Advances in the identification of deciduous molar tooth germs.

In forensic anthropology, correct identification of human deciduous teeth is of paramount importance for age-at-death estimation and relies on detailed anatomical descriptions. Yet literature is scarce on indications: details on the morphology of molar tooth germs of fetuses and newborns, developing from multiple mineralized centers that will eventually coalesce, are scant. This paper presents new anatomical elements for practitioners to identify human molar tooth germs at early developmental stages. 126 deciduous molars from 22 modern skeletons of fetuses and newborns (with a known age-at-death ranging between 0 days and 2 months and 21 days postnatal), without reported or observed dental pathological signs, were selected from the Collezione Antropologica LABANOF (CAL) documented skeletal collection. Gross anatomical descriptions of the morphology and configuration of the centers were provided, considering the number of mineralized centers, the shape and the outline of the occlusal plane at different stages. Three different developmental stages were observed in the maxillary first and second molar and the mandibular first molar, whereas in the mandibular second molar four stages were observed. For each stage, we provide additional detailed morphological descriptions, sketches outlining the shape of the tooth germ, and a picture of the tooth; also, indications for siding the teeth are presented. This information can be used by forensic anthropologists and odontologists for a proper identification when tooth germs are not found in anatomical connection within the dental sockets. Further analyses that encompass more age groups on a larger sample would allow to map the entire crown development of deciduous molars.

Insulin-like growth factor 1 modulates bioengineered tooth morphogenesis.

Regenerative therapy to replace missing teeth is a critical area of research. Functional bioengineered teeth have been produced by the organ germ method using mouse tooth germ cells. However, these bioengineered teeth are significantly smaller in size and exhibit an abnormal crown shape when compared with natural teeth. The proper sizes and shapes of teeth contribute to their normal function. Therefore, a method is needed to control the morphology of bioengineered teeth. Here, we investigated whether insulin-like growth factor 1 (IGF1) can regulate the sizes and shapes of bioengineered teeth, and assessed underlying mechanisms of such regulation. IGF1 treatment significantly increased the size of bioengineered tooth germs, while preserving normal tooth histology. IGF1-treated bioengineered teeth, which were developed from bioengineered tooth germs in subrenal capsules and jawbones, showed increased sizes and cusp numbers. IGF1 increased the number of fibroblast growth factor (Fgf4)-expressing enamel knots in bioengineered tooth germs and enhanced the proliferation and differentiation of dental epithelial and mesenchymal cells. This study is the first to reveal that IGF1 increases the sizes and cusp numbers of bioengineered teeth via the induction of enamel knot formation, as well as the proliferation and differentiation of dental epithelial and mesenchymal cells.

Bioengineered Tooth Buds Exhibit Features of Natural Tooth Buds.

Tooth loss is a significant health issue currently affecting millions of people worldwide. Artificial dental implants, the current gold standard tooth replacement therapy, do not exhibit many properties of natural teeth and can be associated with complications leading to implant failure. Here we propose bioengineered tooth buds as a superior alternative tooth replacement therapy. We describe improved methods to create highly cellularized bioengineered tooth bud constructs that formed hallmark features that resemble natural tooth buds such as the dental epithelial stem cell niche, enamel knot signaling centers, transient amplifying cells, and mineralized dental tissue formation. These constructs were composed of postnatal dental cells encapsulated within a hydrogel material that were implanted subcutaneously into immunocompromised rats. To our knowledge, this is the first report describing the use of postnatal dental cells to create bioengineered tooth buds that exhibit evidence of these features of natural tooth development. We propose future bioengineered tooth buds as a promising, clinically relevant tooth replacement therapy.

The characteristics of adjacent anatomy of mandibular third molar germs: a CBCT study to assess the risk of extraction.

This study aims to investigate the characteristics of adjacent anatomy of mandibular third molar germs (MTMGs). Three hundred Chinese patients aged 12 to 17 years old who received cone-beam computed tomography (CBCT) were enrolled. The continuity of cortical outline of inferior alveolar canals (IACs) in the region of MTMGs, the integrity of lingual bone cortex and the relationship between hard tissue part of MTMGs and IACs were investigated by observing CBCT data via the NNT viewer software. The age, degree of dental development, gender and location were recorded as variables. The associations between different variables and the observed data were analysed. The possibilities of disrupted cortical outline of IACs or the hard tissue part of MTMGs contacting IACs were significantly lower in 12 or 13 age groups, lower in Nolla stage ≤ 6. Males were significantly less than females in the incidence of disrupted cortical outline of IACs. As to the perforation of lingual bone cortex, no significant differences were observed in gender, ages, location or development stages. According to the CBCT images, anatomical factors contributed the lest to the risk of inferior alveolar nerve and lingual nerve injury in the 12 to 13 age group during removing the MTMG removal.

Determination of the mesio-distal tooth width via 3D imaging techniques with and without ionizing radiation: CBCT, MSCT, and µCT versus MRI.

Objective: The purpose of this study was to estimate the feasibility and accuracy of mesio-distal width measurements with magnetic resonance imaging (MRI) in comparison to conventional 3D imaging techniques [multi-slice CT (MSCT), cone-beam CT (CBCT), and µCT]. The measured values of the tooth widths were compared to each other to estimate the amount of radiation necessary to enable orthodontic diagnostics. Material and Methods: Two pig skulls were measured with MSCT, CBCT, µCT, and MRI. Three different judges were asked to determine the mesio-distal tooth width of 14 teeth in 2D tomographic images and in 3D segmented images via a virtual ruler in every imaging dataset. Results: Approximately 19% (27/140) of all test points in 2D tomographic slice images and 12% (17/140) of the test points in 3D segmented images showed a significant difference (P ≤ 0.05). The largest significant difference was 1.6mm (P < 0.001). There were fewer significant differences in the measurement of the tooth germs than in erupted teeth. Conclusions: Measurement of tooth width by MRI seems to be clinically equivalent to the conventional techniques (CBCT and MSCT). Tooth germs are better illustrated than erupted teeth on MRI. Three-dimensional segmented images offer only a slight advantage over 2D tomographic slice images. MRI, which avoids radiation, is particularly appealing in adolescents if these data can be corroborated in further studies.

Tooth replacement without a dental lamina: the search for epithelial stem cells in Polypterus senegalus.

Most actinopterygians replace their teeth continuously throughout life. To address the question of where and how replacement teeth form in actinopterygians, it is advisable to investigate well-chosen representatives within the lineage. The African bichir, Polypterus senegalus, belongs to the earliest diverged group of the actinopterygian lineage with currently living representatives. Its well characterized dentition, together with its phylogenetic position, make this species an attractive model to answer following questions: (1) when and where does the replacement tooth form and how is it connected with the dental organ of the predecessor, and (2) is there any evidence for the presence of epithelial stem cells, hypothesized to play a role in replacement? Serial sections show that one tooth family can contain up to three members, which are all interconnected by dental epithelium. Replacement teeth develop without the presence of a successional dental lamina. We propose that this is the plesiomorphic condition for tooth replacement in actinopterygians. BrdU pulse-chase experiments reveal cells in the outer and middle dental epithelium, proliferating at the time of initiation of a new replacement tooth. It is tempting to assume that these cell layers provide a stem cell niche. The observed absence of label-retaining cells after long chase times (up to 8 weeks) is held against the light of divergent views on cell cycling properties of stem cells. At present, our data do not support, neither reject, the hypothesis on involvement of epithelial stem cells within the process of continuous tooth replacement.

Ameloblasts express type I collagen during amelogenesis.

Enamel and enameloid, the highly mineralized tooth-covering tissues in living vertebrates, are different in their matrix composition. Enamel, a unique product of ameloblasts, principally contains enamel matrix proteins (EMPs), while enameloid possesses collagen fibrils and probably receives contributions from both odontoblasts and ameloblasts. Here we focused on type I collagen (COL1A1) and amelogenin (AMEL) gene expression during enameloid and enamel formation throughout ontogeny in the caudate amphibian, Pleurodeles waltl. In this model, pre-metamorphic teeth possess enameloid and enamel, while post-metamorphic teeth possess enamel only. In first-generation teeth, qPCR and in situ hybridization (ISH) on sections revealed that ameloblasts weakly expressed AMEL during late-stage enameloid formation, while expression strongly increased during enamel deposition. Using ISH, we identified COL1A1 transcripts in ameloblasts and odontoblasts during enameloid formation. COL1A1 expression in ameloblasts gradually decreased and was no longer detected after metamorphosis. The transition from enameloid-rich to enamel-rich teeth could be related to a switch in ameloblast activity from COL1A1 to AMEL synthesis. P. waltl therefore appears to be an appropriate animal model for the study of the processes involved during enameloid-to-enamel transition, especially because similar events probably occurred in various lineages during vertebrate evolution.

PrPc is temporospatially expressed in molar development of rats.

Odontogenesis, tooth development, is derived from two tissue components: ectoderm and neural crest-derived mesenchyme. Cyto-differentiation of odontogenic cells during development involves time-dependent and sequential regulation of genetic programs. This study was conducted to detect molecules implicated in cyto-differentiation of developing molar germs of rats. Differential display-PCR revealed that PrP(c) was differentially expressed between cap/early bell-staged germs (maxillary 3rd molar germs) and root formation-staged germs (maxillary 2nd molar germs) at postnatal day 9. Both levels of PrP(c) mRNA and protein expression were higher in the root formation stage than the cap/early bell stage and increased in a time-dependent manner. Immunofluorescence revealed for the first time that PrP(c) was not localized in the enamel organ, but localized in dental follicular cells for the development of the periodontal ligament and cementum as well as odontoblasts, both of which are of neural crest origin. These results suggest that the physiological functions of the PrP(c) in tooth development may be implicated in the differentiation of neural crest-derived mesenchyme including the periodontal tissues for root formation rather than epithelial tissue.

Pediatric maxillary fractures.

Pediatric craniofacial structures differ from those of adults in many ways. Because of these differences, management of pediatric craniofacial fractures is not the same as those in adults. The most important differences that have clinical relevance are the mechanical properties, craniofacial anatomy, healing capacity, and dental morphology. This article will review these key differences and the management of pediatric maxillary fractures. From the mechanical properties' perspective, pediatric bones are much more resilient than adult bones; as such, they undergo plastic deformation and ductile failure. From the gross anatomic perspective, the relative proportion of the cranial to facial structures is much larger for the pediatric patients and the sinuses are not yet developed. The differences related to dentition and dental development are more conical crowns, larger interdental spaces, and presence of permanent tooth buds in the pediatric population. The fracture pattern, as a result of all the above, does not follow the classic Le Fort types. The maxillomandibular fixation may require circum-mandibular wires, drop wires, or Ivy loops. Interfragmentary ligatures using absorbable sutures play a much greater role in these patients. The use of plates and screws should take into consideration the future development with respect to growth centers and the location of the permanent tooth buds. Pediatric maxillary fractures are not common, require different treatments, and enjoy better long-term outcomes.

Morphogenesis and bone integration of the mouse mandibular third molar.

The mouse third molar (M3) develops postnatally and is thus a unique model for studying the integration of a non-mineralized tooth with mineralized bone. This study assessed the morphogenesis of the mouse M3, related to the alveolar bone, comparing M3 development with that of the first molar (M1), the most common model in odontogenesis. The mandibular M3 was evaluated from initiation to eruption by morphology and by assessing patterns of proliferation, apoptosis, osteoclast distribution, and gene expression. Three-dimensional reconstruction and explant cultures were also used. Initiation of M3 occurred perinatally, as an extension of the second molar (M2) which grew into a region of soft mesenchymal tissue above the M2, still far away from the alveolar bone. The bone-free M3 bud gradually became encapsulated by bone at the cap stage at postnatal day 3. Osteoclasts were first visible at postnatal day 4 when the M3 came into close contact with the bone. The number of osteoclasts increased from postnatal day 8 to postnatal day 12 to form a space for the growing tooth. The M3 had erupted by postnatal day 26. The M3, although smaller than the M1, passed through the same developmental stages over a similar time span but showed differences in initiation and in the timing of bone encapsulation.

Differential expression of cxcl-14 during eruptive movement of rat molar germs.

Tooth eruption at the early postnatal period is strictly controlled by the molecules secreted mainly from follicular tissues, which recruit monocytes for osteoclast formation. In this study, it was hypothesized that different molecules can be expressed according to the stages of tooth eruption. Rat molar germs together with follicles were extracted and DD-PCR was performed from the root formation stage 2nd molars germs (after eruptive movement) and cap stage 3rd molar germs (before movement) at postnatal day 9. Cxcl-14, a potent chemoattractant, was detected as one of the differentially expressed molecules from DD-PCR. Its expression increased significantly at the root formation stage, compared with the cap or crown formation stage at both transcription and translation levels. The expression patterns of cxcl-14 were consistent with those of MCP-1 and CSF-1, and opposite to OPG. Immunofluorescence showed that cxcl-14 was localized in the dental follicular tissues only at the root formation stage overlaying the proximo-occlusal region of the molar germs. Many osteoclasts appeared on the surface of the alveolar bone which overlayed the occlusal region of the root formation stage 2nd molar germs and underwent resorption. Cxcl-14 expression was reduced considerably at both the translation and transcription levels by an alendronate treatment. These results suggest that cxcl-14 may be implicated in the formation of the eruptive pathway of tooth germs via osteoclastogenesis.

Differential expression of LAR tyrosine phosphatase in the rat developing molar tooth germ.

OBJECTIVE: This study examined the molecules implicated in the cytodifferentiation of dental hard tissue cells. METHODS: Rat pups at postnatal days 4, 7 and 10 were used. Differential display-polymerase chain reaction (DD-PCR), Western blot and immunofluorescent localisation were performed to search differentially expressed genes in tooth development. RESULTS: Leukocyte-common antigen-related tyrosine phosphatase (lar-tp) was differentially detected between the rat maxillary 2nd and 3rd molar germs on postnatal day 10, which were at the dental hard tissue formation and cap/early bell developmental stages, respectively. Both the mRNA and protein expression levels of lar-tp were higher in the 3rd molar germs than in the 2nd. In addition, the levels in the 2nd molar germs at postnatal days 4, 7 and 10, which corresponded to the early/late bell, crown and root stages, respectively, decreased in a time dependent manner. The immunoreactivity against intracellular P-subunit of lar-tp was detected in the ameloblasts and odontoblasts as well as in the undifferentiated inner enamel epithelia and dental papilla cells. However, strong immunoreactivity against extracellular E-subunit was observed only in the undifferentiated inner enamel epithelia and dental papilla cells in the 3rd molar germs and in the stratum intermedium in the 2nd molar germs. CONCLUSION: This is the first identification of lar-tp in the molar tooth development and suggests that this molecule may be involved in the cytodifferentiation of dental hard tissue cells.

Correlation between the pattern of facial growth and the position of the mandibular third molar.

PURPOSE: The purpose of this study was to examine the correlation between variables that determine the position of the third molar (M3) and correlation between the pattern of facial growth and examined variables. MATERIALS AND METHODS: Material for the study consisted of 130 panoramic radiographs and lateral radiographs of subjects' heads in whom orthodontic therapy had been started. Analysis of the position of the mandibular M3 and its relation to the bone and other teeth were determined by 4 variables: evaluation of the space for both M3, spatial relation between the second molar and M3, vertical position of the M3 in relation to the alveolar ridge, and inclination of the germ of the M3. RESULTS: In boys, significant correlation was determined on the right side of the jaw, between the retromolar area and the vertical position of M3. Significant correlation was determined in girls, between the retromolar area and the vertical position of M3 on both sides, retromolar area and M3 inclination on the left side, and retromolar area and spatial relation of M3 on the right side. Analysis of correlation between facial growth pattern and 4 variables that determinate the position of M3 showed positive correlation only for the lower right M3 inclination in subjects with anterior facial rotation. CONCLUSION: The present study showed that the amount of retromolar space for the position of the M3 is not a sign of its normal development. No significant differences were determined between the position of M3 and type of facial growth.

Establishment, maintenance and modifications of the lower jaw dentition of wild Atlantic salmon (Salmo salar L.) throughout its life cycle.

In this paper we elucidate the pattern of initiation of the first teeth and the pattern of tooth replacement on the dentary of wild Atlantic salmon (Salmo salar L.), throughout nearly all stages of its life cycle, using serially sectioned heads and jaws, cleared and stained animals, and X-rays. The dentary teeth are set in one row. Tooth germs appear around hatching, first in odd positions, followed by even positions. From position 8 further backwards, teeth are added in adjacent positions. The first replacement teeth appear in animals of about 30 mm fork length. On the dentary of early life stages (alevins and fry), every position in the tooth row holds a functional (i.e. attached and erupted) tooth and a replacement tooth. The alternating pattern set up anteriorly in the dentary by the first-generation teeth changes in juveniles (parr) whereby teeth are in a similar functional (for the erupted teeth) or developmental stage (for the replacement teeth) every three positions. This pattern is also observed in marine animals during their marine life phase and in both sexes of adult animals prior to spawning (grilse and salmon), but every position now holds either a functional tooth or a mineralised replacement tooth. This is likely due to the fact that replacement tooth germs have to grow to a larger size before mineralisation starts. In the following spring, the dentary tooth pattern of animals that have survived spawning (kelts) is highly variable. The abundance of functional teeth in post-spawning animals nevertheless indicates that teeth are not lost over winter. We confirm the earlier reported lack of evidence for the existence of an edentulous life phase, preceding the appearance of so-called breeding teeth during upstream migration to the spawning grounds, and consider breeding teeth to be just another tooth generation in a regularly replacing dentition. This study shows how Atlantic salmon maintains a functional adaptive dentition throughout its complex life cycle.

Primary enamel knot cell death in Apaf-1 and caspase-9 deficient mice.

During molar development, apoptosis occurs in a well-characterised pattern suggesting several roles for cell death in odontogenesis. However, molecular mechanisms of dental apoptosis are only poorly understood. In this study, Apaf-1 and caspase-9 knockouts were used to uncover the engagement of these members of the apoptotic machinery during early tooth development, concentrating primarily on their function in the apoptotic elimination of primary enamel knot cells. Molar tooth germ morphology, proliferation and apoptosis were investigated on frontal histological sections of murine heads at embryonic days (ED) 15.5, the stage when the primary enamel knot is eliminated apoptotically. In molar tooth germs of both knockouts, no apoptosis was observed according to morphological (haematoxylin-eosin) as well as biochemical criteria (TUNEL). Morphology of the mutant tooth germs, however, was not changed. Additionally, knockout mice showed no changes in proliferation compared to wild type mice. According to our findings on knockout embryos, Apaf-1 and caspase-9 are involved in apoptosis during tooth development; however, they seem dispensable and not necessary for proper tooth shaping. Compensatory or other mechanisms of cell death may act to eliminate the primary enamel knot cells in the absence of Apaf-1 and caspase-9.

[A histological innovative study of Balb/c mouse tooth germ transplanted to nude mouse in vivo].

PURPOSE: To observed the changes of the developed mouse's dental germs after transplanting into nude mouse and find some theoretical foundation for establishing an innovative experimental model. METHODS: Eight tooth germs from four 5th day postnatal Balb/c mice were transplanted to the back muscles of the adult nude mice. At seventh and fourteenth day after grafting, the germs were collected, fixed, demineralized, dehydrated, and embedded in wax. Serial sections of 5 microm thick were made following the routine methods, stained with haematoxylin-eosin dying solution, and observed under a light microscope. The mandibular first molars were taken out from the 12th and 19th day postnatal mouse. Serial sections of 5 microm thick were made following the routine methods, then compared with the germs after graft. RESULTS: All implantations were located in the superficial muscles with abundant capillary vessels. The dental germs could further developed after grafting under the microscope, but slower than dental germs self-development. The layer of dentin was thin, plenty of dentin with disorganized dentin tubule formed after grafting. Although the location of Hertwig's epithelial root sheath hardly moved, the roots developed further. The floor of pulp chamber could form and the pulp chambers were shrinking. The degree of calcification in the area of root increased very clearly. The inflammatory reaction was found at 7th day while hardly noted at 14th day after grafting. CONCLUSIONS: It suggests that late development of mouse tooth germs could further develop after heterotopic transplantation within the superficial muscles of the nude mouse. This model is useful for study of tooth root development in short time.

The HOX genes are expressed, in vivo, in human tooth germs: in vitro cAMP exposure of dental pulp cells results in parallel HOX network activation and neuronal differentiation.

Homeobox-containing genes play a crucial role in odontogenesis. After the detection of Dlx and Msx genes in overlapping domains along maxillary and mandibular processes, a homeobox odontogenic code has been proposed to explain the interaction between different homeobox genes during dental lamina patterning. No role has so far been assigned to the Hox gene network in the homeobox odontogenic code due to studies on specific Hox genes and evolutionary considerations. Despite its involvement in early patterning during embryonal development, the HOX gene network, the most repeat-poor regions of the human genome, controls the phenotype identity of adult eukaryotic cells. Here, according to our results, the HOX gene network appears to be active in human tooth germs between 18 and 24 weeks of development. The immunohistochemical localization of specific HOX proteins mostly concerns the epithelial tooth germ compartment. Furthermore, only a few genes of the network are active in embryonal retromolar tissues, as well as in ectomesenchymal dental pulp cells (DPC) grown in vitro from adult human molar. Exposure of DPCs to cAMP induces the expression of from three to nine total HOX genes of the network in parallel with phenotype modifications with traits of neuronal differentiation. Our observations suggest that: (i) by combining its component genes, the HOX gene network determines the phenotype identity of epithelial and ectomesenchymal cells interacting in the generation of human tooth germ; (ii) cAMP treatment activates the HOX network and induces, in parallel, a neuronal-like phenotype in human primary ectomesenchymal dental pulp cells.

Histological description of tooth formation in adult Eretmodus cf. cyanostictus (Teleostei, Cichlidae).

The Eretmodini, a tribe of closely related cichlids (Teleostei, Cichlidae) originating from Lake Tanganyika, possess oral tooth shapes ranging from conical (in Tanganicodus) over cylindrical (in Spathodus) to spatulate (in Eretmodus). Prior to a study aiming to understand how these distinctly different tooth shapes can be acquired in such closely related taxa, a detailed histological study was required of tooth formation in a representative of the eretmodines. Here, we present a histological description of replacement tooth development in Eretmodus cf. cyanostictus. Using light-microscopic observations on semithin as well as on ground sections, microradiographs and stereo-microscopic observations of both alizarine red S stained and unstained jaws we can conclude that tooth formation in adult E. cf. cyanostictus roughly corresponds with what is known for teleost tooth development in general. Remarkable features include the localization and shape of the epithelial downgrowth, the transient presence of a layer intermediate between inner dental epithelium (IDE) and outer dental epithelium (ODE), the asymmetric shape of the enamel organ, the fact that the pulp cavity recedes in front of the forming enameloid during enameloid formation, and finally, the pattern of matrix mineralisation and maturation, and the presence of pigment in the enameloid. The observation that the enamel organ in adult E. cf. cyanostictus develops asymmetrically is significant for understanding tooth shape variation in the Eretmodini.

Origin and developmental fate of vestigial tooth primordia in the upper diastema of the field vole (Microtus agrestis, Rodentia).

OBJECTIVE: Odontogenesis in voles is a convenient model to test hypotheses on tooth development generated from investigations in the mouse. Similar to other rodents, the functional dentition of the vole includes a toothless diastema. At its mesial end, a vestigial tooth bud has been found in the upper jaw of vole embryos. The aim of this study was to analyse the developmental dynamics of vestigial tooth structures in the upper diastema of the field vole and to compare it with the situation in the mouse. DESIGN: The development of odontogenic structures in the upper diastema of the field vole was investigated using serial histological sections and three-dimensional (3D) computer-aided reconstruction. RESULTS: A transient continuous dental lamina in the upper diastema of the field vole extended mesially to the first molar primordium, but was not continuous with the dental lamina in the incisor region. At its mesial limit, a large vestigial tooth primordium was regularly present. A further distinct vestigial bud was located mesially to the first molar primordium. The segmentation of the dental lamina suggested a potential to give rise to further vestiges in the upper diastema of the vole. CONCLUSIONS: In the prospective diastema of the vole exists as in the mouse a continuous dental lamina. Beside the prominent vestigial tooth bud in the mesial diastema, a further large bud was transiently located in front of the molars. The incorporation of dental epithelium into the first upper molar (M(1)) primordium in the vole differs from that in the mouse.