Hertwig's Epithelial Root Sheath Formation Research Articles

Malformations of the tooth root in humans.

The most common root malformations in humans arise from either developmental disorders of the root alone or disorders of radicular development as part of a general tooth dysplasia. The aim of this review is to relate the characteristics of these root malformations to potentially disrupted processes involved in radicular morphogenesis. Radicular morphogenesis proceeds under the control of Hertwig's epithelial root sheath (HERS) which determines the number, length, and shape of the root, induces the formation of radicular dentin, and participates in the development of root cementum. Formation of HERS at the transition from crown to root development appears to be very insensitive to adverse effects, with the result that rootless teeth are extremely rare. In contrast, shortened roots as a consequence of impaired or prematurely halted apical growth of HERS constitute the most prevalent radicular dysplasia which occurs due to trauma and unknown reasons as well as in association with dentin disorders. While odontoblast differentiation inevitably stops when growth of HERS is arrested, it seems to be unaffected even in cases of severe dentin dysplasias such as regional odontodysplasia and dentin dysplasia type I. As a result radicular dentin formation is at least initiated and progresses for a limited time. The only condition affecting cementogenesis is hypophosphatasia which disrupts the formation of acellular cementum through an inhibition of mineralization. A process particularly susceptible to adverse effects appears to be the formation of the furcation in multirooted teeth. Impairment or disruption of this process entails taurodontism, single-rooted posterior teeth, and misshapen furcations. Thus, even though many characteristics of human root malformations can be related to disorders of specific processes involved in radicular morphogenesis, precise inferences as to the pathogenesis of these dysplasias are hampered by the still limited knowledge on root formation.

Odontoblast β-catenin signaling regulates fenestration of mouse Hertwig's epithelial root sheath.

The interaction between Hertwig's epithelial root sheath (HERS) and the adjacent mesenchyme is vitally important in mouse tooth root development. We previously generated odontoblast-specific Ctnnb1 (encoding β-catenin) deletion mice, and demonstrated that odontoblast β-catenin signaling regulates odontoblast proliferation and differentiation. However, the role of odontoblast β-catenin signaling in regulation of HERS behavior has not been fully investigated. Here, using the same odontoblast- specific Ctnnb1 deletion mice, we found that ablation of β-catenin signaling in odontoblasts led to aberrant HERS formation. Mechanistically, odontoblast-specific Ctnnb1 deletion resulted in elevated bone morphogenetic protein 7 (Bmp7) expression and reduced expression of noggin and follistatin, both of which encode extracellular inhibitors of BMPs. Furthermore, the levels of phosphorylated Smad1/5/8 were increased in HERS cells. In vitro tissue culture confirmed that BMP7 treatment disrupted the HERS structure. Taken together, we demonstrated that odontoblast β-catenin signaling may act through regulation of BMP signaling to maintain the integrity of HERS cells.

Cell dynamics in cervical loop epithelium during transition from crown to root: implications for Hertwig's epithelial root sheath formation

Some clinical cases of hypoplastic tooth root are congenital. Because the formation of Hertwig's epithelial root sheath (HERS) is an important event for root development and growth, we have considered that understanding the HERS developmental mechanism contributes to elucidate the causal factors of the disease. To find integrant factors and phenomenon for HERS development and growth, we studied the proliferation and mobility of the cervical loop (CL). We observed the cell movement of CL by the DiI labeling and organ culture system. To examine cell proliferation, we carried out immunostaining of CL and HERS using anti-Ki67 antibody. Cell motility in CL was observed by tooth germ slice organ culture using green fluorescent protein mouse. We also examined the expression of paxillin associated with cell movement. Imaging using DiI labeling showed that, at the apex of CL, the epithelium elongated in tandem with the growth of outer enamel epithelium (OEE). Cell proliferation assay using Ki67 immunostaining showed that OEE divided more actively than inner enamel epithelium (IEE) at the onset of HERS formation. Live imaging suggested that mobility of the OEE and cells in the apex of CL were more active than in IEE. The expression of paxillin was observed strongly in OEE and the apex of CL. The more active growth and movement of OEE cells contributed to HERS formation after reduction of the growth of IEE. The expression pattern of paxillin was involved in the active movement of OEE and HERS. The results will contribute to understand the HERS formation mechanism and elucidate the cause of anomaly root.

Reduction of Egf signaling decides transition from crown to root in the development of mouse molars

Mouse, rat, and human molars begin to form their roots after the completion of crown morphogenesis. Though several signaling pathways and transcription factors have been implicated in the regulation of molar crown development, relatively little is known about the regulatory mechanisms involved in the transition from crown to root development. Tooth root formation is initiated by the development of Hertwig's epithelial root sheath (HERS) from the cervical loop in the enamel organ. In this study we examined the change in epidermal growth factor (Egf) signaling during this transition process. Immunohistochemical studies showed that the expression of Egf receptors in the enamel organ disappear gradually in the process and are not observed in HERS. Here, to examine the effect of Egf on the transition, we used the organ culture method to examine the root development. In the presence of Egf, stellate reticulum (SR) cells between the inner and outer epithelial layers in the enamel organ actively proliferated and maintained the enamel organ, and the formation of HERS was not observed. On the other hand, in either the absence of Egf or the presence of the inhibitor of Egf receptors, the SR cells disappeared and HERS formation started. Subsequently, root formation proceeded in the culture period. Therefore, disappearance of SR area may be a key event that controls the timing of onset of HERS formation, and Egf may be one of regulatory factors involved in the change from cervical loop epithelium to HERS during root development.

BMP4 signaling regulates formation of Hertwig’s epithelial root sheath during tooth root development

Although Hertwig's epithelial root sheath (HERS) performs an important function in the formation of the tooth root, the developmental mechanisms that control HERS growth and differentiation remain to be thoroughly elucidated. Bone morphogenetic protein 4 (BMP4), which is secreted by mesenchymal cells, acts on the dental epithelium as a regulator of cell differentiation during crown formation. In an effort to determine whether BMP4 specifically regulates the development of HERS in the dental epithelium, we assessed the localizations of BMP4, BMP receptor-IB (BMPR-IB), and BMPR-II during molar root formation in the mouse. HERS cells were shown to express BMPR-IB and BMPR-II. BMP4-positive cells were detected densely in the dental papillae around HERS, thereby suggesting that BMP4 participated in HERS formation. Beads soaked in BMP4, NOGGIN, or phosphate-buffered saline (PBS) were implanted into the pulp cavity under culture conditions, and the length of HERS was evaluated with regard to the proliferating cells. After 12 h, both groups exhibited a similar HERS developmental pattern, with the length and shape of HERS bearing a close resemblance to one another. However, after 48 h, the observed HERS elongation was significantly shorter in the BMP4-treated group. In addition, proliferative cell nuclear antigens were detectable only in the NOGGIN- and PBS-treated groups. These findings demonstrate that mesenchymally expressed BMP4 regulates HERS development by preventing elongation and maintaining cell proliferation. BMP4 may, therefore, prove useful as a root-formation regulatory agent in a variety of tissue-engineering applications.

Molecular Mechanisms Regulating Transition from Crown to Root Formation in the Development of Mouse Molars

The tooth root is a foundation structure embedded in the alveolar bone of the jaws. Root formation starts after the completion of crown morphogenesis and proceeds slowly while coordinating the formation of the periodontal tissue that supports the tooth. Although several signaling pathways and transcription factors have been implicated in the regulation of molar crown development, relatively little is known about the regulatory mechanisms governing the transition from crown to root development. The formation of Hertwig's epithelial root sheath (HERS), consisting of 2 epithelial layers, is a key event in the initiation of root development. Recently, a new hypothesis about the mechanism of HERS formation was proposed based on the results of experiments using Fibroblast growth factor (Fgf) -10-deficient mice. In this review, we discuss this new hypothesis of HERS development and the signaling change for HERS formation during the transition process from crown to root development.

The Role of BMP4 in HERS during Tooth Root Development

Tooth development is regulated by reciprocal interactions between epithelial and mesenchymal cells. Several morphological studies suggest that growth factors secreted from these cells regulate tooth development. These growth factors induce undifferentiated cells in the tooth germ to differentiate into tooth-forming cells. Many growth factors are reportedly expressed in the developing tooth, but the key molecule underlying tooth root development is still unknown. There is a considerable amount of data on the expression and function of bone morphogenetic proteins (BMPs) during tooth development, but the relationship between BMPs and tooth root development has not been analyzed extensively. This prompted us to examine the expression patterns of BMP4 and BMP receptors (BMPRs) at the apical region of the tooth germ during mouse tooth root development. We found that BMP4 was strictly expressed in the apical mesenchyme, and that BMPRs were localized in Hertwig's epithelial root sheath (HERS). We also investigated the possible roles of BMP4 in HERS during the early stage of tooth root development. Our organ-culture results indicated that BMP4 regulates HERS formation via harmonious interactions with its own antagonist, noggin, by preventing elongation and maintaining cell proliferation. BMP4 may therefore be useful in various tissue-engineering applications as a regulator of tooth root formation.

Nfic Gene Disruption Inhibits Differentiation of Odontoblasts Responsible for Root Formation and Results in Formation of Short and Abnormal Roots in Mice

Nuclear factor I genes play an important role in the development of the brain, lung, and roots of teeth. We had reported that Nfic-deficient mice form normal crowns, but abnormal roots of molar teeth. However, the mechanism by which the disruption of Nfic gene causes abnormal root formation remains unknown. To understand this mechanism, the root formation in Nfic-deficient mice was examined and compared to that of wild-type mice by morphological, immunohistochemical, and in situ hybridization analyses. Nfic-deficient mice formed normal Hertwig's epithelial root sheath (HERS) but severely disrupted odontoblast differentiation, leading to the formation of aberrant odontoblasts in the early stage of root formation. They became dissociated and polygonal in shape, lost their orientation and polarity, and did not express dentin sialophosphoprotein. The abnormal roots contained trapped aberrant odontoblasts, thereby resembling osteodentin in overall morphology. No osteoclasts were associated with abnormal roots. Further, the abnormal roots exhibited a decreased number of cementoblasts and cementum formation on the root surface. The loss of Nfic did not interfere with the formation of HERS, but it caused disrupted odontoblast differentiation, which resulted in the formation of short and abnormal roots, and decreased cementum. This finding suggests that root dentin is required for normal cementum formation. Therefore, Nfic may be a key regulator of root odontoblast differentiation and root formation.