocalized on tooth tissue sections. Smooth muscle cells were investigated by anti-alpha smooth muscle actin (αSMA) antibodies. VEGF showed developmentally regulated epithelial and mesenchymal mRNA expression domains including the enamel knot signaling centers that correlated with the growth and navigation of the blood vessels expressing Vegfr2 and VEGFR2 to the dental papilla and enamel organ. Developing blood vessels were present in the jaw mesenchyme including the presumptive dental mesenchyme before the appearance of the epithelial dental placode and dental neurites. Similarly, formation of a blood vessel plexus around the bud stage tooth germ and ingrowth of vessels into dental papilla at E14 preceded ingrowth of neurites. Subsequently, pioneer blood vessels in the dental papilla started to receive smooth muscle coverage at the early embryonic bell stage. Establishment and patterning of the blood vessels and nerves during tooth formation are developmentally regulated, stepwise processes that likely involve differential patterning mechanisms. Development of tooth vascular supply is proposed to be regulated by local, tooth-specific regulation by epithelial-mesenchymal tissue interactions and involving tooth target expressed VEGF signaling. Further investigations on tooth vascular development by local VEGF signaling, as well as how tooth innervation and development of blood vessels are integrated with advancing tooth organ formation by local signaling mechanisms, are warranted.

Modeling Edar expression reveals the hidden dynamics of tooth signaling center patterning.

When patterns are set during embryogenesis, it is expected that they are straightly established rather than subsequently modified. The patterning of the three mouse molars is, however, far from straight, likely as a result of mouse evolutionary history. The first-formed tooth signaling centers, called MS and R2, disappear before driving tooth formation and are thought to be vestiges of the premolars found in mouse ancestors. Moreover, the mature signaling center of the first molar (M1) is formed from the fusion of two signaling centers (R2 and early M1). Here, we report that broad activation of Edar expression precedes its spatial restriction to tooth signaling centers. This reveals a hidden two-step patterning process for tooth signaling centers, which was modeled with a single activator-inhibitor pair subject to reaction-diffusion (RD). The study of Edar expression also unveiled successive phases of signaling center formation, erasing, recovering, and fusion. Our model, in which R2 signaling center is not intrinsically defective but erased by the broad activation preceding M1 signaling center formation, predicted the surprising rescue of R2 in Edar mutant mice, where activation is reduced. The importance of this R2-M1 interaction was confirmed by ex vivo cultures showing that R2 is capable of forming a tooth. Finally, by introducing chemotaxis as a secondary process to RD, we recapitulated in silico different conditions in which R2 and M1 centers fuse or not. In conclusion, pattern formation in the mouse molar field relies on basic mechanisms whose dynamics produce embryonic patterns that are plastic objects rather than fixed end points.

Expression of Phosphate Transporters during Dental Mineralization.

The importance of phosphate (Pi) as an essential component of hydroxyapatite crystals suggests a key role for membrane proteins controlling Pi uptake during mineralization in the tooth. To clarify the involvement of the currently known Pi transporters (Slc17a1, Slc34a1, Slc34a2, Slc34a3, Slc20a1, Slc20a2, and Xpr1) during tooth development and mineralization, we determined their spatiotemporal expression in murine tooth germs from embryonic day 14.5 to postnatal day 15 and in human dental samples from Nolla stages 6 to 9. Using real-time polymerase chain reaction, in situ hybridization, immunohistochemistry, and X-gal staining, we showed that the expression of Slc17a1, Slc34a1, and Slc34a3 in tooth germs from C57BL/6 mice were very low. In contrast, Slc34a2, Slc20a1, Slc20a2, and Xpr1 were highly expressed, mostly during the postnatal stages. The expression of Slc20a2 was 2- to 10-fold higher than the other transporters. Comparable results were obtained in human tooth germs. In mice, Slc34a2 and Slc20a1 were predominantly expressed in ameloblasts but not odontoblasts, while Slc20a2 was detected neither in ameloblasts nor in odontoblasts. Rather, Slc20a2 was highly expressed in the stratum intermedium and the subodontoblastic cell layer. Although Slc20a2 knockout mice did not show enamel defects, mutant mice showed a disrupted dentin mineralization, displaying unmerged calcospherites at the mineralization front. This latter phenotypical finding raises the possibility that Slc20a2 may play an indirect role in regulating the extracellular Pi availability for mineralizing cells rather than a direct role in mediating Pi transport through mineralizing plasma cell membranes. By documenting the spatiotemporal expression of Pi transporters in the tooth, our data support the possibility that the currently known Pi transporters may be dispensable for the initiation of dental mineralization and may rather be involved later during the tooth mineralization scheme.

Accelerated dentinogenesis by inhibiting the mitochondrial fission factor, dynamin related protein 1.

Undifferentiated odontogenic epithelium and dental papilla cells differentiate into ameloblasts and odontoblasts, respectively, both of which are essential for tooth development. These differentiation processes involve dramatic functional and morphological changes of the cells. For these changes to occur, activation of mitochondrial functions, including ATP production, is extremely important. In addition, these changes are closely related to mitochondrial fission and fusion, known as mitochondrial dynamics. However, few studies have focused on the role of mitochondrial dynamics in tooth development. The purpose of this study was to clarify this role. We used mouse tooth germ organ cultures and a mouse dental papilla cell line with the ability to differentiate into odontoblasts, in combination with knockdown of the mitochondrial fission factor, dynamin related protein (DRP)1. In organ cultures of the mouse first molar, tooth germ developed to the early bell stage. The amount of dentin formed under DRP1 inhibition was significantly larger than that of the control. In experiments using a mouse dental papilla cell line, differentiation into odontoblasts was enhanced by inhibiting DRP1. This was associated with increased mitochondrial elongation and ATP production compared to the control. These results suggest that DRP1 inhibition accelerates dentin formation through mitochondrial elongation and activation. This raises the possibility that DRP1 might be a therapeutic target for developmental disorders of teeth.

Distribution of Cathepsin K in Late Stage of Tooth Germ Development and Its Function in Degrading Enamel Matrix Proteins in Mouse.

Cathepsin K (CTSK) is a member of cysteine proteinase family, and is predominantly expressed in osteoclastsfor degradationof bone matrix proteins. Given the similarity in physical properties of bone and dental mineralized tissues, including enamel, dentin and cementum, CTSK is likely to take part in mineralization process during odontogenesis. On the other hand, patients with pycnodysostosis caused by mutations of the CTSK gene displayedmultipledental abnormalities, such as hypoplasia of the enamel, obliterated pulp chambers, hypercementosis and periodontal disease. Thereforeitis necessary to study the metabolic role of CTSK in tooth matrix proteins. In this study, BALB/c mice at embryonic day 18 (E18), post-natal day 1 (P1), P5, P10 and P20 were used (5 mice at each time point)for systematic analyses of CTSK expression in the late stage of tooth germ development. We found that CTSK was abundantly expressed in the ameloblasts during secretory and maturation stages (P5 and P10) by immunohistochemistry stainings.During dentinogenesis, the staining was also intense in the mineralization stage (P5 and P10),but not detectable in the early stage of dentin formation (P1) and after tooth eruption (P20).Furthermore, through zymography and digestion test in vitro, CTSK was proved to be capable of hydrolyzing Emdogain and also cleaving Amelogenininto multiple products. Our resultsshed lights on revealing new functions of CTSK and pathogenesis of pycnodysostosis in oral tissues.

Involvement of IGF-2, IGF-1R, IGF-2R and PTEN in development of human tooth germ - an immunohistochemical study.

Insulin-Like Growth Factor 2 (IGF-2) is a peptide hormone essential for prenatal growth and development. IGF-2 exerts its mitogenic effects via Insulin-Like Growth Factor 1 Receptor (IGF-1R), and is eliminated by binding to Insulin-Like Growth Receptor 2 (IGF-2R). IGF-2 is also negatively regulated by Phosphatase and Tensin Homolog (PTEN), a phosphatase mutated in various tumors. Not much is known about the interplay between these factors during human odontogenesis. In this study, expression patterns of IGF-2, IGF-1R, IGF-2R and PTEN were analyzed by double immunofluorescence in incisor human tooth germs during the foetal period of development between the 7th and 20th gestational week. Throughout the investigated period, IGF-2 was mostly expressed in enamel organ, whereas mild to moderate expression of PTEN could be seen in dental papilla and parts of enamel organ. Expression of IGF-1R was ubiquitous and displayed strong intensity throughout the entire enamel organ. In contrast, expression of IGF-2R had rather erratic pattern in enamel organ and dental papilla alike. Expression patterns of IGF-2, IGF-1R, IGF-2R and PTEN in highly proliferative cervical loops, as well as in differentiating pre-ameloblasts and pre-odontoblasts of cusp tip region during the early and late bell stages when enamel organ acquires definitive shape, indicate importance of these factors in crown morphogenesis of human incisor. Taken together, our data suggest the involvement of IGF-2, IGF-1R, IGF-2R and PTEN in temporo-spatial patterning of basic cellular processes (proliferation, differentiation) during normal tooth development. They are also relevant for improving knowledge of molecular basis of human odontogenesis.

Facioscapulohumeral muscular dystrophy (FSHD) region gene 1 (FRG1) expression and possible function in mouse tooth germ development.

Abnormal expression of Facioscapulohumeral muscular dystrophy (FSHD) region gene 1 (FRG1) is involved in the pathogenesis of FSHD. FRG1 is also important for the normal muscular and vascular development. Our previous study showed that FRG1 is one of the highly expressed genes in the mandible on embryonic day 10.5 (E10.5) than on E12.0. In this study, we investigated the temporospatial expression pattern of FRG1 mRNA and protein during the development of the mouse lower first molar, and also evaluated the subcellular localization of the FRG1 protein in mouse dental epithelial (mDE6) cells. The FRG1 expression was identified in the dental epithelial and mesenchymal cells at the initiation and bud stages. It was detected in the inner enamel epithelium at the cap and early bell stages. At the late bell and root formation stages, these signals were detected in ameloblasts and odontoblasts during the formation of enamel and dentin matrices, respectively. The FRG1 protein was localized in the cytoplasm in the mouse tooth germ in vivo, while FRG1 was detected predominantly in the nucleus and faintly in the cytoplasm in mDE6 cells in vitro. In mDE6 cells treated with bone morphogenetic protein 4 (BMP4), the protein expression of FRG1 increased in cytoplasm, suggesting that FRG1 may translocate to the cytoplasm. These findings suggest that FRG1 is involved in the morphogenesis of the tooth germ, as well as in the formation of enamel and dentin matrices and that FRG1 may play a role in the odontogenesis in the mouse following BMP4 stimulation.

Slow cycling cells in the continuous dental lamina of Scyliorhinus canicula: new evidence for stem cells in sharks.

In the lesser spotted catshark (Scyliorhinus canicula), as in most non-mammalian vertebrates, the dentition renews throughout life. To contribute to our understanding of how continuous tooth replacement is achieved, we searched for evidence for the presence of stem cells in this species. Three-dimensional reconstructions of juvenile (2-3 weeks post-hatch) specimens showed that tooth families merge imperceptibly with so-called interdental zones within a continuous and permanent dental lamina. Interdental regions are composed of three layers, continuous with cervical loop, middle, and outer dental epithelium of the tooth families, respectively. A BrdU pulse-chase experiment revealed that cell proliferation is initiated in the lingual part of the dental lamina and the resulting population shifts one tooth position towards the oral epithelium in around four to five weeks. In the longest chase time (114 days) label-retaining and arguably non-differentiated cells were present at the lingual border of the dental lamina. These were found in the outer and middle dental epithelium, both within and between tooth families. This area of the dental lamina did not show expression or distribution of Sox2. Our data support the hypothesis that stem cells reside at the lingual border of the continuous dental lamina, more specifically in the middle dental epithelium at the level of the tooth families, and in its extension between the tooth families. To demonstrate their true stemness and their role in continuous tooth replacement, it remains to be shown that these cells have the potential to give rise to a complete new successor.

Altered distribution of Ghrelin protein in mice molar development.

OBJECTIVE: Ghrelin, an appetite-stimulating hormone, plays diverse regulatory functions in cell growth, proliferation, differentiation and apoptosis during mammalian development. There is limited information currently available regarding Ghrelin expression during mammalian tooth development, thus we aimed to establish the spatiotemporal expression of Ghrelin during murine molar odontogenesis. DESIGN: Immunohistochemistry was performed to detect the expression pattern of Ghrelin in mandible molar from E15.5 to PN7 during murine tooth development. RESULTS: The results showed that Ghrelin initially expressed in the inner enamel epithelium and the adjacent mesenchymal cells below, further with persistent expression in the ameloblasts and odontoblasts throughout the following developmental stages. In addition, Ghrelin was also present in Hertwig's epithelial root sheath at the beginning of tooth root formation. CONCLUSIONS: These results suggest that Ghrelin was present in tooth organs throughout the stages of tooth development, especially in ameloblasts and odontoblasts with little spatiotemporal expression differences. However, the potential regulatory roles of this hormone in tooth development still need to be validated by functional studies.

Spatial and temporal expression of histone demethylase, Kdm2a, during murine molar development.

The histone demethylase, lysine (K)-specific demethylase 2A (Kdm2a), is highly conserved and expressed ubiquitously. Kdm2a can regulate cell proliferation and osteo/dentinogenic, adipogenic and chondrogenic differentiation of mesenchymal stem cells (MSCs) derived from dental tissue. We used quantitative real-time RT-PCR analysis and immunohistochemistry to detect Kdm2a expression during development of the murine molar at embryonic days E12, E14, E16 and E17 and postnatal days P3 and P14. Immunohistochemistry results showed no positive staining of Kdm2a at E12. At E14, Kdm2a was expressed weakly in the inner enamel epithelium, stellate reticulum cells and dental sac. At E16, Kdm2a was expressed mainly in the inner and outer enamel epithelium, stratum intermedium and dental sac, but weaker staining was found in cervical loop and dental papilla cells adjacent to the basement membrane. At E17, the strongest Kdm2a staining was detected in the ameloblasts and stronger Kdm2a staining also was detected in the stratum intermedium, outer enamel epithelium and dental papilla cells compared to the expression at E16. Postnatally, we found that Kdm2a was localized in secretory and mature ameloblasts and odontoblasts, and dentin was unstained. Real-time RT-PCR showed that Kdm2a mRNA levels in murine germ cells increased from E12 to E14 and from E14 to E16; no significant change occurred at E16, E17 or P3, then the levels decreased at P14 compared to P3. Kdm2a expression may be closely related to cell proliferation, to ameloblast and odontoblast differentiation and to the secretion of extracellular enamel and dentin during murine tooth development.

Osteogenic Profile of Mesenchymal Cell Populations Contributing to Alveolar Bone Formation.

Teeth develop within the surrounding periodontal tissues, involving the alveolar bone, periodontal ligament and cementum. The alveolar bone originates through the process of intramembranous ossification involving mesenchymal cells from the tooth germ. As most available data are related to endochondral ossification, we examined the molecular background of alveolar bone development. We investigated the osteogenic profile of mesenchymal cells dissected from mouse mandible slices at the stage of early alveolar bone formation. Relative monitoring of gene expression was undertaken using PCR Arrays; this included the profiles of 84 genes associated with osteogenesis. To examine the tooth-bone interface, stages with detectable changes in bone remodelling during development (E13.0, E14.0 and E15.0) were chosen and compared with each other. These results showed a statistically significant increase in the expression of the genes Fgf3, Ctsk, Icam-1, Mmp9, Itga3 and Tuft1, and of a wide range of collagens (Col1a2, Col3a1, Col7a1, Col12a1, Col14a1). Decreased expression was detected in the case of Col2a1, Sox9, Smad2 and Vegfb. To confirm these changes in gene expression, immunofluorescence analyses of Mmp9 and Sox9 proteins were performed in situ. Our research has identified several candidate genes that may be crucial for the initiation of alveolar bone formation and is the basis for further functional studies.

Towards unraveling the human tooth transcriptome: the dentome.

The goal of the study was to characterize the transcriptome profiles of human ameloblasts and odontoblasts, evaluate molecular pathways and advance our knowledge of the human "dentome". Laser capture microdissection was used to isolate odontoblasts and ameloblasts from human tooth buds (15-20week gestational age) from 4 fetuses. RNA was examined using Agilent 41k whole genome arrays at 2 different stages of enamel formation, presecretory and secretory. Probe detection was considered against the array negative control to control for background noise. Differential expression was examined using Significance Analysis of Microarrays (SAM) 4.0 between different cell types and developmental stages with a false discovery rate of 20%. Pathway analysis was conducted using Ingenuity Pathway Analysis software. We found that during primary tooth formation, odontoblasts expressed 14,802 genes, presecretory ameloblasts 15,179 genes and secretory ameloblasts 14,526 genes. Genes known to be active during tooth development for each cell type (eg COL1A1, AMELX) were shown to be expressed by our approach. Exploring further into the list of differentially expressed genes between the motile odontoblasts and non-motile presecretory ameloblasts we found several genes of interest that could be involved in cell movement (FN1, LUM, ASTN1). Furthermore, our analysis indicated that the Phospholipase C and ERK5 pathways, that are important for cell movement, were activated in the motile odontoblasts. In addition our pathway analysis identified WNT3A and TGFB1 as important upstream contributors. Recent studies implicate these genes in the development of Schimke immuno-osseous dysplasia. The utility of laser capture microdissection can be a valuable tool in the examination of specific tissues or cell populations present in human tooth buds. Advancing our knowledge of the human dentome and related molecular pathways provides new insights into the complex mechanisms regulating odontogenesis and biomineralization. This knowledge could prove useful in future studies of odontogenic related pathologies.

Expression and roles of CCN2 in dental epithelial cells.

Connective tissue growth factor (CCN2) regulates diverse cellular functions, including tooth development. In order to delineate the precise role of CCN2 in the epithelium during odontogenesis, we investigated how it is expressed and what roles it may have in primary cultures of epithelial cells derived from developing tooth germ of the bovine fetus. Ccn2 mRNA and protein were strongly expressed in the inner dental epithelium, which is consistent with the expression of transforming growth factor-ß2 mRNA and proliferating cell nuclear antigen. Bone morphogenetic protein 4 (BMP4) and fibroblast growth factor 2 (FGF2) were also expressed in the inner dental epithelium, indicating that CCN2 functionally interacts with these factors in the epithelium. The stimulatory effects of FGF2 on cell proliferation and BMP4 on cell differentiation were additively up-regulated by CCN2 in a newly-established dental epithelium cell culture. Taken together, our data provide clear evidence that CCN2 is synthesized by inner dental epithelial cells, and appears to act as an autocrine factor, which regulates dental epithelial cell proliferation and differentiation in concert with growth factors.

Excess NF-κB induces ectopic odontogenesis in embryonic incisor epithelium.

Nuclear factor kappa B (NF-κB) signaling plays critical roles in many physiological and pathological processes, including regulating organogenesis. Down-regulation of NF-κB signaling during development results in hypohidrotic ectodermal dysplasia. The roles of NF-κB signaling in tooth development, however, are not fully understood. We examined mice overexpressing IKKß, an essential component of the NF-κB pathway, under keratin 5 promoter (K5-Ikkß). K5-Ikkß mice showed supernumerary incisors whose formation was accompanied by up-regulation of canonical Wnt signaling. Apoptosis that is normally observed in wild-type incisor epithelium was reduced in K5-Ikkß mice. The supernumerary incisors in K5-Ikkß mice were found to phenocopy extra incisors in mice with mutations of Wnt inhibitor, Wise. Excess NF-κB activity thus induces an ectopic odontogenesis program that is usually suppressed under physiological conditions.

Expression of Mmp-20 in dental germs of human fetus / Expresion de Mmp-20 en gérmenes dentales de fetos humanos

During experiments in animal studies, it has been observed that enamelysin (MMP-20) is expressed during tooth development in the late secretory stage of amelogenesis but not in the mature enamel.The aim of this research was to determine the location of MMP-20 in human tooth germs in the different structures of the enamel organ.The detection of MMP-20 was performed by immunohistochemistry in 20 specimens obtained from human fetuses. Immunostaining of MMP-20 was observed from the presecretor stadium in stellate reticulum and intermediate stratum and in the basal portion of ameloblasts in the secretory stage in stellate reticulum, stratum intermedium, secretory ameloblasts, odontoblasts and dental papilla. The results of this research show the location of MMP-20 in tooth germ development in humans and provides the foundation for future research about the process of dental organ formation.

En estudios realizados en animales de experimentación se ha observado que la enamelisina (MMP-20) se expresa durante el desarrollo dental durante el estadio de secreción tardío de la amelogénesis pero no en el esmalte maduro. El objetivo de la presente investigación fue determinar la localización de MMP-20 en gérmenes dentarios humanos en las diferentes estructuras del órgano del esmalte. Se analizaron 20 especímenes obtenidos de fetos humanos, efectuando la detección de MMP-20 por Inmunohistoquímica. Se observó inmunolocalización de MMP-20 desde el estadio presecretor en retículo estrellado y estrato intermedio, así como en porción basal de ameloblastos; en el estadio secretor en retículo estrellado, estrato intermedio, ameloblastos secretores, odontoblastos y papila dental. Los resultados de la presente investigación muestran la localización de la MMP-20 en el desarrollo del germen dentario en humanos y aporta las bases para futuras investigaciones acerca del proceso de formación de los órganos dentales.

YAP overexpression affects tooth morphogenesis and enamel knot patterning.

Teeth develop through distinct morphological stages. At the cap stage, a compactly clustered and concentrically arranged cell mass, the enamel knot, appears at the tip of the enamel organ. Cells in this knot express sets of key molecules, and as such have been proposed to act as a signaling center directing tooth morphogenesis and tooth cusp formation. YAP is a transcriptional co-activator of the Hippo signaling pathway that is essential for the proper regulation of organ growth. In this study, we analyzed the tooth phenotype in transgenic mice that overexpressed a constitutively active form of YAP in the dental epithelium. We found that overexpression of YAP resulted in deformed tooth morphogenesis with widened dental lamina. In addition, the enamel knot was mislocated to the upper portion of the enamel organ, where it remained devoid of proliferating cells and contained apoptotic cells with intense Edar transcripts and reduced E-cadherin expression. Interestingly, some signaling molecules, such as Shh, Fgf4, and Wnt10a, were not expressed in this mislocated enamel knot, but remained at the tip of the enamel organ. Analysis of these data suggests that the signaling center is induced by reciprocal epithelial-mesenchymal interactions, and its induction may be independent of the enamel knot.