The Development of a Measuring System for Intraoral SpO2.

Blood oxygen saturation (SpO2) is an essential indicator of a patient's general condition. However, conventional measurement methods have some issues such as time delay and interference by ambient light. Improved measurement methods must be developed, and there are no reports on intraoral measurements of SpO2 using wearable devices. Therefore, we aimed to establish an intraoral SpO2 measurement method for the first time. Twelve healthy adults participated in this study. The following steps were taken: (1) to identify the optimal measurement location, mid-perfusion index (PI) values were measured at six places on the mucosa of the maxilla, (2) to validate the optimal measurement pressure, PI values were obtained at different pressures, and (3) using the proposed mouthpiece device, SpO2 values in the oral cavity and on the finger were analyzed during breath-holding. The highest PI values were observed in the palatal gingiva of the maxillary canine teeth, with high PI values at pressures ranging from 0.3 to 0.8 N. In addition, changes in SpO2 were detected approximately 7 s faster in the oral cavity than those on the finger, which is attributed to their proximity to the heart. This study demonstrates the advantage of the oral cavity for acquiring biological information using a novel device.

Comparison of Maxillofacial Morphology Between Modified Furlow's and Modified two-Flap Palatoplasty in Orofacial Clefts During the Primary Dentition Period.

OBJECTIVE: To assess the effect of two palatoplasty procedures, modified Furlow's palatoplasty (F procedure) and modified two-flap palatoplasty (T procedure), on the maxillofacial morphology of unilateral cleft lip and palate (UCLP) and bilateral cleft lip and palate (BCLP) during the primary dentition period. DESIGN: Retrospective cohort study. SETTING: Department of Orthodontics, School of Dentistry, Showa University. PARTICIPANTS: We enrolled 106 pediatric patients (63 boys, 43 girls; aged 4.43 ± 0.34 years) with non-syndromic orofacial clefts who underwent cheiloplasty and palatoplasty. INTERVENTIONS: Patients were divided into four groups according to cleft type (BCLP or UCLP) and palatoplasty procedure type (F or T procedure). MAIN OUTCOME MEASURES: Maxillofacial morphology was assessed by examining parameters on lateral cephalograms. RESULTS: Multiple comparisons revealed significant differences among N-A, N-ANS, and ANS-PNS distances and SNA and ANB angles among the groups. There were significant differences in N-A, N-ANS, ANS-PNS, SNA, and SNB among the UCLP and BCLP groups. The calculated effect sizes were all within 0.3-0.5. The statistical power was as follows: N-A, 86.41%; N-ANS, 79.77%; ANS-PNS, 97.49%; SNA, 96.88%; and ANB, 99.25%. CONCLUSIONS: Although UCLP and BCLP both exhibited differences in craniofacial distances and angles, the procedure type (either F or T procedure) had no significant effect on the maxillofacial morphology (as determined by lateral cephalograms). However, to rule out additional bias, patient-specific factors should be considered that may be affected by maxillofacial development when deciding surgical approaches.

Autocrine regulation of mesenchymal progenitor cell fates orchestrates tooth eruption.

Formation of functional skeletal tissues requires highly organized steps of mesenchymal progenitor cell differentiation. The dental follicle (DF) surrounding the developing tooth harbors mesenchymal progenitor cells for various differentiated cells constituting the tooth root-bone interface and coordinates tooth eruption in a manner dependent on signaling by parathyroid hormone-related peptide (PTHrP) and the PTH/PTHrP receptor (PPR). However, the identity of mesenchymal progenitor cells in the DF and how they are regulated by PTHrP-PPR signaling remain unknown. Here, we show that the PTHrP-PPR autocrine signal maintains physiological cell fates of DF mesenchymal progenitor cells to establish the functional periodontal attachment apparatus and orchestrates tooth eruption. A single-cell RNA-seq analysis revealed cellular heterogeneity of PTHrP+ cells, wherein PTHrP+ DF subpopulations abundantly express PPR. Cell lineage analysis using tamoxifen-inducible PTHrP-creER mice revealed that PTHrP+ DF cells differentiate into cementoblasts on the acellular cementum, periodontal ligament cells, and alveolar cryptal bone osteoblasts during tooth root formation. PPR deficiency induced a cell fate shift of PTHrP+ DF mesenchymal progenitor cells to nonphysiological cementoblast-like cells precociously forming the cellular cementum on the root surface associated with up-regulation of Mef2c and matrix proteins, resulting in loss of the proper periodontal attachment apparatus and primary failure of tooth eruption, closely resembling human genetic conditions caused by PPR mutations. These findings reveal a unique mechanism whereby proper cell fates of mesenchymal progenitor cells are tightly maintained by an autocrine system mediated by PTHrP-PPR signaling to achieve functional formation of skeletal tissues.

A New Intraoral Six-Degrees-of-Freedom Jaw Movement Tracking Method Using Magnetic Fingerprints.

We proposed a novel jaw movement tracking method that can measure in six degrees of freedom. The magnetic field generated by a permanent magnet paired with a small, low-power-consumption Hall effect magnetic sensor is used to estimate the relative distance between two objects-in this instance, the lower and upper jaws. By installing a microelectromechanical system (MEMS) orientation sensor in the device, we developed a mouthpiece-type sensing device that can measure voluntary mandibular movements in three-dimensional orientation and position. An evaluation of individuals wearing this device demonstrated its ability to measure mandibular movement with an accuracy of approximately 3 mm. Using the movement recording feature with six degrees of freedom also enabled the evaluation of an individual's jaw movements over time in three dimensions. In this method, all sensors are built onto the mouthpiece and the sensing is completed in the oral cavity. It does not require the fixation of a large-scale device to the head or of a jig to the teeth, unlike existing mandibular movement tracking devices. These novel features are expected to increase the accessibility of routine measurements of natural jaw movement, unrestricted by an individual's physiological movement and posture.

Three-Dimensional Assessment of Craniofacial Features in Patients With Down Syndrome During the Mixed Dentition Period: A Case-Control Study.

OBJECTIVES: Down syndrome (DS) is a common congenital chromosomal disorder related to trisomy 21. Lateral cephalometric radiography studies have shown that patients with DS have characteristic craniofacial morphology; however, no 3-dimensional analysis studies have been performed to investigate the craniofacial features, including volumetric aspects, of patients with DS. The present study was performed to evaluate the craniofacial features, including volumetric aspects, of patients with DS and to compare these findings with control participants using cone beam computed tomography (CBCT). MATERIALS AND METHODS: The study sample consisted of 12 patients with DS and 12 control participants. All participants were examined by means of CBCT; the resulting images were used for evaluation of maxillary and mandibular volume, cranial base, and craniofacial measurements. Differences between patients with DS and control participants were statistically analyzed using Student t test. RESULTS: Compared to control participants, patients with DS exhibited statistically significant reductions in maxillary and mandibular volumes. Both sagittal and axial cranial base linear measurements were shorter in patients with DS than in control participants. In contrast, the cranial base angle was enhanced in patients with DS, compared with control participants. Moreover, condylion (Co)-gnathion, anterior nasal spine-menton, and Co-subspinale (point A) measurements were shorter in patients with DS than in control participants; the sella-nasion-mandibular plane angle was significantly reduced in patients with DS, compared with control participants. CONCLUSION: Our results suggest that patients with DS have distinct skeletal volume and craniofacial morphology features, relative to individuals without DS.

Longitudinal Follow-Up of Facial Growth of Patients With Unilateral Cleft Lip and Palate Following Modified Veau-Wardill-Kilner Palatoplasty.

OBJECTIVE: To evaluate facial growth after modified Veau-Wardill-Kilner/pushback (PB) palatoplasty from childhood to adulthood in patients with unilateral cleft lip and palate (UCLP). DESIGN: Retrospective longitudinal study. SETTING: Single center. PATIENTS: Sixty-two (29 males and 33 females) consecutive patients with nonsyndromic UCLP. INTERVENTIONS: Pushback palatoplasty and subsequent cleft palate treatment. MAIN OUTCOME MEASURES: Lateral and posterior-anterior cephalograms were taken at 3 different phases: phase 1 (before first-stage orthodontic treatment; N = 58; average age, 4.9 ± 1.1 years), phase 2 (before second-stage orthodontic treatment; N = 58; 15.9 ± 1.1 years), and phase 3 (after orthodontic retention; N = 51; 22.1 ± 3.2 years). RESULTS: The majority of patients had skeletal class III morphology in all 3 phases due to retrognathic maxilla. Maxillary growth did not improve in phase 2 despite first-stage orthodontic treatment in phase 1. Maxillary morphology improved in phase 3 but retardation occurred, although 77.42% of patients received orthognathic surgery during second-stage orthodontic treatment. Mandibular growth was slightly reduced in phases 1 and 2 and the mandible remained retrognathic in phase 3, following mandibular setback orthognathic surgery. The horizontal occlusal cant was slightly upward and toward the cleft side with respect to the reference plane, and the upper midline was deviated to the cleft side in phases 1 and 3. CONCLUSIONS: Patients with UCLP who undergo phased PB palatoplasty, orthodontic treatment, and orthognathic surgeries do not maintain skeletal class I facial morphology.

Neural crest-derived cells in nasal conchae of adult mice contribute to bone regeneration.

Neural crest-derived cells (NCDCs), a class of adult stem cells not restricted to embryonic tissues, are attractive tissue regenerative therapy candidates because of their ease of isolation, self-renewing properties, and multipotency. Although adult NCDCs can undergo osteogenic differentiation in vitro, whether they induce bone formation in vivo remains unclear. Previously, our group reported findings showing high amounts of NCDCs scattered throughout nasal concha tissues of adult mice. In the present study, NCDCs in nasal conchae labeled with enhanced green fluorescent protein (EGFP) were collected from adult P0-Cre/CAG-CAT-EGFP double transgenic mice, then cultured in serum-free medium to increase the number. Subsequently, NCDCs were harvested and suspended in type I atelocollagen gel, then an atelocollagen sponge was used as a scaffold for the cell suspension. Atelocollagen scaffolds with NCDCs were placed on bone defects created in a mouse calvarial bone defect model. Over the ensuing 12 weeks, micro-CT and histological analysis findings showed that mice with scaffolds containing NCDCs had slightly greater bone formation as compared to those with a scaffold alone. Furthermore, Raman spectroscopy revealed spectral properties of bone in mice that received scaffolds with NCDCs similar to those of native calvarial bone. Bone regeneration is important not only for gaining bone mass but also chemical properties. These results are the first to show the validity of biomolecule-free adult nasal concha-derived NCDCs for bone regeneration, including the chemical properties of regenerated bone tissue.

A three-dimensional analysis of primary failure of eruption in humans and mice.

OBJECTIVES: Primary failure of eruption (PFE) is a genetic disorder exhibiting the cessation of tooth eruption. Loss-of-function mutations in parathyroid hormone (PTH)/parathyroid hormone-related peptide (PTHrP) receptor (PTH/PTHrP receptor, PPR) were reported as the underlying cause of this disorder in humans. We showed in a PFE mouse model that PTHrP-PPR signaling is responsible for normal dental follicle cell differentiation and tooth eruption. However, the mechanism underlying the eruption defect in PFE remains undefined. In this descriptive study, we aim to chronologically observe tooth eruption and root formation of mouse PFE molars through 3D microCT analyses. SETTING AND SAMPLE POPULATION: Two individuals with PFE were recruited at Showa University. A mouse PFE model was generated by deleting PPR specifically in PTHrP-expressing dental follicle and divided into three groups, PPRfl/fl ;R26RtdTomato/+ (Control), PTHrP-creER;PPRfl/+ ;R26RtdTomato/+ (cHet), and PTHrP-creER;PRRfl/fl ;R26RtdTomato/+ (cKO). MATERIALS AND METHODS: Images from human PFE subjects were acquired by CBCT. All groups of mouse samples were studied at postnatal days 14, 25, 91, and 182 after a tamoxifen pulse at P3, and superimposition of 3D microCT images among three groups was rendered. RESULTS: Mouse and human PFE molars exhibited a similar presentation in the 3D CT analyses. The quantitative analysis in mice demonstrated a statistically significant decrease in the eruption height of cKO first and second molars compared to other groups after postnatal day 25. Additionally, cKO molars demonstrated significantly shortened roots with dilacerations associated with the reduced interradicular bone height. CONCLUSIONS: Mouse PFE molars erupt at a much slower rate compared to normal molars, associated with shortened and dilacerated roots and defective interradicular bones.

Cdc42 regulates cranial suture morphogenesis and ossification.

Cdc42 (cell division cycle 42) is ubiquitously expressed small GTPases belonging to the Rho family of proteins. Previously, we generated limb bud mesenchyme-specific Cdc42 inactivated mice (Cdc42 conditional knockout mice; Cdc42 fl/fl; Prx1-Cre), which showed short limbs and cranial bone deformities, though the mechanism related to the cranium phenotype was unclear. In the present study, we investigated the role of Cdc42 in cranial bone development. Our results showed that loss of Cdc42 caused a defect of intramembranous ossification in cranial bone tissues which is related to decreased expressions of cranial suture morphogenesis genes, including Indian hedgehog (Ihh) and bone morphogenetic proteins (BMPs). These findings demonstrate that Cdc42 plays a crucial role in cranial osteogenesis, and is controlled by Ihh- and BMP-mediated signaling during cranium development.

Orthognathic surgery induces genomewide changes longitudinally in DNA methylation in saliva.

OBJECTIVE: Orthognathic surgery dramatically changes morphology of the maxillofacial deformity and improves the malocclusion morphologically and functionally. We investigated the influence of orthognathic surgery on genomewide DNA methylation in saliva. METHODS: Saliva was obtained from nine patients undergoing orthognathic surgery and two healthy reference individuals before and 3 months after orthognathic surgery. Genomewide DNA methylation profiling of saliva (341,482 CpG dinucleotides) was conducted using Infinium HumanMethylation450 BeadChips. RESULTS: Comparison between pre- and postsurgery saliva samples revealed significant changes in DNA methylation patterns at 2,381 CpG sites (p < 0.01) with suggestive significance. The differentially methylated probe sets were significantly associated with the cancer pathway (p = 2.8 × 10-7 ; a false discovery rate q-value = 3.7 × 10-4 ) and PI3K-Akt signalling pathway (p = 2.4 × 10-5 ; a false discovery rate q-value = 3.1 × 10-2 ). CONCLUSION: Pathway enrichment analysis of genes with suggestive significance demonstrated that altered DNA methylation in saliva of patients undergoing orthognathic surgery, possibly as a response to surgical stress or bone injury. Further studies with a large sample size and long-term observation are needed to validate the phenomena identified in this study.

Analysis of the relationship between the morphology of the palate and facial skeletal patterns in Class III malocclusion using structural equation modelling.

INTRODUCTION: The present study investigated the relationship between facial skeletal patterns and morphology of the palate in adult patients with Class III malocclusion using structural equation modelling (SEM). SETTING AND SAMPLE POPULATION: One hundred cone beam computed tomography images of Class III adults were evaluated for skeletal measurements. MATERIALS AND METHODS: The skeletal measurements were classified into the vertical, anteroposterior and transverse group based on factor analysis. 3D scanning model of the maxilla was analysed by Generalized procrustes analysis (GPA) and principal component analysis (PCA). Structural equation modelling was used to analyse relationship among the skeletal and morphometric factors. RESULTS: According to the factor analysis, latent variables were extracted by each skeletal variable. First principal component (PC1) and PC2 of palatal morphology were used to analyse relationship with skeletal variables. As results of the structural equation model, the transverse latent variable had the most influence on PC1, followed by vertical and anteroposterior variables. This result means that as the facial width increases, the palate becomes narrower, deeper and longer. CONCLUSIONS: The relationship between the skeletal pattern with Class III malocclusion and palatal morphology was analysed through SEM. The transverse facial skeletal pattern showed the highest correlation with PC1 of palatal morphology.

Association between the FGFR1 rs13317 single nucleotide polymorphism and orbitale-nasion depth based on cephalometric images.

The fibroblast growth factor receptor 1 (FGFR1) gene plays an important role in craniofacial morphogenesis. In our previous study, an association between FGFR1 single nucleotide polymorphisms (SNPs) and craniofacial morphology was demonstrated in Japanese and Korean subjects. The present study aimed to evaluate the relationship between a common FGFR1 SNP (rs13317) with craniofacial morphology, increasing the number of measurements and examining Egyptian subjects (n = 191) in addition to the Japanese (n = 211) and Korean (n = 226) subjects. Genotyping for rs13317 was performed using the TaqMan assay, and its associations with 81 craniofacial measurements derived from lateral and posteroanterior cephalograms were analyzed by multiple regression analysis controlling sex and facial size. The results from each of the populations were then statistically combined. In the Egyptian subjects, rs13317 was significantly associated with the nasion-orbitale depth (P = 0.00040), and a suggestive association was also observed in the Japanese (P = 0.037) and Korean subjects (P = 0.045). The combined analysis revealed that only the nasion-orbitale depth showed a significant association (P = 0.000062) and that several measurements showed a suggestive association. Our results strongly indicate that rs13317 is associated with a smaller depth between the nasion and orbitale, representing a relative protrusion of the cheekbones and retrusion of the nasal root. A similar characteristic is also observed in individuals with Pfeiffer syndrome, which is caused by a dysfunctional FGFR1 mutation.

Whole-genome sequencing in a pair of monozygotic twins with discordant cleft lip and palate subtypes.

OBJECTIVE: Orofacial clefts (OFCs) are common and etiologically complex birth defects. This study explored potential genetic differences in a pair of Japanese monozygotic (MZ) twins with different forms of OFC using whole-genome sequencing. SUBJECTS AND METHODS: One co-twin (MZ-1) presented with nonsyndromic bilateral cleft lip and palate; the other co-twin (MZ-2) had nonsyndromic bilateral cleft lip and unilateral left-sided cleft alveolus. Neither parent had an OFC. Craniofacial morphologic features and potential genetic differences were compared using standard cephalometry and whole-genome sequencing, respectively. RESULTS: Morphologically, MZ-1 had a smaller vertical mandibular height, compared to MZ-2. However, no discordant genetic differences were detected. Moreover, both twins and their parents harbored rare candidate gene variants (GRHL3; TPM1) considered to be associated with OFCs. CONCLUSION: The observed differences between MZ-1 and MZ-2 in craniofacial morphology assessed by cephalograms might be directly attributable to the effects of the OFC on growth and/or differences in surgical history, given the lack of any differences in genetic background. However, comparisons of discordant MZ twins should continue to identify novel candidates that might control OFC or that might partly explain the missing heritability for this common birth defect, in addition to understanding craniofacial growth and development.

Common polymorphisms in WNT10A affect tooth morphology as well as hair shape.

Hair and teeth are appendages of ectodermal origin, and there are common molecular backgrounds involved in their formation. To date, it has been revealed that a non-synonymous polymorphism in EDAR has effects on the morphological variation in both hair and teeth. Previous association studies have confirmed that single-nucleotide polymorphisms (SNPs) in/near THADA, FRAS1, WNT10A, NAF1 and FGFR2 are associated with hair morphology. In this study, we thus examined whether these SNPs are also associated with dental characteristics. We measured metric dental traits including crown size and also evaluated non-metric dental traits using plaster casts obtained from subjects (272 Japanese and 226 Koreans). DNA samples were prepared from the subjects and genotyped for the hair morphology-associated SNPs. We observed a significant association of crown size with an SNP in WNT10A (rs7349332), but not with SNPs in other genes. Therefore, we further examined four SNPs within and around WNT10A, among which rs10177996 had the strongest association with dental traits. World distribution of the derived allele in rs10177996, which is associated with larger teeth, showed that Eurasians have a higher allele frequency than Africans. Together with previous studies on hair morphology, this study demonstrated that common variations in WNT10A have pleiotropic effects on the morphology of ectodermal appendages.

Assessment of the root apex position of impacted maxillary canines on panoramic films.

INTRODUCTION: In this study, we aimed to analyze the root apex positions of impacted maxillary canines on panoramic radiographs according to their labiopalatal positions and to propose a new panoramic radiographic predictor of impacted maxillary canines. METHODS: Paired panoramic films and cone-beam computed tomography images of 104 patients with unilateral impacted maxillary canines were selected. The sector locations of the impacted canine root apices on the panoramic radiographs were compared with the labiopalatal positions of impacted maxillary canines on cone-beam computed tomography. Statistical correlations between panoramic and cone-beam computed tomography findings were examined with the chi-square test. RESULTS: A statistically significant association was observed between the sector locations of impacted canine root apices and the labiopalatal positions of the canines (P <0.001). The root apices of labially impacted canines were more frequently located in sector 2 (corresponding to the lateral incisor position), those of palatally impacted canines were more frequently located in sector 4 (corresponding to the first premolar), and those of midalveolus impacted canines were more frequently located in sector 3 (corresponding to the canine). CONCLUSIONS: On panoramic films, the root apices of palatally impacted canines tended toward the root apices of the maxillary first premolars, whereas those of labially impacted canines tended toward the root apices of the maxillary lateral incisors. Therefore, the sector locations of impacted canine root apices on panoramic radiographs could be used for the early prediction of maxillary canine impactions and determination of the labiopalatal positions of impacted canines.

Rho GTPase protein Cdc42 is critical for postnatal cartilage development.

Cdc42, a small Rho GTPase family member, has been shown to regulate multiple cellular functions in vitro, including actin cytoskeletal reorganization, cell migration, proliferation, and gene expression. However, its tissue-specific roles in vivo remain largely unknown, especially in postnatal cartilage development, as cartilage-specific Cdc42 inactivated mice die within a few days after birth. In this study, we investigated the physiological functions of Cdc42 during cartilage development after birth using tamoxifen-induced cartilage-specific inactivated Cdc42 conditional knockout (Cdc42 (fl/fl); Col2-CreERT) mice, which were generated by crossing Cdc42 flox mice (Cdc42 (fl/fl)) with tamoxifen-induced type II collagen (Col2) Cre transgenic mice using a Cre/loxP system. The gross morphology of the Cdc42 cKO mice was shorter limbs and body, as well as reduced body weight as compared with the controls. In addition, severe defects were found in growth plate chondrocytes of the long bones, characterized by a shorter proliferating zone (PZ), wider hypertrophic zone (HZ), and loss of columnar organization of proliferating chondrocytes, resulting in delayed endochondral bone formation associated with abnormal bone growth. Our findings demonstrate the importance of Cdc42 for cartilage development during both embryonic and postnatal stages.

Localization and osteoblastic differentiation potential of neural crest-derived cells in oral tissues of adult mice.

In embryos, neural crest cells emerge from the dorsal region of the fusing neural tube and migrate throughout tissues to differentiate into various types of cells including osteoblasts. In adults, subsets of neural crest-derived cells (NCDCs) reside as stem cells and are considered to be useful cell sources for regenerative medicine strategies. Numerous studies have suggested that stem cells with a neural crest origin persist into adulthood, especially those within the mammalian craniofacial compartment. However, their distribution as well as capacity to differentiate into osteoblasts in adults is not fully understood. To analyze the precise distribution and characteristics of NCDCs in adult oral tissues, we utilized an established line of double transgenic (P0-Cre/CAG-CAT-EGFP) mice in which NCDCs express green fluorescent protein (GFP) throughout their life. GFP-positive cells were scattered like islands throughout tissues of the palate, gingiva, tongue, and buccal mucosa in adult mice, with those isolated from the latter shown to form spheres, typical cell clusters composed of stem cells, under low-adherent conditions. Furthermore, GFP-positive cells had markedly increased alkaline phosphatase (a marker enzyme of osteoblast differentiation) activity and mineralization as shown by alizarin red staining, in the presence of bone morphogenetic protein (BMP)-2. These results suggest that NCDCs reside in various adult oral tissues and possess potential to differentiate into osteoblastic cells. NCDCs in adults may be a useful cell source for bone regeneration strategies.

Quantitative skeletal maturation estimation using cone-beam computed tomography-generated cervical vertebral images: a pilot study in 5- to 18-year-old Japanese children.

OBJECTIVES: The purpose of this study was to establish multivariable regression models for the estimation of skeletal maturation status in Japanese boys and girls using the cone-beam computed tomography (CBCT)-based cervical vertebral maturation (CVM) assessment method and hand-wrist radiography. MATERIALS AND METHODS: The analyzed sample consisted of hand-wrist radiographs and CBCT images from 47 boys and 57 girls. To quantitatively evaluate the correlation between the skeletal maturation status and measurement ratios, a CBCT-based CVM assessment method was applied to the second, third, and fourth cervical vertebrae. Pearson's correlation coefficient analysis and multivariable regression analysis were used to determine the ratios for each of the cervical vertebrae (p < 0.05). RESULTS: Four characteristic parameters ((OH2 + PH2)/W2, (OH2 + AH2)/W2, D2, AH3/W3), as independent variables, were used to build the multivariable regression models: for the Japanese boys, the skeletal maturation status according to the CBCT-based quantitative cervical vertebral maturation (QCVM) assessment was 5.90 + 99.11 × AH3/W3 - 14.88 × (OH2 + AH2)/W2 + 13.24 × D2; for the Japanese girls, it was 41.39 + 59.52 × AH3/W3 - 15.88 × (OH2 + PH2)/W2 + 10.93 × D2. CONCLUSIONS: The CBCT-generated CVM images proved very useful to the definition of the cervical vertebral body and the odontoid process. The newly developed CBCT-based QCVM assessment method showed a high correlation between the derived ratios from the second cervical vertebral body and odontoid process. CLINICAL RELEVANCE: There are high correlations between the skeletal maturation status and the ratios of the second cervical vertebra based on the remnant of dentocentral synchondrosis.

Identification of gene expression profile of neural crest-derived cells isolated from submandibular glands of adult mice.

Neural crest cells in the embryo migrate to reach target sites as neural crest-derived cells (NCDCs) where they differentiate into a variety of derivatives. Some NCDCs are maintained in an undifferentiated state throughout the life of the animal and are considered to be a useful cell source for regenerative medicine. However, no established method to obtain NCDCs sufficient for regenerative medicine from adults with high purity has been presented, since their distribution in adult tissues is not fully understood. It is critical to identify reliable markers for NCDCs in adults, as the expressions of P0 and Wnt1, the most reliable NCDC markers, are shut off in the embryonic stage. To analyze the characteristics of NCDCs in adult tissues, we utilized a double transgenic mouse strain, P0-Cre/CAG-CAT-EGFP transgenic mice (P0 mice), in which NCDCs were shown to express EGFP and we were able to recognize GFP-positive cells in those. We focused on the submandibular glands (SMGs), which are known to be derived from the neural crest. GFP-positive cells were shown to be scattered like islands in the SMGs of adult P0 mice. We surgically removed SMGs from adult mice and digested samples into single cell suspensions. GFP-positive cells separated using flow cytometry expressed a high level of Sox10, a marker of embryonic neural crest cells, suggesting successful isolation of NCDCs. To identify candidate marker genes in isolated NCDCs, we performed DNA microarray analyses and real-time PCR analysis of GFP-positive and -negative cells isolated from P0 mice, then selected genes showing differential gene expression patterns. As compared to GFP-negative cells, GFP-positive cells expressed Gpr4 and Ednrb at higher levels, whereas Pdgfra and Pdgfrb were expressed at lower levels. Furthermore, DNA microarray analysis showed that GFP-positive cells were positive for aquaporin 5, a marker for acinar cells. Together, our results indicate that NCDCs in adult SMGs have characteristic gene expression profiles specially their cell surface molecules. Cell sorting using a combination of these specific cell surface proteins would be a useful strategy for isolation of NCDCs from SMGs with high purity.

Mastication markedly affects mandibular condylar cartilage growth, gene expression, and morphology.

INTRODUCTION: Mandibular growth is believed to be strongly related to mastication. Furthermore, mandibular condylar cartilage is known to be derived from neural crest cells. We examined whether the degree of chewing affects condylar cartilage growth of the mandible. METHODS: Mice were fed diets with varying hardness. Genes specific to neural crest-derived cells were measured by real-time polymerase chain reaction to compare the expression changes between the mandibular and tibia cartilages. The mandibular condylar cartilage was then evaluated histologically, and proliferation was evaluated using proliferating cell nuclear antigen. Immunostaining was conducted for osteopontin, type X collagen, and Musashi1, and real-time polymerase chain reaction was used to assess the expression levels of osteopontin and type X collagen. RESULTS: Markers including P75, Wnt-1, Musashi1, and Nestin were upregulated in the mandibular condylar cartilage as compared with the tibial cartilage. Histologic assessment of the mandibular cartilage showed that the hypertrophic chondrocyte zone was statistically significantly thicker in mice fed a hard diet. Chondrocyte proliferation and Musashi1 expression were lower in mice fed a hard diet. After 4 weeks, numerous osteopontin and type X collagen-positive cells were observed in mice fed a mixed diet. CONCLUSIONS: Mastication affects the balance between differentiation and proliferation in the mandibular condylar cartilage. This phenomenon might be attributed to the presence of neural crest-derived cells.