The eternal tooth germ is formed at the apical end of continuously growing teeth

doi:10.1016/j.archoralbio.2004.09.008

Archives of Oral Biology

Volume 50, Issue 2, February 2005, Pages 153-157

https://doi.org/10.1016/j.archoralbio.2004.09.008 Get rights and content

Summary

Rodent incisors are known to be continuously growing teeth that are maintained by both the cell-proliferation at the apical end and the attrition of the incisal edge. This type of tooth had a special epithelial structure for the maintenance of stem cells, showing the bulbous epithelial protrusion at the apical end. The morphological transition of the epithelial–mesenchymal compartment by serial transverse sections of the apical end toward the incisal direction is likely to reflect the development of the tooth germ in the prenatal stage. Based on the present histological and previous molecular biological studies, the special structure at the apical end is obviously different from the cervical loop giving rise to Hertwig's epithelial root sheath (HERS), in human, mouse and rat molar tooth germs. Hence, we propose a new concept that the eternal tooth bud producing various dental progeny is formed at the apical end of continuously growing teeth, and a new term “apical bud” for indicating this specialized epithelial structure. Furthermore, BrdU labelling analysis suggested that the guinea-pig molars, which were continuously growing teeth, also possessed plural specific proliferative regions and “apical bud” at the apical end.

Introduction

Rodent incisors are continuously growing teeth, and all stages of odontogenesis including amelogenesis and dentinogenesis can be surveyed if we prepare the sections of the tooth from the apical end to the incisal edge.1, 2, 3 This phenomenon is maintained by both the cell-proliferation at the apical end and the attrition of the incisal edge. Recent molecular biological studies have clearly demonstrated the existence of the niche for the self-renewing adult stem cells in these rodent incisors and the molecular signals regulating the maintenance and cell fate decision of adult stem cells by the epithelial–mesenchymal interaction through fibroblast growth factor (FGF) signalling.4, 5 The term “cervical loop” has been so far used for indicating the epithelial tissue situated at the proliferative end of the rodent incisor.4, 5, 6 However, the “cervical loop” is the term referring to the junctional zone where the inner enamel epithelium meets the external enamel epithelium at the rim of the enamel organ.⁷ Thus, there is no suitable term for indicating this specialized epithelial compartment at the apical end of rodent incisors.

The stem cells divide slowly to give rise to one daughter cell that remains in the apical region and another cell enters the zone of rapidly dividing inner enamel epithelial cells (transit-amplifying cell population) to differentiate into ameloblasts to deposit the enamel matrix. In addition to the previous cell-proliferation assays in rodent incisors,1, 2, 4, 8, 9 our recent cell kinetic studies by double staining of 5-bromo-2′-deoxyuridine (BrdU) and Ki67 as the markers of dividing cells have clearly shown the presence of adult stem cells in the apical end of rodent incisors.¹⁰ Thus, the apical region of these teeth is totally different from “cervical loop” in rodent molars or human teeth from the viewpoint of the morphology and the biological significance.

The continuously growing teeth are represented not only by rodent incisors but also molars in certain other species, including rabbits, guinea-pigs, and field voles. In these animals, the structural similarity of the dental epithelium has been detected.11, 12, 13 Basically, these specific structures are composed of a large amount of the stellate reticulum and the basal epithelium. The present study aims to clarify the morphological features of the apical end of continuously growing teeth from both rodent incisors and guinea-pig molars and the structural similarities and differences between these two species.

Section snippets

Materials and methods

All experiments were performed following the Guidelines of the Niigata University Intramural Animal Use and Care Committee. The incisors were dissected carefully from the mandibles of 2- or 3-day-old mice after the amputation of heads under deep anaesthesia. Dissected teeth were fixed in 2% paraformaldehyde + 2.5% glutaraldehyde + 1% acrolein in 0.01 M phosphate buffer saline (PBS) (pH 7.2) overnight at 4 °C. They were then post-fixed in 1% OsO₄ with 1.5% potassium ferrocyanide for 2 h, dehydrated

Morphological features of the stem cell component in continuously growing incisors

Sagittal semithin sections of the rodent incisor tooth germ including the apical end showed that the labial epithelial compartment appeared as an oval-shaped structure. The morphology of serial transverse sections of the apical epithelial compartment showed the morphological features equivalent to bud, cap and bell stages-tooth germs of molar teeth. These structures were composed of the cells of inner and outer enamel epithelium, and stellate reticulum. Cutting more incisally, the mesial and

Discussion

Adult stem cells are present in many vertebrate regenerative tissues including the hematopoietic system, nervous system, gut, gonads, skin, olfactory epithelium and tooth.4, 14, 15, 16 Adult stem cells have been shown to undergo asymmetric cell division resulting in one daughter cell remaining in the stem cell compartment and another undergoing further cell divisions and giving rise to differentiated cells.4, 14 It is reasonable to suppose that a variety of continuously growing teeth possess

Acknowledgements

This work was supported by 21st century COE program (Osaka University), Grant-in-Aid to promote 2001-Multidisciplinary Research Projects in 2001–2005, KAKENHI (B) (16390523 to H.O. and 16390527 to H.H.), and KAKENHI (C) (15592159 to K.N.-O.) from MEXT, Japan

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Citation Excerpt :
The incisors and molars of rats exhibit two different development patterns. The incisors form enamel and erupt continuously throughout their lives, while the molars will no longer erupt after having formed completed roots (Ohshima et al., 2005). Tummers et al. (Tummers & Thesleff, 2003) suggested that the cervical-loop was a determinant of the developmental pattern of the teeth.
Rodent incisors and molars show different eruption patterns. Dental follicles and their interaction with dental epithelia play key roles in tooth eruption. However, little is known about the differences between incisor dental follicle (IF) and molar dental follicle (MF) during tooth eruption of rodents. This study aimed to investigate the differences between IF and MF during tooth eruption under induction with cervical-loop cells (CLC) and Hertwig’s epithelial root sheath (HERS) cells of rats.
CLC, HERS, IF, MF cells were isolated from 10 postnatal day 7 rats and identified by immunofluorescence staining. CLC or HERS cells-derived conditioned medium (CM) was obtained to induce IF and MF cells. Cell proliferation, mineralization, gene and protein expression related to tooth eruption were detected, and histological analysis was also performed.
The osteogenic differentiation and mineralization abilities of IF cells were stronger than those of MF cells. Both CLC and HERS cells-derived CM enhanced these abilities of IF cells, whereas they showed the opposite effect on MF cells. At 7, 10, and 15 d after birth, IF cells expressed more OPG and less RANKL than MF cells.
IF and MF cells present distinct characteristics in tooth eruption, CLC and HERS cells have significant inductive effects on them.
Stem cell properties of Gli1-positive cells in the periodontal ligament
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In this chapter, recent studies investigating the roles of Gli1-positive cells during tooth development are reviewed. Epithelial stem cells at the posterior apex of the rodent incisor, which is termed as the apical bud, facilitate a continuous tooth eruption throughout the life span of rodents [43–45]. Shh is expressed in the inner enamel epithelium and ameloblasts near the apical bud of the mouse incisor (Fig. 2A and B) [46].
The periodontal ligament (PDL), which surrounds the tooth root, contains mesenchymal stem cells (MSCs) capable of differentiating into osteoblasts, cementoblasts, and fibroblasts under normal conditions. These MSCs are thought to have important roles in the repair and regeneration of injured periodontal tissues. However, since there is no useful marker for MSCs in the PDL, the characteristics and distributions of these cells remain unclear. Gli1, an essential hedgehog signaling transcription factor, functions in undifferentiated cells during embryogenesis. Previous studies have demonstrated that the dental epithelial and mesenchymal cells positive for Gli1 in developing teeth have stem cell properties, including the ability to form colonies and pluripotency. Therefore, the focus of this review is the stem cell properties of Gli1-positive cells in the PDL, with an emphasis on the differentiation ability of osteoblasts for the regeneration of periodontal tissues.
Lineage tracing analysis identified Gli1-positive PDL cells as MSCs that contribute to the formation of periodontal tissues and can regenerate alveolar bone.
Gli1 is a potential stem cell marker in the PDL. A more definitive understanding of the functions of Gli1-positive cells could be useful for the development of regenerative methods using the MSCs in the PDL.
Potassium dihydrogen phosphate promotes the proliferation and differentiation of human periodontal ligament stem cells via nuclear factor kappa B pathway
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Periodontal ligament stem cells (PDLSCs) are vital for the regeneration of periodontal tissues. Potassium dihydrogen phosphate (KH₂PO₄) has recently been applied as a component of the mineralization inducing medium (MM), which can be used to induce osteogenic differentiation of dental stem cells. However, whether KH₂PO₄ has effects on PDLSCs has not been studied.
PDLSCs were isolated by magnetic activated cell sorting and cultured. Alkaline phosphatase (ALP) activity and ALP protein expression of PDLSCs treated with different concentrations of KH₂PO₄ were examined to make sure the optimal concentration of KH₂PO₄ for the following experiments. The effects of KH₂PO₄ on the proliferation and differentiation of PDLSCs were investigated by flow cytometry, cell counting kit-8 assay, alizarin red staining, real-time RT-PCR, and Western blot. The involvement of nuclear factor kappa B (NF-κB) pathway in KH₂PO₄-treated PDLSCs was analyzed by Western blot and alizarin red staining.
ALP activity assay and ALP protein expression examination revealed that 1.8 mmol/L KH₂PO₄ was the optimal concentration for the induction of hPDLSCs by KH₂PO₄. The proliferation and mineralization capacity of PDLSCs treated with KH₂PO₄ were enhanced as compared with the control group. PDLSCs treated with KH₂PO₄ showed an improved proliferation capacity in logarithmic growth phase at day 7. As PDLSCs were treated with KH₂PO₄, the expression of odonto/osteogenic markers (OCN/OCN, DSP/DSPP, OSX/OSX, RUNX2/RUNX2, and ALP/ALP) in cells were up-regulated at day 3 or 7. Moreover, the expression of IκBα in cytoplasm was down-regulated, along with an increased expression of p-P65 in cytoplasm and an up-regulated expression of P65 in nucleus. When treated with BMS345541 (the specific NF-κB inhibitor), the odonto/osteogenic differentiation of KH₂PO₄-treated PDLSCs was significantly attenuated.
KH₂PO₄ can improve the proliferation and odonto/osteogenic differentiation capacity of PDLSCs via NF-κB pathway, and thus represents a potential target involved in the regeneration of periodontium for clinical treatments.
Three-dimensional configuration of apical epithelial compartments including stem cell niches in guinea pig cheek teeth
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Continuously growing rodent incisors have an apically located epithelial stem cell compartment, known as an “apical bud” (AB). Few studies have described the morphological features of ABs and stem cell niches in continuously growing premolars/molars. We attempted to clarify the relationship between the three-dimensional configuration of ABs and the stem cell niches in guinea pig cheek teeth.
We perfusion-fixed four-week-old guinea pigs, then decalcified their premolars/molars to produce serial paraffin sections, which we immunostained for Sox2. We reconstructed the serial sections using image processing and analysis software. We processed undecalcified samples for scanning electron microscopy by KOH digestion.
Two types of epithelia with M and Δ shapes surrounded the S-shaped dental papilla in the apical region of the premolars/molars, and there were three Sox2-positive epithelial bulges above the M- and Δ-shaped epithelia. Sox2-positive epithelial stem cell niches were restricted to the apical side, and cell proliferation and differentiation immediately proceeded in the crown-analogue dentin. The Sox2-positive epithelial stem cell niches were sparsely distributed and extended to the occlusal side. We also detected continuously proliferating cells in the cervical loop and Hertwig’s epithelial root sheath of the root-analogue dentin.
Our findings suggest that guinea pig cheek teeth have three ABs, and the complex configuration of these types of teeth may be attributed to the prompt formation of crown-analogue dentin followed by the long-term formation of root-analogue dentin.
Experimental tooth movement temporally changes neural excitation and topographical map in rat somatosensory cortex
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During orthodontic treatment, binding teeth, may change the topographically organized representation of teeth in the cerebral cortex. To test the hypothesis that experimental tooth movement (ETM) changes the somatotopy of an individual tooth arrangement in the somatosensory cortex, we examined the spatiotemporal features of cortical excitatory propagation in response to mechanical stimulation of the maxillary incisor or molar using optical imaging in late adolescent rats without or with ETM. The ETM models consisted of 1d, 3d, and 7d ETM in which a closed-coil spring was ligated between the maxillary first molar and incisors. In controls, incisor and molar mechanical stimulation evoked excitation in the rostral and dorsocaudal regions of the primary somatosensory cortex (S1), respectively. In addition, the secondary somatosensory cortex and insular oral region (S2/IOR) were also activated. Incisor stimulation-induced excitatory regions in S1 of 3d and 7d ETM shifted without changing the maximum excitatory area or peak amplitude; the incisor stimulation-responding region moved toward the dorsocaudal region, which responded to molar stimulation in the control. This shift in excitatory region was not observed in 1d ETM. One day after removal of the coil spring that was attached for 6 days, the excitatory region shift in S1 was recovered to the control region. On the other hand, 1d ETM exhibited facilitation of the excitatory area and peak amplitude upon molar stimulation, and the facilitation of excitatory propagation disappeared in 3d and 7d ETM. These results may explain the clinical finding that abnormal sensation temporally occurs during orthodontic treatment.
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A 6-year-old female black-tailed prairie dog (Cynomys ludovicianus) was presented with a space-occupying lesion in the left submandibular region. On computed tomography, a low attenuating, poorly circumscribed mass infiltrated the left mandibular bone, with osteolytic change. Microscopically, the lesion was composed of odontogenic epithelium proliferating in nests and embedded in abundant dental papilla-like ectomesenchyme, including dentine and enamel. Multifocal amyloid deposition was observed. Immunohistochemically, the neoplastic epithelial cells were positive for cytokeratin (CK) AE1/AE3, CK14 and p63. Some epithelial cells were positive for amelogenin and some adjacent to the amyloid deposits co-expressed S100. The ectomesenchymal cells expressed vimentin and strong S100 immunoreactivity was observed in odontoblast-like cells. The amyloid was immunolabelled with amelogenin. The tumour was diagnosed as amyloid-producing odontoameloblastoma.

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The eternal tooth germ is formed at the apical end of continuously growing teeth

Summary

Introduction

Section snippets

Materials and methods

Morphological features of the stem cell component in continuously growing incisors

Discussion

Acknowledgements

Cell

Cell

Cellular renewal in the enamel organ and the odontoblast layer of the rat incisor as followed by radioautography using 5H-thymidine

Anat Rec

Movement of entire cell populations during renewal of the rat incisor as shown by radioautography after labelling with 3H-thymidine. The concept of a continuously differentiating cross-sectional segment (with an appendix on the development of the periodontal ligament)

Am J Anat

The relationship between odontoblasts and pulp capillaries in the process of enamel- and cementum-related dentin formation in rat incisors

Cell Tissue Res

Localization of putative stem cells in dental epithelium and their association with Notch and FGF signalling

J Cell Biol

FGF10 maintains stem cell compartment in developing mouse incisors

Development

Cell–matrix interactions and cell-cell junctions during epithelial histo-morphogenesis in the developing mouse incisor

Int J Dev Biol

Cellular renewal in the enamel organ and the odontoblast layer of the rat incisor as followed by radioautography using ⁵H-thymidine

Movement of entire cell populations during renewal of the rat incisor as shown by radioautography after labelling with ³H-thymidine. The concept of a continuously differentiating cross-sectional segment (with an appendix on the development of the periodontal ligament)