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Analysis of various genetically modified mice, with supernumerary teeth, has revealed the following two intrinsic molecular mechanisms that increase the number of teeth. One plausible explanation for supernumerary tooth formation is the rescue of tooth rudiments. Topical application of candidate molecules could lead to whole tooth formation under suitable conditions. Congenital tooth agenesis is caused by the cessation of tooth development due to the deletion of the causative gene and suppression of its function. The arrest of tooth development in Runx2 knockout mice, a mouse model of congenital tooth agenesis, is rescued in double knockout mice of Runx2 and Usag-1. The Usag-1 knockout mouse is a supernumerary model mouse. Targeted molecular therapy could be used to generate teeth in patients with congenital tooth agenesis by stimulating arrested tooth germs. The third dentition begins to develop when the second successional lamina is formed from the developing permanent tooth in humans and usually regresses apoptotically. Targeted molecular therapy, therefore, seems to be a suitable approach in whole-tooth regeneration by the stimulation of the third dentition. A second mechanism of supernumerary teeth formation involves the contribution of odontogenic epithelial stem cells in adults. Cebpb has been shown to be involved in maintaining the stemness of odontogenic epithelial stem cells and suppressing epithelial-mesenchymal transition. Odontogenic epithelial stem cells are differentiated from one of the tissue stem cells, enamel epithelial stem cells, and odontogenic mesenchymal cells are formed from odontogenic epithelial cells by epithelial-mesenchymal transition. Both odontogenic epithelial cells and odontogenic mesenchymal cells required to form teeth from enamel epithelial stem cells were directly induced to form excess teeth in adults. An approach for the development of targeted therapeutics has been the local application of monoclonal neutralizing antibody/siRNA with cationic gelatin for USAG-1 or small molecule for Cebpb.

Uterine sensitization–associated gene-1 (USAG-1) deficiency leads to enhanced bone morphogenetic protein (BMP) signaling, leading to supernumerary teeth formation. Furthermore, antibodies interfering with binding of USAG-1 to BMP, but not lipoprotein receptor–related protein 5/6 (LRP5/6), accelerate tooth development. Since USAG-1 inhibits Wnt and BMP signals, the essential factors for tooth development, via direct binding to BMP and Wnt coreceptor LRP5/6, we hypothesized that USAG-1 plays key regulatory roles in suppressing tooth development. However, the involvement of USAG-1 in various types of congenital tooth agenesis remains unknown. Here, we show that blocking USAG-1 function through USAG-1 knockout or anti–USAG-1 antibody administration relieves congenital tooth agenesis caused by various genetic abnormalities in mice. Our results demonstrate that USAG-1 controls the number of teeth by inhibiting development of potential tooth germs in wild-type or mutant mice missing teeth. Anti–USAG-1 antibody administration is, therefore, a promising approach for tooth regeneration therapy., 先天性無歯症に対する分子標的薬の開発 --USAG-1を標的分子とした歯再生治療--. 京都大学プレスリリース. 2021-02-15.

Objective Fibrous dysplasia (FD) is a benign bone disease characterized by fibro-osseous lesions. FD is caused by somatic mutations in the gene, guanine nucleotide-binding protein, alpha stimulating activity polypeptide 1 (GNAS), which encodes the G protein subunit, Gsα. FD manifests early in life, but the growth of lesions usually ceases in adulthood. FD lesions often exhibit somatic mutation mosaicism. In this study, the relationship between lesion growth and mutation prevalence within a lesion was investigated. Design Lesions from five FD patients were characterized by radiographical, histological and immunohistochemical methods. To accurately calculate the prevalence of mutations within lesions, GNAS codon 201 in genomic DNA isolated from fresh surgical FD specimens was sequenced. Results Uniquely, a lesion in one 46-year-old patient was still growing, enabling simultaneous analysis of both stable-old and active-new FD lesions in the same patient. Immunohistochemical analysis indicated that a newer, proximal lesion was growing while an older, distal lesion was not. The mutation prevalence differed between these lesions; it was low in the old and high in the new lesion. Thus, the frequency of mutated cells had decreased in the older lesion. Conclusions This is the first direct evidence for the age-dependent demise of mutated cells in FD, helping to explain why FD lesion growth generally ceases in adulthood.

Adult Cebpb KO mice incisors present amelogenin-positive epithelium pearls, enamel and dentin allopathic hyperplasia, fewer Sox2-positive cells in labial cervical loop epitheliums, and reduced Sox2 expression in enamel epithelial stem cells. Thus, Cebpb acts upstream of Sox2 to regulate stemness. In this study, Cebpb KO mice demonstrated cementum-like hard tissue in dental pulp, loss of polarity by ameloblasts, enamel matrix in ameloblastic layer, and increased expression of epithelial-mesenchymal transition (EMT) markers in a Cebpb knockdown mouse enamel epithelial stem cell line. Runx2 knockdown in the cell line presented a similar expression pattern. Therefore, the EMT enabled disengaged odontogenic epithelial stem cells to develop supernumerary teeth. Cebpb and Runx2 knockdown in the cell line revealed higher Biglycan and Decorin expression, and Decorin-positive staining in the periapical region, indicating their involvement in supernumerary tooth formation. Cebpb and Runx2 acted synergistically and played an important role in the formation of supernumerary teeth in adult incisors.

While the prevalence of supernumerary teeth (ST) is high in permanent dentition, the etiology of ST in humans remains unclear. However, multiple murine models of ST have elaborated on dated mechanisms traditionally ascribed to ST etiology: one involves the rescue of rudimental teeth, and the second considers the contribution of odontogenic epithelial stem cells. It remains unclear whether these mechanisms of ST formation in mice are applicable to humans. The third dentition is usually regressed apoptotic—that is, the teeth do not completely form in humans. Recently, it was suggested that ST result from the rescue of regression of the third dentition in humans. The present investigation evaluates the proportion of collected general ST cases that evinced a third dentition based on the clinical definition of ST derived from the third dentition. We also investigated the contribution of SOX2-positive odontogenic epithelial stem cells to ST formation in humans. We collected 215 general ST cases from 15,008 patients. We confirmed that the general characteristics of the collected ST cases were similar to the results from previous reports. Of the 215 cases, we narrowed our analysis to the 78 patients who had received a computed tomography scan. The frequency of ST considered to have been derived from the third dentition was 26 out of 78 cases. Evidence of a third dentition was especially apparent in the premolar region, was more common in men, and was more likely among patients with ≥3 ST. SOX2-positive odontogenic epithelial stem cells within the surrounding epithelial cells of developing ST were observed in non–third dentition cases and not in third dentition cases. In conclusion, the third dentition is the main cause of ST in humans. The odontogenic epithelial stem cells may contribute to ST formation in cases not caused by a third dentition.

OBJECTIVES The aim of this study was to examine the relationship between morphologic factors of mandibular protrusion patients and clinical indices of obstructive sleep apnea (OSA). METHODS Fifty-two Japanese patients divided into 2 groups: 1 jaw surgery group (30 patients) and 2 jaw surgery group (22 patients). Morphologic changes were studied using cephalograms taken before surgery and 1 year after surgery. Functional changes studied using impulse oscillometry and pulse oximetry during sleep, both of which are clinically useful measures in assessing OSA, taken before surgery and 1 year after surgery. RESULT Lower face cage area significantly decreased in 1 jaw group than in 2 jaw group patients. Positive significant correlation was found between changes in 3% oxygen desaturation index (ODI) and changes of tongue area and vertical position of the hyoid bone in 1 jaw surgery group. Multiple regression analysis indicates that tongue area and airway area were independently significant predictors of 3% ODI in 1 jaw group patients. CONCLUSION In 2 jaw surgery, maxillary surgery compensated for the effect of mandibular setback surgery. Mandibular setback surgery to mandibular protrusion patients was performed within the range of adequate movement distance, but precautions for risk of postoperative obstructive sleep apnea syndrome should be considered.

There have been some reports concerning supernumerary teeth located in the maxilla; however, eruption into the nasal cavity leading to a dentigerous cyst is quite rare. The authors report a 6-year-old boy who was noticed to have a foreign body in the right nasal cavity during a school physical examination. There was no subjective nasal symptom, such as bleeding or a feeling of rhinostenosis. On radiological examination, a tooth-like radiopaque object was noted beneath the surface of the right nasal floor, and part of a supernumerary tooth in the left maxillary incisor area was also seen. Ectopic supernumerary teeth were diagnosed and removed under general anesthesia. Histopathological examination identified the tooth-like structure as a mature tooth, and the isolated tissue in the nasal cavity was coated by soft tissue, confirmed as a feature of a cyst. Three years after surgery, the patient was doing well and had healed uneventfully.

Objective Fibrous dysplasia of bone (FD) is a benign developmental disorder of bone in which normal bone is replaced by fibrous tissue, containing trabeculae of immature woven bone. This disease has been classified into three types: monostotic (MFD), polyostotic (PFD), and McCune–Albright syndrome (MAS). Occurring in multiple adjacent craniofacial bones (craniofacial FD) is considered to be an MFD lesion. Surgical reduction is performed for form revision after having observed the progress until adulthood, but postoperative recurrence cases are not rare, there is a report of malignant progression to osteosarcoma, and the clinical phenotypes are generally various. This study aimed to identify the relationship between clinical futures of FD and expression of bone morphogenetic protein (BMP) subtypes. Methods We studied 10 cases (9 MFD cases and 1 McCune–Albright syndrome case) diagnosed with fibrous dysplasia in our hospital, and immunohistochemical examinations of the excised sample from each case with anti-BMP-2, 4, 6, and 7 antibodies were performed. Results 5 MFD cases and 1 MAS case underwent reoperation, and the case that had the most number of operations was 9 times in the MFD case. Malignant transformations were not identified. BMP-4, 6, and 7 expressions were positive in all cases, but the expression of BMP-2 was positive only in 6 MFD cases. BMP-2 negative cases tended to undergo reoperation. Conclusions The variable expression of BMP-2 demonstrated in the current study was suggested to be a useful indication for clinical activity and convalescence of the lesions.

Purpose The aim of this study was to examine and compare morphological and functional outcomes after either isolated mandibular setback or bimaxillary surgery in males and females. Materials and methods A retrospective study was done on 52 patients, in whom surgical correction for mandibular prognathism was performed either by isolated mandibular setback (30 cases) or by bimaxillary surgery (22 cases). Morphological changes were studied using cephalograms and functional changes studied using impulse oscillometry (IOS) taken before surgery (T0), 3 months (T1) and 1 year after surgery (T2). Also 3% oxygen desaturation index (ODI) was measured at T0 and T2. Result Posterior airway space decreased significantly in both groups and both sexes but more so in males after mandibular setback surgery and in females after bimaxillary surgery. Changes in supine R20 (central airway resistance at 20 Hz) and supine R5 (total airway resistance at 5 Hz) in IOS statistically significantly increased in the period T0–T1 in males compared with females after mandibular setback surgery (p Conclusion Gender dimorphism is present according to morphological and functional outcomes, with males at a higher risk for obstructive sleep apnea (OSA) after mandibular setback surgery and females after bimaxillary surgery; however, compensatory changes act as a barrier against this.

Adult Cebpb KO mice incisors present amelogenin-positive epithelium pearls, enamel and dentin allopathic hyperplasia, fewer Sox2-positive cells in labial cervical loop epitheliums, and reduced Sox2 expression in enamel epithelial stem cells. Thus, Cebpb acts upstream of Sox2 to regulate stemness. In this study, Cebpb KO mice demonstrated cementum-like hard tissue in dental pulp, loss of polarity by ameloblasts, enamel matrix in ameloblastic layer, and increased expression of epithelial-mesenchymal transition (EMT) markers in a Cebpb knockdown mouse enamel epithelial stem cell line. Runx2 knockdown in the cell line presented a similar expression pattern. Therefore, the EMT enabled disengaged odontogenic epithelial stem cells to develop supernumerary teeth. Cebpb and Runx2 knockdown in the cell line revealed higher Biglycan and Decorin expression, and Decorin-positive staining in the periapical region, indicating their involvement in supernumerary tooth formation. Cebpb and Runx2 acted synergistically and played an important role in the formation of supernumerary teeth in adult incisors.

This study aimed to carry out a case-control research study to assess occurrence of clicking of the temporomandibular joint (TMJ) in order to establish the relationship between TMJ clicking and the genotype of “ANKH inorganic pyrophosphate transport regulator” (ANKH) polymorphisms. A sample of 41 first-year dental residents was selected. Each was examined using standard clinical procedures and genotyping techniques. The participation rate was 91.8 %. The prevalence of TMJ clicking was 51.2 % (95 % CI: 35.7–66.7 %). Occurrence of TMJ clicking was not related to age, gender and genotypes of ANKH-OR as well as ANKH-TR polymorphisms (p ≥ 0.165). A similar distribution of ANKH genotypes in TMJ clicking and asymptomatic individuals has been demonstrated by this study. A high percentage of TMJ clicking has been confirmed. Future investigations are indicated.

Objectives: This investigation aimed to conduct a case-control study of mandibular morphology and dental anomalies to propose a relationship between mandibular/dental phenotypes and deficiency of CCAAT/enhancer-binding protein beta (CEBPB). Materials and methods: Skulls of CEBPB(-/-) , CEBPB(+/-) and CEBPB(+/+) mice were inspected with micro-computed tomography. Mandibular morphology was assessed with a method of Euclidean distance matrix analysis. Results: Elongation of the coronoid process was identified in CEBPB(+/-) (P ≤ 0.046) and CEBPB(-/-) 12-month-olds (P ≤ 0.028) but not in 14-day-olds (P ≥ 0.217) and 0-day-olds (P ≥ 0.189) of either genotype. Formation of supernumerary teeth in CEBPB(-/-) adult mice was demonstrated (χ(2 ) =( ) 6.00, df = 1, P = 0.014). Conclusions: CEBPB deficiency was related to elongation of the coronoid process and formation of supernumerary teeth. The mandibular and dental phenotypes of CEBPB deficiency were unseen by the 14th day after birth. Future investigations into the influence of CEBPB on mandibular and dental development are needed.

Background Wnt5a and Mrfzb1 genes are involved in the regulation of tooth size, and their expression levels are similar to that of Bmp7 during morphogenesis, including during the cap and early bell stages of tooth formation. We previously reported that Usag-1-deficient mice form supernumerary maxillary incisors. Thus, we hypothesized that BMP7 and USAG-1 signaling molecules may play important roles in tooth morphogenesis. In this study, we established double genetically modified mice to examine the in vivo inter-relationships between Bmp7 and Usag-1. Results We measured the volume and cross-sectional areas of the mandibular incisors using micro-computed tomography (micro-CT) in adult Bmp7- and Usag-1-LacZ knock-in mice and their F2 generation upon interbreeding. The mandibular incisors of adult Bmp7+/− mice were significantly larger than those of wild-type (WT) mice. The mandibular incisors of adult Usag-1−/− mice were the largest of all genotypes examined. In the F2 generation, the effects of these genes were additive; Bmp7+/− was most strongly associated with the increase in tooth size using generalized linear models, and the total area of mandibular supernumerary incisors of Usag-1−/−Bmp7+/− mice was significantly larger than that of Usag-1−/−Bmp7 +/+ mice. At embryonic day 15 (E15), BrdU assays demonstrated that the labeling index of Bmp7+/− embryos was significantly higher than that of WT embryos in the cervical loop. Additionally, the labeling index of Usag-1−/− embryos was significantly the highest of all genotypes examined in dental papilla. Conclusions Bmp7 heterozygous mice exhibited significantly increased tooth sizes, suggesting that tooth size was controlled by specific gene expression. Our findings may be useful in applications of regenerative medicine and dentistry.

Supernumerary teeth and tooth agenesis are common morphological anomalies in humans. We previously obtained evidence that supernumerary maxillary incisors form as a result of the successive development of the rudimentary maxillary incisor tooth germ in Usag-1 null mice. The development of tooth germs is arrested in Runx2 null mice, and such mice also exhibit lingual epithelial buds associated with the upper molars and incisors. The aim of this study is to investigate the potential crosstalk between Usag-1 and Runx2 during tooth development. In the present study, three interesting phenomena were observed in double null Usag-1-/-/Runx2-/- mice: the prevalence of supernumerary teeth was lower than in Usag-1 null mice; tooth development progressed further compared than in Runx2 null mice; and the frequency of molar lingual buds was lower than in Runx2 null mice. Therefore, we suggest that RUNX2 and USAG-1 act in an antagonistic manner. The lingual bud was completely filled with odontogenic epithelial Sox2-positive cells in the Usag-1+/+/Runx2-/- mice, whereas almost no odontogenic epithelial Sox2-positive cells contributed to supernumerary tooth formation in the rudimentary maxillary incisors of the Usag-1-/-/Runx2+/+ mice. Our findings suggest that RUNX2 directly or indirectly prevents the differentiation and/or proliferation of odontogenic epithelial Sox2-positive cells. We hypothesize that RUNX2 inhibits the bone morphogenetic protein (BMP) and/or Wnt signaling pathways regulated by USAG-1, whereas RUNX2 expression is induced by BMP signaling independently of USAG-1.

[Extract] The tooth is a complex organ that consists of enamel, dentin, cementum, and pulp. Missing teeth is frequently occurring problem in aging populations. To treat these defects, the current approach involves prostheses, autotransplantation, and dental implants. The exploration of new strategies for tooth replacement has become a hot topic. Using the foundations of experimental embryology, developmental and molecular biology, tooth regeneration is becoming realistic possibility. Several different methods have been proposed to achieve biological tooth replacement. These include scaffold-based tooth regeneration, cell pellet engineering, stimulation of the formation of a third dentition, and gene-manipulated tooth regeneration. The idea that a third dentition might be locally induced to replace missing teeth is an attractive concept (Young et al., 2005; Edward & Mason, 2006; Takahashi et al., 2008, 2013). This approach is generally presented in terms of adding molecules to induce de novo tooth initiation in the mouth. Tooth development is the result of reciprocal and reiterative signaling between oral ectoderm-derived dental epithelium and cranial neural crest cell-derived dental mesenchyme under genetic control (Thesleff, 2006). More than 200 genes are known to be expressed during tooth development (http://bite-it.helsinki.fi/). A number of mouse mutants are now starting to provide some insights into the mechanisms of supernumerary tooth formation. Multiple supernumerary teeth may have genetic components in their etiology and partially represent the third dentition in humans. Such candidate molecules might be those that are involved in embryonic tooth induction, in successional tooth formation, or in the control of the number of teeth. This means that it may be possible to induce de novo tooth formation by the in situ repression or activation of a single candidate molecule. In this review, we provide an overview of the collective knowledge of tooth regeneration, especially regarding the control of the number of teeth for molecularly targeted therapy by the stimulation of a third dentition.

[Extract] The tooth is a complex biological organ that consists of multiple tissues, including enamel, dentin, cementum, and pulp. Missing teeth is a common and frequently occurring problem in aging populations. To treat these defects, the current approach involves fixed or removable prostheses, autotransplantation, and dental implants. The exploration of new strategies for tooth replacement has become a hot topic. Using the foundations of experimental embryology, developmental and molecular biology, and the principles of biomimetics, tooth regeneration is becoming a realistic possibility. Several different methods have been proposed to achieve biological tooth replacement[1-8]. These include scaffold-based tooth regeneration, cell pellet engineering, chimeric tooth engineering, stimulation of the formation of a third dentition, and gene-manipulated tooth regeneration. The idea that a third dentition might be locally induced to replace missing teeth is an attractive concept[5,8,9]. This approach is generally presented in terms of adding molecules to induce de novo tooth initiation in the mouth. It might be combined with gene-manipulated tooth regeneration; that is, endogenous dental cells in situ can be activated or repressed by a gene-delivery technique to produce a tooth. Tooth development is the result of reciprocal and reiterative signaling between oral ectoderm-derived dental epithelium and cranial neural crest cell-derived dental mesenchyme under genetic control [10-12]. More than 200 genes are known to be expressed during tooth development (http://bite-it.helsinki.fi/). A number of mouse mutants are now starting to provide some insights into the mechanisms of supernumerary tooth formation. Multiple supernumerary teeth may have genetic components in their etiology and partially represent the third dentition in humans. Such candidate molecules or genes might be those that are involved in embryonic tooth induction, in successional tooth formation, or in the control of the number of teeth. This means that it may be possible to induce de novo tooth formation by the in situ repression or activation of a single candidate gene. In this review, we present an overview of the collective knowledge of tooth regeneration, especially regarding the control of the number of teeth for gene therapy by the stimulation of a third dentition.

Background. The expression term of the gene transfected in cells needs to belong enough inorder to make a gene therapy clinically effective. The controlled release of the transfected gene can be utilized. The new biodegradable hydrogel material created by 20 w/w% aldehyded dextran and 10 w/w%ε-poly(L-lysine) (ald-dex/PLL) was developed. We examined whether it could be as a nonviral carrier of the gene transfer.Methods. A plasmid (Lac-Z) was mixed with ald-dex/PLL. Anin vitrostudy was performed to assess the expression of Lac-Z with X-gal stain after gene transfer into the cultured 293 cells and bone marrow cells. As a control group, PLL was used as a cationic polymer.Results. We confirmed that the transfection efficiency of the ald-dex/PLL had a higher transfection efficiency than PLL in 293 cells (plasmid of 2 μg: ald-dex/PLL 1.1%, PLL 0.23%, plasmid of 16 μg: ald-dex/PLL 1.23%, PLL 0.48%). In bone marrow cells, we confirmed the expression of Lac-Z by changing the quantity of aldehyded dextran. In the groups using ald-dextran of the quantity of 1/4 and 1/12 of PLL, their transfection efficiency was 0.43% and 0.41%, respectively.Conclusions. This study suggested a potential of using ald-dex/PLL as a non-carrier for gene transfer.

Objectives: This study aimed to carry out a histological examination of the temporomandibular joint (TMJ) in ank mutant mice and to identify polymorphisms of the human ANKH gene in order to establish the relationship between the type of temporomandibular disorders (TMD) and ANKH polymorphisms. Materials and Methods: Specimens from the TMJ of ank mutant and wild-type mice were inspected with a haematoxylin and eosin staining method. A sample of 55 TMD patients were selected. Each was examined with standard clinical procedures and genotyping techniques. Results: The major histological finding in ank mutant mice was joint space narrowing. Within TMD patients, closed lock was more prevalent among ANKH-OR homozygotes (p = 0.011, OR = 7.7, 95% CI 1.6–36.5) and the elder (p = 0.005, OR = 2.4, 95% CI 1.3–4.3). Conclusions: Fibrous ankylosis was identified in the TMJ of ank mutant mice. In the human sample, ANKH-OR polymorphism was found to be a genetic marker associated with TMJ closed lock. Future investigations correlating genetic polymorphism to TMD are indicated.

Maxillofacial dysmorphogenesis is found in 5% of the population. To begin to understand the mechanisms required for maxillofacial morphogenesis, we employed the inhibitors of the differentiation 2 (Id2) knock-out mouse model, in which Id proteins, members of the regulator of basic helix-loop-helix (bHLH) transcription factors, modulate cell proliferation, apoptosis, and differentiation. We now report that spatially-restricted growth defects are localized at the skull base of Id2 KO mice. Curiously, at birth, neither the mutant Id2 KO nor wild-type (WT) mice differed, based upon cephalometric and histological analyses of cranial base synchondroses. In postnatal week 2, a narrower hypertrophic zone and an inhibited proliferative zone in presphenoid synchondrosis (PSS) and spheno-occipital synchondrosis (SOS) with maxillary hypoplasia were identified in the Id2 mutant mice. Complementary studies revealed that exogenous bone morphogenetic proteins (BMPs) enhanced cartilage growth, matrix deposition, and chondrocyte proliferation in the WT but not in the mutant model. Id2-deficient chondrocytes expressed more Smad7 transcripts. Based on our results, we assert that Id2 plays an essential role, acting downstream of BMP signaling, to regulate cartilage formation at the postnatal stage by enhancing BMP signals through inhibiting Smad7 expression. As a consequence, abnormal endochondral ossification was observed in cranial base synchondroses during the postnatal growth period, resulting in the clinical phenotype of maxillofacial dysmorphogenesis.

Bone morphogenetic proteins (BMPs) are highly conserved signaling molecules that are part of the transforming growth factor (TGF)-beta superfamily, and function in the patterning and morphogenesis of many organs including development of the dentition. The functions of the BMPs are controlled by certain classes of molecules that are recognized as BMP antagonists that inhibit BMP binding to their cognate receptors. In this study we tested the hypothesis that USAG-1 (uterine sensitization-associated gene-1) suppresses deciduous incisors by inhibition of BMP-7 function. We learned that USAG-1 and BMP-7 were expressed within odontogenic epithelium as well as mesenchyme during the late bud and early cap stages of tooth development. USAG-1 is a BMP antagonist, and also modulates Wnt signaling. USAG-1 abrogation rescued apoptotic elimination of odontogenic mesenchymal cells. BMP signaling in the rudimentary maxillary incisor, assessed by expressions of Msx1 and Dlx2 and the phosphorylation of Smad protein, was significantly enhanced. Using explant culture and subsequent subrenal capsule transplantation of E15 USAG-1 mutant maxillary incisor tooth primordia supplemented with BMP-7 demonstrated in USAG-1+/- as well as USAG-1-/- rescue and supernumerary tooth development. Based upon these results, we conclude that USAG-1 functions as an antagonist of BMP-7 in this model system. These results further suggest that the phenotypes of USAG-1 and BMP-7 mutant mice reported provide opportunities for regenerative medicine and dentistry.

Background Although runt-related transcription factor 2 (RUNX2) has been considered a determinant of cleidocranial dysplasia (CCD), some CCD patients were free of RUNX2 mutations. CCAAT/enhancer-binding protein beta (Cebpb) is a key factor of Runx2 expression and our previous study has reported two CCD signs including hyperdontia and elongated coronoid process of the mandible in Cebpb deficient mice. Following that, this work aimed to conduct a case-control study of thoracic, zygomatic and masticatory muscular morphology to propose an association between musculoskeletal phenotypes and deficiency of Cebpb, using a sample of Cebpb-/-, Cebpb+/- and Cebpb+/+ adult mice. Somatic skeletons and skulls of mice were inspected with soft x-rays and micro-computed tomography (μCT), respectively. Zygomatic inclination was assessed using methods of coordinate geometry and trigonometric function on anatomic landmarks identified with μCT. Masseter and temporal muscles were collected and weighed. Expression of Cebpb was examined with a reverse transcriptase polymerase chain reaction (RT-PCR) technique. Results Cebpb-/- mice displayed hypoplastic clavicles, a narrow thoracic cage, and a downward tilted zygomatic arch (p