Msx1 is required for the induction of Patched by Sonic hedgehog in the mammalian tooth germ.
We have used the mouse developing tooth germ as a model system to explore the transmission of Sonic hedgehog (Shh) signal in the induction of Patched (Ptc). In the early developing molar tooth germ, Shh is expressed in the dental epithelium, and the transcripts of Shh downstream target genes Ptc and Gli1 are expressed in dental epithelium as well as adjacent mesenchymal tissue. The homeobox gene Msx1 is also expressed in the dental mesenchyme of the molar tooth germ at this time. We show here that the expression of Ptc, but not Gli1, was downregulated in the dental mesenchyme of Msx1 mutants. In wild-type E11.0 molar tooth mesenchyme SHH-soaked beads induced the expression of Ptc and Gli1. However, in Msx1 mutant dental mesenchyme SHH-soaked beads were able to induce Gli1 but failed to induce Ptc expression, indicating a requirement for Msx1 in the induction of Ptc by SHH. Moreover, we show that another signaling molecule, BMP4, was able to induce Ptc expression in wild-type dental mesenchyme, but induced a distinct expression pattern of Ptc in the Msx1 mutant molar mesenchyme. We conclude that in the context of the tooth germ Msx1 is a component of the Shh signaling pathway that leads to Ptc induction. Our results also suggest that the precise pattern of Ptc expression in the prospective tooth-forming region is controlled and coordinated by at least two inductive signaling pathways. (+info)
An immunocytochemical study of amelogenin proteins in the developing tooth enamel of the gar-pike, Lepisosteus oculatus (Holostei, Actinopterygii).
Previous studies have demonstrated the morphological similarity of the enamel-like layer found in the teeth of the coelacanth, lungfish and gar-pike to the enamel of tetrapods. In order to clarify the phylogenetic continuity between both structures, tooth germs of the gar-pike were immunocytochemically studied using an anti-bovine amelogenin polyclonal antibody. Intense immunoreaction was shown over the enamel-like matrix layer. Certain cell organelles associated with the secretory pathway of the ameloblasts were recognized as immunoreactive. These results indicate that the enamel-like layer of the gar-pike is a tissue homologous with the mammalian enamel because both possess a common, amelogenin-like substance. (+info)
Antagonistic signals between BMP4 and FGF8 define the expression of Pitx1 and Pitx2 in mouse tooth-forming anlage.
Members of the Pitx/RIEG family of homeodomain-containing transcription factors have been implicated in vertebrate organogenesis. In this study, we examined the expression and regulation of Pitx1 and Pitx2 during mouse tooth development. Pitx1 expression is detected in early development in a widespread pattern, in both epithelium and mesenchyme, covering the tooth-forming region in the mandible, and is then maintained in the dental epithelium from the bud stage to the late bell stage. Pitx2 expression, on the other hand, is restricted to the dental epithelium throughout odontogenesis. Interestingly, from E9.5 to E10.5, the expression domains of Pitx1 and Pitx2, in the developing mandible, overlap with that of Fgf8 but are exclusive to the zone of Bmp4 expression. Bead implantation experiments demonstrate that ectopic expression of Fgf8 can induce/maintain the expression of both Pitx1 and Pitx2 at E9.5. In contrast, Bmp4-expressing tissues and BMP4-soaked beads were able to repress Pitx1 expression in mandibular mesenchyme and Pitx2 expression in the presumptive dental epithelium, respectively. However, the effects of FGF8 and BMP4 are transient. It thus appears that the early expression patterns of Pitx1 and Pitx2 in the developing mandible are regulated by the antagonistic effects of FGF8 and BMP4 such that the Pitx1 and Pitx2 expression patterns are defined. These results indicate that the epithelial-derived signaling molecules are responsible not only for restricting specific gene expression in the dental mesenchyme, but also for defining gene expression in the dental epithelium. (+info)
Proteinases in developing dental enamel.
For almost three decades, proteinases have been known to reside within developing dental enamel. However, identification and characterization of these proteinases have been slow and difficult, because they are present in very small quantities and they are difficult to purify directly from the mineralizing enamel. Enamel matrix proteins such as amelogenin, ameloblastin, and enamelin are cleaved by proteinases soon after they are secreted, and their cleavage products accumulate in the deeper, more mature enamel layers, while the full-length proteins are observed only at the surface. These results suggest that proteinases are necessary for "activating" enamel proteins so the parent proteins and their cleavage products may perform different functions. A novel matrix metalloproteinase named enamelysin (MMP-20) was recently cloned from tooth tissues and was later shown to localize primarily within the most recently formed enamel. Furthermore, recombinant porcine enamelysin was demonstrated to cleave recombinant porcine amelogenin at virtually all of the sites that have previously been described in vivo. Therefore, enamelysin is at least one enzyme that may be important during early enamel development. As enamel development progresses to the later stages, a profound decrease in the enamel protein content is observed. Proteinases have traditionally been assumed to degrade the organic matrix prior to its removal from the enamel. Recently, a novel serine proteinase named enamel matrix serine proteinase-1 (EMSP1) was cloned from enamel organ epithelia. EMSP1 localizes primarily to the early maturation stage enamel and may, therefore, be involved in the degradation of proteins prior to their removal from the maturing enamel. Other, as yet unidentified, proteinases and proteinase inhibitors are almost certainly present within the forming enamel and await discovery. (+info)
A new function of BMP4: dual role for BMP4 in regulation of Sonic hedgehog expression in the mouse tooth germ.
The murine tooth development is governed by sequential and reciprocal epithelial-mesenchymal interactions. Multiple signaling molecules are expressed in the developing tooth germ and interact each other to mediate the inductive tissue interactions. Among them are Sonic hedgehog (SHH), Bone Morphogenetic Protein-2 (BMP2) and Bone Morphogenetic Protein-4 (BMP4). We have investigated the interactions between these signaling molecules during early tooth development. We found that the expression of Shh and Bmp2 is downregulated at E12.5 and E13.5 in the dental epithelium of the Msx1 mutant tooth germ where Bmp4 expression is significantly reduced in the dental mesenchyme. Inhibition of BMP4 activity by noggin resulted in repression of Shh and Bmp2 in wild-type dental epithelium. When implanted into the dental mesenchyme of Msx1 mutants, beads soaked with BMP4 protein were able to restore the expression of both Shh and Bmp2 in the Msx1 mutant epithelium. These results demonstrated that mesenchymal BMP4 represents one component of the signal acting on the epithelium to maintain Shh and Bmp2 expression. In contrast, BMP4-soaked beads repressed Shh and Bmp2 expression in the wild-type dental epithelium. TUNEL assay indicated that this suppression of gene expression by exogenous BMP4 was not the result of an increase in programmed cell death in the tooth germ. Ectopic expression of human Bmp4 to the dental mesenchyme driven by the mouse Msx1 promoter restored Shh expression in the Msx1 mutant dental epithelium but repressed Shh in the wild-type tooth germ in vivo. We further demonstrated that this regulation of Shh expression by BMP4 is conserved in the mouse developing limb bud. In addition, Shh expression was unaffected in the developing limb buds of the transgenic mice in which a constitutively active Bmpr-IB is ectopically expressed in the forelimb posterior mesenchyme and throughout the hindlimb mesenchyme, suggesting that the repression of Shh expression by BMP4 may not be mediated by BMP receptor-IB. These results provide evidence for a new function of BMP4. BMP4 can act upstream to Shh by regulating Shh expression in mouse developing tooth germ and limb bud. Taken together, our data provide insight into a new regulatory mechanism for Shh expression, and suggest that this BMP4-mediated pathway in Shh regulation may have a general implication in vertebrate organogenesis. (+info)
Interactions between the Wnt (wingless) and hedgehog signaling pathways were first described as playing a role in establishing boundaries between ectodermal cells in Drosophila segmentation. During the initiation of mammalian tooth development, boundaries that distinguish oral from dental ectoderm must be formed to correctly position the sites of tooth formation. We describe a reciprocal relationship between the expression of Wnt-7b in presumptive oral ectoderm and Shh in presumptive dental ectoderm in mouse embryos that mark boundaries between these cells with different developmental fates. By using a murine retrovirus to ectopically express Wnt-7b in presumptive dental ectoderm in mandibular arch explants, we show that Shh expression in the ectoderm and Ptc expression in the underlying ectomesenchyme are down-regulated, and tooth development is subsequently arrested. This suggests that Wnt-7b acts to repress Shh expression in oral ectoderm, thus maintaining the boundaries between oral and dental ectodermal cells. Implantation of beads soaked in Shh protein into Wnt-7b-infected explants resulted in complete rescue of tooth development, confirming that the repressive action of Wnt-7b specifically affects Shh signaling. (+info)
Nestin expression in embryonic and adult human teeth under normal and pathological conditions.
Nestin is an intermediate filament most related to neurofilaments and expressed predominantly in the developing nervous system and muscles. In the present study we examined the in vivo distribution of nestin in human teeth during embryonic development and in permanent teeth under normal and pathological conditions. The results show that nestin is first expressed at the bell stage and that its distribution is restricted in pulpal cells located at the cusp area of the fetal teeth. In young permanent teeth, nestin is found only in functional odontoblasts, which produce the hard tissue matrix of dentin. Expression is progressively down-regulated and nestin is absent from older permanent teeth. In carious and injured teeth, nestin expression is up-regulated in a selective manner in odontoblasts surrounding the injury site, showing a link between tissue repair competence and nestin up-regulation under pathological conditions. In an in vitro assay system of human dental pulp explants, nestin is up-regulated after local application of bone morphogenic protein-4. A similar effect is seen in cultures of primary pulp cells during their differentiation into odontoblasts. Taken together, these results suggest that nestin plays a potential role in odontoblast differentiation during normal and pathological conditions and that bone morphogenic protein-4 is involved in nestin up-regulation. (+info)
Molecular signaling and pulpal nerve development.
The purpose of this review is to discuss molecular factors influencing nerve growth to teeth. The establishment of a sensory pulpal innervation occurs concurrently with tooth development. Epithelial/mesenchymal interactions initiate the tooth primordium and change it into a complex organ. The initial events seem to be controlled by the epithelium, and subsequently, the mesenchyme acquires odontogenic properties. As yet, no single initiating epithelial or mesenchymal factor has been identified. Axons reach the jaws before tooth formation and form terminals near odontogenic sites. In some species, local axons have an initiating function in odontogenesis, but it is not known if this is also the case with mammals. In diphyodont mammals, the primary dentition is replaced by a permanent dentition, which involves a profound remodeling of terminal pulpal axons. The molecular signals underlying this remodeling remain unknown. Due to the senescent deterioration of the dentition, the target area of tooth nerves shrinks with age, and these nerves show marked pathological-like changes. Nerve growth factor and possibly also brain-derived neurotrophic factor seem to be important in the formation of a sensory pulpal innervation. Neurotrophin-3 and -4/5 are probably not involved. In addition, glial cell line-derived neurotrophic factor, but not neurturin, seems to be involved in the control of pulpal axon growth. A variety of other growth factors may also influence developing tooth nerves. Many major extracellular matrix molecules, which can influence growing axons, are present in developing teeth. It is likely that these molecules influence the growing pulpal axons. (+info)