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Background/Objectives: Tooth agenesis (TA) is among the most common malformations in humans. Although several causative mutations have been described, the genetic cause often remains elusive. Here, we test whether whole genome sequencing (WGS) could bridge this diagnostic gap. Methods: In four families with TA, we assessed the dental phenotype using the Tooth Agenesis Code after intraoral examination and radiographic and photographic documentation. We performed WGS of index patients and subsequent segregation analysis. Results: We identified two variants of uncertain significance (a potential splice variant in PTH1R, and a 2.1 kb deletion abrogating a non‐coding element in FGF7) and three pathogenic variants: a novel frameshift in the final exon of PITX2, a novel deletion in PAX9, and a known nonsense variant in WNT10A. Notably, the FGF7 variant was found in the patient, also featuring the WNT10A variant. While mutations in PITX2 are known to cause Axenfeld‐Rieger syndrome 1 (ARS1) predominantly featuring ocular findings, accompanied by dental malformations, we found the PITX2 frameshift in a family with predominantly dental and varying ocular findings. Conclusion: Severe TA predicts a genetic cause identifiable by WGS. Final exon PITX2 frameshifts can cause a predominantly dental form of ARS1. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the regulatory mechanisms governing dental mesenchymal cell commitment during tooth development, focusing on odontoblast differentiation and the role of epigenetic regulation in this process. We performed single-cell RNA sequencing (scRNA-seq) of dental cells from embryonic day 14.5 (E14.5) mice to understand the heterogeneity of developing tooth germ cells. Computational analyses including gene regulatory network (GRN) assessment were conducted. We validated our findings using immunohistochemistry (IHC) and in vitro loss-of-function analyses using the DNA methyltransferase 1 (DNMT1) inhibitor Gsk-3484862 in primary dental mesenchymal cells (DMCs) isolated from E14.5 mouse tooth germs. Bulk RNA-seq of Gsk-3484862-treated DMCs was performed to identify potential downstream targets of DNMT1. scRNA-seq analysis revealed diverse cell populations within the tooth germs, including epithelial, mesenchymal, immune, and muscle cells. Using single-cell regulatory network inference and clustering (SCENIC), we identified Dnmt1 as a key regulator of early odontoblast development. IHC analysis showed the ubiquitous expression of DNMT1 in the dental papilla and epithelium. Bulk RNA-seq of cultured DMCs showed that Gsk-3484862 treatment upregulated odontoblast-related genes, whereas genes associated with cell division and the cell cycle were downregulated. Integrated analysis of bulk RNA-seq data with scRNA-seq SCENIC profiles was used to identify the potential Dnmt1 target genes. Dnmt1 may negatively affect odontoblast commitment and differentiation during tooth development. These findings contribute to a better understanding of the molecular mechanisms underlying tooth development and future development of hard-tissue regenerative therapies. • This study aimed to understand key regulators of dental mesenchymal cell development. • scRNA-seq identified Dnmt1 as a regulator in undifferentiated mesenchymal cells. • Dnmt1 may regulate odontoblast commitment and differentiation during tooth development. [ABSTRACT FROM AUTHOR]

Thanks to their exceptional diversity, teeth are among the most distinctive features of vertebrates. Parameters such as tooth size, shape, number, identity, and implantation can have substantial implications for the ecology and certain social behaviors of toothed species. Despite decades of research primarily focused on mammalian dentition, particularly using the laboratory mouse model, squamate reptiles ("lizards" and snakes) offer a wide array of tooth types and dentition variations. This diversity, which includes differences in size, shape, function, and replacement capacity, provides invaluable opportunities for investigating these fundamental properties. The central bearded dragon (Pogona vitticeps), a popular pet species with well-established husbandry practices, is of particular interest. It features a broad spectrum of morphs and spontaneous mutants and exhibits a wide range of heterodont phenotypes, including variation in the size, shape, number, implantation, and renewal of teeth at both posterior and anterior positions. These characteristics position the species as a crucial model organism for developmental studies in tooth research and for gaining deeper insights into evolutionary patterns of vertebrate dentitions. In this article, we provide an overview of the current understanding of squamate dentition, its diversity, development, and replacement. Furthermore, we discuss the significant advantages offered by squamate species as model organisms for investigating the evolutionary and developmental aspects of vertebrate dentition. [Display omitted] • Teeth are a hallmark features of vertebrates • Large ecomorphological diversity of squamate dentitions remains largely unexplored • Central bearded dragon is an emerging model for evo-devo and dental research • Squamate dentition: diversity, development, and replacement importance • Squamate models offer crucial insights into vertebrate tooth evolution [ABSTRACT FROM AUTHOR]

Ameloblasts are the specialized dental epithelial cell type responsible for enamel formation. Following completion of enamel development in humans, ameloblasts are lost and biological repair or regeneration of enamel is not possible. In the past, in vitro models to study dental epithelium and ameloblast biology were limited to freshly isolated primary cells or immortalized cell lines, both with limited translational potential. In recent years, large strides have been made with the development of induced pluripotent stem cell and organoid models of this essential dental lineage – both enabling modeling of human dental epithelium. Upon induction with several different signaling factors (such as transforming growth factor and bone morphogenetic proteins) these models display elevated expression of ameloblast markers and enamel matrix proteins. The advent of 3D bioprinting, and its potential combination with these advanced cellular tools, is poised to revolutionize the field – and its potential for tissue engineering, regenerative and personalized medicine. As the advancements in these technologies are rapidly evolving, we evaluate the current state-of-the-art regarding in vitro cell culture models of dental epithelium and ameloblast lineage with a particular focus toward their applicability for translational tissue engineering and regenerative/personalized medicine. Future perspectives for in vitro modeling of dental epithelium and ameloblasts. Development of iPSC and organoid models that can reliably generate dental epithelium and ameloblast-like cells, together with advances in 3D bioprinting, provide promising tools for enamel research. Advanced models will provide new avenues for development of enamel repair/regeneration approaches, for testing of dental materials or drugs, studying host-pathogen and/or cell-cell interactions, in vitro modeling of enamel diseases (e.g. amelogenesis imperfecta) and developing novel insights in fundamental tooth biology (e.g. regulation of amelogenesis, lineage specification). Abbreviations: iPSC: induced pluripotent stem cells; TO: tooth organoids; DE: dental epithelium; AB: ameloblast. [ABSTRACT FROM AUTHOR]

Objectives: The aim of this study was to investigate the impact of birth weight on tooth development in children aged 7–8 years. Materials and methods: This retrospective cohort study comprised 75 children born at Bint Al-Huda Hospital, Bojnurd, in 2013–2014. The children were categorized into three groups based on their birth weight: Normal Birth Weight (NBW), Low Birth Weight (LBW), and Very Low Birth Weight (VLBW). Panoramic radiographs were taken for orthodontic examination, and Demirjian’s 8-teeth method was employed to determine dental age. The study compared dental and chronological age within each group. Data analysis utilized SPSS software version 26, employing One-way ANOVA and chi-square tests. Statistical significance was set at P ≤ 0.05. Results: The mean difference in dental and chronological age for Very Low Birth Weight (VLBW) children was 0.22 ± 0.44 years, for Low Birth Weight (LBW) children it was 0.19 ± 0.45 years, and for Normal Birth Weight (NBW) children, it was 0.08 ± 0.46 years. Although the mean difference decreased with increasing birth weight, this trend did not achieve statistical significance (P = 0.55). Furthermore, no significant differences were observed between the weight groups (P = 0.529) or genders (P = 0.191). [ABSTRACT FROM AUTHOR]

Epigenetic modulation, including histone modification, alters gene expression and controls cell fate. Histone deacetylases (HDACs) are identified as important regulators of dental pulp cell (DPC) mineralisation processes. Currently, there is a paucity of information regarding the nature of histone modification and HDAC expression in the dentine–pulp complex during dentinogenesis. The aim of this study was to investigate post-translational histone modulation and HDAC expression during DPC mineralisation and the expression of Class I/II HDACs during tooth development and in adult teeth. HDAC expression (isoforms −1 to −6) was analysed in mineralising primary rat DPCs using qRT-PCR and Western blot with mass spectrometry being used to analyse post-translational histone modifications. Maxillary molar teeth from postnatal and adult rats were analysed using immunohistochemical (IHC) staining for HDACs (1–6). HDAC-1, -2, and -4 protein expression increased until days 7 and 11, but decreased at days 14 and 21, while other HDAC expression increased continuously for 21 days. The Class II mineralisation-associated HDAC-4 was strongly expressed in postnatal sample odontoblasts and DPCs, but weakly in adult teeth, while other Class II HDACs (-5, -6) were relatively strongly expressed in postnatal DPCs and adult odontoblasts. Among Class I HDACs, HDAC-1 showed high expression in postnatal teeth, notably in ameloblasts and odontoblasts. HDAC-2 and -3 had extremely low expression in the rat dentine–pulp complex. Significant increases in acetylation were noted during DPC mineralisation processes, while trimethylation H3K9 and H3K27 marks decreased, and the HDAC-inhibitor suberoylanilide hydroxamic acid (SAHA) enhanced H3K27me3. These results highlight a dynamic alteration in histone acetylation during mineralisation and indicate the relevance of Class II HDAC expression in tooth development and regenerative processes. [ABSTRACT FROM AUTHOR]

Background: Mutations of human WNT10A are associated with odonto‐ectodermal dysplasia syndromes. Here, we present analyses of wnt10a loss‐of‐function mutants in the zebrafish. Results: wnt10a mutant zebrafish embryos display impaired tooth development and a collapsing median fin fold (MFF). Rescue experiments show that wnt10a is essential for MFF maintenance both during embryogenesis and later metamorphosis. The MFF collapse could not be attributed to increased cell death or altered proliferation rates of MFF cell types. Rather, wnt10a mutants show reduced expression levels of dlx2a in distal‐most MFF cells, followed by compromised expression of col1a1a and other extracellular matrix proteins encoding genes. Transmission electron microscopy analysis shows that although dermal MFF compartments of wnt10a mutants initially are of normal morphology, with regular collagenous actinotrichia, positioning of actinotrichia within the cleft of distal MFF cells becomes compromised, coinciding with actinotrichia shrinkage and MFF collapse. Conclusions: MFF collapse of wnt10a mutant zebrafish is likely caused by the loss of distal properties in the developing MFF, strikingly similar to the proposed molecular pathomechanisms underlying the teeth defects caused by the loss of Wnt10 in fish and mammals. In addition, it points to thus fur unknown mechanisms controlling the linear growth and stability of actinotrichia and their collagen fibrils. Key Findings: Description of first genetic zebrafish mutant in Wnt10a.Wnt10a is required for zebrafish tooth development.Wnt10a is required for maintenance of embryonic median fin fold (MFF).Wnt10a is required for formation of adult unpaired fins during metamorphosis.Wnt10a signaling is required for distal specification of epithelial MFF cells and persistent collagen expression and actinotrichia formation. [ABSTRACT FROM AUTHOR]

This review delves into the roles of glycosaminoglycans (GAGs), integral components of proteoglycans, in tooth development. Proteoglycans consist of a core protein linked to GAG chains, comprised of repeating disaccharide units. GAGs are classified into several types, such as hyaluronic acid, heparan sulfate, chondroitin sulfate, dermatan sulfate, and keratan sulfate. Functioning as critical macromolecular components within the dental basement membrane, these GAGs facilitate cell adhesion and aggregation, and play key roles in regulating cell proliferation and differentiation, thereby significantly influencing tooth morphogenesis. Notably, our recent research has identified the hyaluronan-degrading enzyme Transmembrane protein 2 (Tmem2) and we have conducted functional analyses using mouse models. These studies have unveiled the essential role of Tmem2-mediated hyaluronan degradation and its involvement in hyaluronan-mediated cell adhesion during tooth formation. This review provides a comprehensive summary of the current understanding of GAG functions in tooth development, integrating insights from recent research, and discusses future directions in this field. [ABSTRACT FROM AUTHOR]

Objectives: To identify the molecular etiology of distinct dental anomalies found in eight Thai patients and explore the mutational effects on cellular functions. Materials and Methods: Clinical and radiographic examinations were performed for eight patients. Whole exome sequencing, mutant protein modelling, qPCR, western blot analysis, scratch assays, immunofluorescence, confocal analysis, in situ hybridization, and scanning electron micrography of teeth were done. Results: All patients had molars with multiple supernumerary cusps, single‐cusped premolars, and a reduction in root number. Mutation analysis highlighted a heterozygous c.865A>G; p.Ile289Val mutation in CACNA1S in the patients. CACNA1S is a component of the slowly inactivating L‐type voltage‐dependent calcium channel. Mutant protein modeling suggested that the mutation might allow leakage of Ca2+ or other cations, or a tightening, to restrict calcium flow. Immunohistochemistry analysis showed expression of Cacna1s in the developing murine tooth epithelium during stages of crown and root morphogenesis. In cell culture, the mutation resulted in abnormal cell migration of transfected CHO cells compared to wildtype CACNA1S, with changes to the cytoskeleton and markers of focal adhesion. Conclusions: The malformations observed in our patients suggest a role for calcium signaling in organization of both cusps and roots, affecting cell dynamics within the dental epithelium. [ABSTRACT FROM AUTHOR]

Objective: Epigenetic regulation plays important role in stem cell maintenance. Ptip was identified as epigenetic regulator, but the role in dental progenitor cells remains unclear. Subjects and Methods: Dental mesenchymal progenitor cells were targeted by Sp7‐icre and visualized in mTmG; Sp7‐icre mice. The Ptipf/f; Sp7‐icre mice were generated and the phenotype of incisors and molars were shown by micro‐computerized tomography, scanning electron microscope, hematoxylin & eosin staining, and immunofluorescence. Dental mesenchymal progenitor cells were sorted by fluorescence‐activated cell sorting from lower incisors and RNA sequencing was performed. Results: The Sp7‐icre targets dental mesenchymal progenitor cells in incisors and molars. The Ptipf/f; Sp7‐icre mice showed spontaneous fractures in the cusp of upper incisors and lower incisors at 3 weeks (w), compensative overgrowth of lower incisors at 1 month (M), and overgrowth extended to the outside at 2 M. The molars showed shortened roots. The functions of odontoblasts and dental mesenchymal progenitor cells were impaired. Mechanically, loss of Ptip activates the Wnt pathway and upregulates the expression of Wls in dental mesenchymal progenitor cells. Also, the regenerative ability of lower incisors was significantly impaired. Conclusion: We first demonstrated that Ptip was crucial for tooth development via regulating Wnt signaling. [ABSTRACT FROM AUTHOR]

Tooth development and regeneration are regulated through a complex signaling network. Previous studies have focused on the exploration of intracellular signaling regulatory networks, but the regulatory roles of extracellular networks have only been revealed recently. Proteoglycans, which are essential components of the extracellular matrix (ECM) and pivotal signaling molecules, are extensively involved in the process of odontogenesis. Proteoglycans are composed of core proteins and covalently attached glycosaminoglycan chains (GAGs). The core proteins exhibit spatiotemporal expression patterns during odontogenesis and are pivotal for dental tissue formation and periodontium development. Knockout of core protein genes Biglycan, Decorin, Perlecan, and Fibromodulin has been shown to result in structural defects in enamel and dentin mineralization. They are also closely involved in the development and homeostasis of periodontium by regulating signaling transduction. As the functional component of proteoglycans, GAGs are negatively charged unbranched polysaccharides that consist of repeating disaccharides with various sulfation groups; they provide binding sites for cytokines and growth factors in regulating various cellular processes. In mice, GAG deficiency in dental epithelium leads to the reinitiation of tooth germ development and the formation of supernumerary incisors. Furthermore, GAGs are critical for the differentiation of dental stem cells. Inhibition of GAGs assembly hinders the differentiation of ameloblasts and odontoblasts. In summary, core proteins and GAGs are expressed distinctly and exert different functions at various stages of odontogenesis. Given their unique contributions in odontogenesis, this review summarizes the roles of proteoglycans and GAGs throughout the process of odontogenesis to provide a comprehensive understanding of tooth development. [ABSTRACT FROM AUTHOR]

Background: Children with neurodevelopmental disorders (NDDs) often have poor oral health and dental abnormalities. An increasing number of genes have been associated with neurodevelopmental conditions affecting the oral cavity, but the specific dental features associated with many genes remain unknown. Aim: To report the types and frequencies of dental manifestations in children with neurodevelopmental conditions of known genetic cause. Design: A 30‐question survey assesing ectodermal and dental features was administered through Simons Searchlight, with which formed a recontactable cohort of individuals with genetic NDDs often associated with autism spectrum disorder (ASD). Results: Data were collected from a largely paediatric population with 620 affected individuals across 39 genetic conditions and 145 unaffected siblings without NDDs for comparison. Drooling, difficulty accessing dental care, late primary teeth eruption, abnormal primary and permanent teeth formation, misshapen nails, and hair loss were more frequent in individuals with NDDs. Additionally, we evidenced an association between three new pathogenic gene variant/oral manifestation pairs: CSNK2A1/unusual primary teeth, DYRK1A/late primary teeth eruption, and PPP2R5D/sialorrhea. Conclusion: Our results demonstrate that genetic NDDs caused by mutations in CSNK2A1, DYRK1A, and PP2R5D are associated with unique dental manifestations, and knowledge of these features can be helpful to personalize dental care. [ABSTRACT FROM AUTHOR]

Mutations in PAX9 are the most common genetic cause of tooth agenesis (TA). The aim of this study was to systematically review the profiles of the TA and PAX9 variants and establish their genotype-phenotype correlation. Forty articles were eligible for 178 patients and 61 mutations (26 in frame and 32 null mutations). PAX9 mutations predominantly affected molars, mostly the second molar, and the mandibular first premolar was the least affected. More missing teeth were found in the maxilla than the mandible, and with null mutations than in-frame mutations. The number of missing teeth was correlated with the locations of the in-frame mutations with the C-terminus mutations demonstrating the fewest missing teeth. The null mutation location did not influence the number of missing teeth. Null mutations in all locations predominantly affected molars. For the in-frame mutations, a missing second molar was commonly associated with mutations in the highly conserved paired DNA-binding domain, particularly the linking peptide (100% prevalence). In contrast, C-terminus mutations were rarely associated with missing second molars and anterior teeth, but were commonly related to an absent second premolar. These finding indicate that the mutation type and position contribute to different degrees of loss of PAX9 function that further differentially influences the manifestations of TA. This study provides novel information on the correlation of the PAX9 genotype-phenotype, aiding in the genetic counseling for TA.

Orthodontic tooth movement (OTM) involves mechanical-biochemical signal transduction, which results in tissue remodeling of the tooth-periodontium complex and the movement of orthodontic teeth. The dynamic regulation of osteogenesis and osteoclastogenesis serves as the biological basis for remodeling of the periodontium, and more importantly, the prerequisite for establishing periodontal homeostasis. Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) are key effectors of the Hippo signaling pathway, which actively respond to mechanical stimuli during tooth movement. Specifically, they participate in translating mechanical into biochemical signals, thereby regulating periodontal homeostasis, periodontal remodeling, and tooth development. YAP and TAZ have widely been considered as key factors to prevent dental dysplasia, accelerate orthodontic tooth movement, and shorten treatment time. In this review, we summarize the functions of YAP and TAZ in regulating tooth development and periodontal remodeling, with the aim to gain a better understanding of their mechanisms of action and provide insights into maintaining proper tooth development and establishing a healthy periodontal and alveolar bone environment. Our findings offer novel perspectives and directions for targeted clinical treatments. Moreover, considering the similarities and differences in the development, structure, and physiology between YAP and TAZ, these molecules may exhibit functional variations in specific regulatory processes. Hence, we pay special attention to their distinct roles in specific regulatory functions to gain a comprehensive and profound understanding of their contributions. [ABSTRACT FROM AUTHOR]

Upon endoplasmic reticulum (ER) stress, inositol-requiring enzyme 1 (IRE1) is activated, which subsequently converts an unspliced X-box binding protein 1 (XBP1U) mRNA to a spliced mRNA that encodes a potent XBP1S transcription factor. XBP1S is essential for relieving ER stress and secretory cell differentiation. We previously established Twist2-Cre;Xbp1CS/+ mice that constitutively expressed XBP1S in the Twist2-expressing cells as well as in the cells derived from the Twist2-expressing cells. In this study, we analyzed the dental phenotype of Twist2-Cre;Xbp1CS/+ mice. We first generated a mutant Xbp1s minigene that corresponds to the recombinant Xbp1Δ26 allele (the Xbp1CS allele that has undergone Cre-mediated recombination) and confirmed that the Xbp1s minigene expressed XBP1S that does not require IRE1α activation in vitro. Consistently, immunohistochemistry showed that XBP1S was constitutively expressed in the odontoblasts and other dental pulp cells in Twist2-Cre;Xbp1CS/+ mice. Plain X-ray radiography and µCT analysis revealed that constitutive expression of XBP1S altered the dental pulp chamber roof- and floor-dentin formation, resulting in a significant reduction in dentin/cementum formation in Twist2-Cre;Xbp1CS/+ mice, compared to age-matched Xbp1CS/+ control mice. However, there is no significant difference in the density of dentin/cementum between these two groups of mice. Histologically, persistent expression of XBP1S caused a morphological change in odontoblasts in Twist2-Cre;Xbp1CS/+ mice. Nevertheless, in situ hybridization and immunohistochemistry analyses showed that continuous expression of XBP1S had no apparent effects on the expression of the Dspp and Dmp1 genes. In conclusion, these results support that sustained production of XBP1S adversely affected odontoblast function and dentin formation. [ABSTRACT FROM AUTHOR]

Mutations in PAX9 are the most common genetic cause of tooth agenesis (TA). The aim of this study was to systematically review the profiles of the TA and PAX9 variants and establish their genotype-phenotype correlation. Forty articles were eligible for 178 patients and 61 mutations (26 in frame and 32 null mutations). PAX9 mutations predominantly affected molars, mostly the second molar, and the mandibular first premolar was the least affected. More missing teeth were found in the maxilla than the mandible, and with null mutations than in-frame mutations. The number of missing teeth was correlated with the locations of the in-frame mutations with the C-terminus mutations demonstrating the fewest missing teeth. The null mutation location did not influence the number of missing teeth. Null mutations in all locations predominantly affected molars. For the in-frame mutations, a missing second molar was commonly associated with mutations in the highly conserved paired DNA-binding domain, particularly the linking peptide (100% prevalence). In contrast, C-terminus mutations were rarely associated with missing second molars and anterior teeth, but were commonly related to an absent second premolar. These finding indicate that the mutation type and position contribute to different degrees of loss of PAX9 function that further differentially influences the manifestations of TA. This study provides novel information on the correlation of the PAX9 genotype-phenotype, aiding in the genetic counseling for TA. [ABSTRACT FROM AUTHOR]

Background: Fibroblast growth factor receptor‐3 (FGFR3) gain‐of‐function mutations are linked to achondroplasia. Infigratinib, a FGFR1‐3 tyrosine kinase inhibitor, improves skeletal growth in an achondroplasia mouse model. FGFs and their receptors have critical roles in developing teeth, yet effects of infigratinib on tooth development have not been assessed. Dentoalveolar and craniofacial phenotype of Wistar rats dosed with low (0.1 mg/kg) and high (1.0 mg/kg) dose infigratinib were evaluated using micro‐computed tomography, histology, and immunohistochemistry. Results: Mandibular third molars were reduced in size and exhibited aberrant crown and root morphology in 100% of female rats and 80% of male rats at high doses. FGFR3 and FGF18 immunolocalization and extracellular matrix protein expression were unaffected, but cathepsin K (CTSK) was altered by infigratinib. Cranial vault bones exhibited alterations in dimension, volume, and density that were more pronounced in females. In both sexes, interfrontal sutures were significantly more patent with high dose vs vehicle. Conclusions: High dose infigratinib administered to rats during early stages affects dental and craniofacial development. Changes in CTSK from infigratinib in female rats suggest FGFR roles in bone homeostasis. While dental and craniofacial disruptions are not expected at therapeutic doses, our findings confirm the importance of dental monitoring in clinical studies. Key Findings: Prolonged exposure to high‐dose infigratinib, particularly in female rats, dramatically affects crown and root development when administered in earlier stages of tooth development.Exposure to infigratinib is dependent on maturation of liver enzymes.Changes in extracellular matrix proteins suggest role for FGFR3 in bone modeling/remodeling. [ABSTRACT FROM AUTHOR]

Background. Dental papilla cells (DPCs) are one of the key stem cells for tooth development, eventually forming dentin and pulp. Previous studies have reported that PER2 is expressed in a 24-hour oscillatory pattern in DPCs in vitro. In vivo, PER2 is highly expressed in odontoblasts (which are differentiated from DPCs). However, whether PER2 modulates the odontogenic differentiation of DPCs is uncertain. This research was to identify the function of PER2 in the odontogenic differentiation of DPCs and preliminarily explore its mechanisms. Methods. We monitored the expression of PER2 in DPCs differentiated in vivo. We used PER2 overexpression and knockdown studies to assess the role of PER2 in DPC differentiation and performed intracellular ATP content and reactive oxygen species (ROS) assays to further investigate the mechanism. Results. PER2 expression was considerably elevated throughout the odontoblastic differentiation of DPCs in vivo. Overexpressing Per2 boosted levels of odontogenic differentiation markers, such as dentin sialophosphoprotein (Dspp), dentin matrix protein 1 (Dmp1), and alkaline phosphatase (Alp), and enhanced mineralized nodule formation in DPCs. Conversely, the downregulation of Per2 inhibited the differentiation of DPCs. Additionally, downregulating Per2 further affected intracellular ATP content and ROS levels during DPC differentiation. Conclusion. Overall, we demonstrated that PER2 positively regulates the odontogenic differentiation of DPCs, and the mechanism may be related to mitochondrial function as shown by intracellular ATP content and ROS levels. [ABSTRACT FROM AUTHOR]

The dental pulp plays a crucial role in the long-term maintenance of tooth function. The progress of endodontic treatment and pulp tissue regeneration engineering has made pulp-regeneration therapy promising in clinical practice. However, the mechanisms of pulp regeneration and the role of dental stem cells in development and regeneration have not been fully elucidated. Bridging the gaps between clinical operation and basic research is urgently needed. With the application of single-cell sequencing technology in dental research, the landscapes of human dental pulp cells have begun being outlined. However, the specific cellular heterogeneity of dental pulp cells, especially that of dental stem cells, at different spatial and temporal levels, is still unclear. In this study, we used single-cell RNA sequencing analysis of pulp samples at four different developmental stages and combined the findings with immunohistochemical staining to explore the development of dental pulp and stem cells. The results revealed temporal changes in the proportion of pulp cells during development. For example, mononuclear phagocytes accounted for a higher proportion in early samples. Odontoblasts identified by DMP1 had a higher expression of ion channel-related and neurodevelopment-related genes. Subpopulations were identified in fibroblasts, odontoblasts, and mesenchymal stem cells. We identified a subclass of odontoblasts that expresses DGKI and RRBP1 present in early developmental samples. Population of earlier mesenchymal stem cells expressed the SEPTIN gene, which may have greater proliferative and differentiation potential. Furthermore, dental pulp stem cells can differentiate into two directions: mineralization and myogenesis. In summary, the specific cellular heterogeneity of dental pulp cells was revealed at different spatial and temporal levels. These findings may shed light on the mechanism of tooth development. The gene expression profile of developing pulp cells may help to select cells for regenerative engineering and improve the success of dental pulp regeneration. [ABSTRACT FROM AUTHOR]

Reports of natal and neonatal teeth have been well documented in the literature, although the presence of natal primary molars remains rare, and a clear management strategy does not exist. This is a case study of a female newborn delivered at a gestational age of 41 weeks 1 day to a 31-year-old G1P1001 mother. Apgar scores were 8 and 9 at 1 and 5 min, respectively. The infant was delivered by cesarean section and was admitted to the Mass General Hospital Newborn Nursery, where she received routine care. The patient had two posterior molars, which warranted consultation for oral maxillofacial surgery. Due to the gross mobility of the natal teeth and the risk of aspiration in a small breastfeeding newborn, the decision was made to extract the natal teeth immediately. Understanding the management of natal teeth is important for the pediatrician.

Background: The intricate process of tooth formation during embryonic development ensures sufficient nutrition for the growth of healthy dental tissues. Amelogenin and alkaline phosphatase (ALP) are serine proteinases secreted by the ameloblast during the transition and maturation phases of the amelogenesis process. Consumption of saltwater fish is predicted to increase the expression of amelogenin and ALP in ameloblast cells during tooth formation. Only now have the function of each gene, tooth-forming cells, and the proteins they produce in the biomolecular amelogenesis of tooth enamel, which began during prenatal development, been clarified. Purpose: This study aims to determine how saltwater fish powder affects the ability of mother mice to increase the expression of amelogenin and ALP in cell ameloblast. Methods: Using a completely randomized design, this study was experimental and aimed to examine the effects of sardine (Sardinella fimbriata), splendid ponyfish (Leiognathus splendens), and tuna (Euthynnus affinis) powder. As samples, twenty-four female mice (Mus musculus) were used. Two groups of mice were created: group 1 (2.14 mg/0.5 ml) and the control group. The expression of amelogenin and ALP was determined using immunohistochemistry (IHC) and t-test (p0.05). Results: Expression of ameloblast was significantly different between the treatment and control groups (p0.05). Conclusion: The consumption of saltwater fish reduces the amelogenin and ALP expressions of mouse fetal ameloblast cells during tooth development in vivo.

Epigenetic modulation, including histone modification, alters gene expression and controls cell fate. Histone deacetylases (HDACs) are identified as important regulators of dental pulp cell (DPC) mineralisation processes. Currently, there is a paucity of information regarding the nature of histone modification and HDAC expression in the dentine–pulp complex during dentinogenesis. The aim of this study was to investigate post-translational histone modulation and HDAC expression during DPC mineralisation and the expression of Class I/II HDACs during tooth development and in adult teeth. HDAC expression (isoforms −1 to −6) was analysed in mineralising primary rat DPCs using qRT-PCR and Western blot with mass spectrometry being used to analyse post-translational histone modifications. Maxillary molar teeth from postnatal and adult rats were analysed using immunohistochemical (IHC) staining for HDACs (1–6). HDAC-1, -2, and -4 protein expression increased until days 7 and 11, but decreased at days 14 and 21, while other HDAC expression increased continuously for 21 days. The Class II mineralisation-associated HDAC-4 was strongly expressed in postnatal sample odontoblasts and DPCs, but weakly in adult teeth, while other Class II HDACs (-5, -6) were relatively strongly expressed in postnatal DPCs and adult odontoblasts. Among Class I HDACs, HDAC-1 showed high expression in postnatal teeth, notably in ameloblasts and odontoblasts. HDAC-2 and -3 had extremely low expression in the rat dentine–pulp complex. Significant increases in acetylation were noted during DPC mineralisation processes, while trimethylation H3K9 and H3K27 marks decreased, and the HDAC-inhibitor suberoylanilide hydroxamic acid (SAHA) enhanced H3K27me3. These results highlight a dynamic alteration in histone acetylation during mineralisation and indicate the relevance of Class II HDAC expression in tooth development and regenerative processes.

Abstract Background Tooth development, as one of the major mineralized tissues in the body, require fine-tuning of mineralization micro-environment. The interaction between dental epithelium and mesenchyme plays a decisive role in this process. With epithelium–mesenchyme dissociation study, we found interesting expression pattern of insulin-like growth factor binding protein 3 (IGFBP3) in response to disruption of dental epithelium–mesenchyme interaction. Its action and related mechanisms as regulator of mineralization micro-environment during tooth development are investigated. Results Expressions of osteogenic markers at early stage of tooth development are significantly lower than those at later stage. BMP2 treatment further confirmed a high mineralization micro-environment is disruptive at early stage, but beneficial at later stage of tooth development. In contrast, IGFBP3's expression increased gradually from E14.5, peaked at P5, and decreased afterwards, demonstrating an inverse correlation with osteogenic markers. RNA-Seq and Co-immunoprecipitation showed that IGFBP3 regulates the Wnt/beta-catenin signaling pathway activity by enhancing DKK1 expression and direct protein–protein interaction. The suppression of the mineralization microenvironment effectuated by IGFBP3 could be reversed by the DKK1 inhibitor WAY-262611, further demonstrating that IGFBP3 exerted its influence via DKK1. Conclusion A deeper understanding of tooth development mechanisms is essential for tooth regeneration, which have great implications for dental care. The current study demonstrated that the IGFBP3 expression is regulated in accordance with the needs of the mineralization microenvironment during tooth development, and IGFBP3 exerts its modulating action on osteogenic/odontogenic differentiation of hDPSCs by DKK1-Wnt/ beta-catenin axis.

The role of regulated cell death in organ development, particularly the impact of non-apoptotic cell death, remains largely uncharted. Ferroptosis, a non-apoptotic cell death pathway known for its iron dependence and lethal lipid peroxidation, is currently being rigorously investigated for its pathological functions. The balance between ferroptotic stress (iron and iron-dependent lipid peroxidation) and ferroptosis supervising pathways (anti-lipid peroxidation systems) serves as the key mechanism regulating the activation of ferroptosis. Compared with other forms of regulated necrotic cell death, ferroptosis is critically related to the metabolism of lipid and iron which are also important in organ development. In our study, we examined the role of ferroptosis in organogenesis using an ex vivo tooth germ culture model, investigating the presence and impact of ferroptotic stress on tooth germ development. Our findings revealed that ferroptotic stress increased during tooth development, while the expression of glutathione peroxidase 4 (Gpx4), a crucial anti-lipid peroxidation enzyme, also escalated in dental epithelium/mesenchyme cells. The inhibition of ferroptosis was found to partially rescue erastin-impaired tooth morphogenesis. Our results suggest that while ferroptotic stress is present during tooth organogenesis, its effects are efficaciously controlled by the subsequent upregulation of Gpx4. Notably, an overabundance of ferroptotic stress, as induced by erastin, suppresses tooth morphogenesis.

The circadian clock plays a critical role in dentomaxillofacial development. Tooth biomineralization is characterized by the circadian clock; however, the mechanisms underlying the coordination of circadian rhythms with tooth development and biomineralization remain unclear. The p75 neurotrophin receptor (p75NTR) is a clock factor that regulates the oscillatory components of the circadian rhythm. This study aims to investigate the impact of p75NTR on the rhythmic mineralization of teeth and elucidate its underlying molecular mechanisms. We generated p75NTR knockout mice to examine the effects of p75NTR deficiency on tooth mineralization. Ectomesenchymal stem cells (EMSCs), derived from mouse tooth germs, were used for in vitro experiments. Results showed a reduction in tooth mineral density and daily mineralization rate in p75NTR knockout mice. Deletion of p75NTR decreased the expression of DMP1, DSPP, RUNX2, and ALP in tooth germ. Odontogenic differentiation and mineralization of EMSCs were activated by p75NTR. Histological results demonstrated predominant detection of p75NTR protein in odontoblasts and stratum intermedium cells during rapid formation phases of dental hard tissue. The mRNA expression of p75NTR exhibited circadian variations in tooth germs and EMSCs, consistent with the expression patterns of the core clock genes Bmal1 and Clock. The upregulation of BMAL1/CLOCK expression by p75NTR positively regulated the mineralization ability of EMSCs, whereas BMAL1 and CLOCK exerted a negative feedback regulation on p75NTR by inhibiting its promoter activity. Our findings suggest that p75NTR is necessary to maintain normal tooth biomineralization. Odontogenic differentiation and mineralization of EMSCs is regulated by the p75NTR-BMAL1/CLOCK signaling axis. These findings offer valuable insights into the associations between circadian rhythms, tooth development, and biomineralization.

Teeth consist of 3 mineralized tissues: enamel, dentin, and cementum. Tooth malformation, the most common craniofacial anomaly, arises from complex genetic and environmental factors affecting enamel structure, size, shape, and tooth eruption. Hyaluronic acid (HA), a primary extracellular matrix component, contributes to structural and physiological functions in periodontal tissue. Transmembrane protein 2 (TMEM2), a novel cell surface hyaluronidase, has been shown to play a critical role during embryogenesis. In this study, we demonstrate Tmem2 messenger RNA expression in inner enamel epithelium and presecretory, secretory, and mature ameloblasts. Tmem2 knock-in reporter mice reveal TMEM2 protein localization at the apical and basal ends of secretory ameloblasts. Micro–computed tomography analysis of epithelial-specific Tmem2 conditional knockout (Tmem2 - CKO) mice shows a significant reduction in enamel layer thickness and severe enamel deficiency. Enamel matrix protein expression was remarkably downregulated in Tmem2 - CKO mice. Scanning electron microscopy of enamel from Tmem2 - CKO mice revealed an irregular enamel prism structure, while the microhardness and density of enamel were significantly reduced, indicating impaired ameloblast differentiation and enamel matrix mineralization. Histological evaluation indicated weak adhesion between cells and the basement membrane in Tmem2 - CKO mice. The reduced and irregular expressions of vinculin and integrin β1 suggest that Tmem2 deficiency attenuated focal adhesion formation. In addition, abnormal HA accumulation in the ameloblast layer and weak claudin 1 immunoreactivity in Tmem2 - CKO mice indicate impaired tight junction gate function. Irregular actin filament assembly was also observed at the apical and basal ends of secretory ameloblasts. Last, we demonstrated that Tmem2 -deficient mHAT9d mouse ameloblasts exhibit defective adhesion to HA-containing substrates in vitro. Collectively, our data highlight the importance of TMEM2 in adhesion to HA-rich extracellular matrix, cell-to-cell adhesion, ameloblast differentiation, and enamel matrix mineralization. [ABSTRACT FROM AUTHOR]

The aim of this study was to compare the accuracy of the Willems method, Nolla method, Cameriere dental age estimation method(CDM), Greulich-Pyle Atlas(GPA) method, and Cameriere bone age estimation method(CBM) for age estimation in healthy children aged between ten and 14.9 years, according to the pubertal growth stage(PGS). The accuracy was determined as the difference between estimated age(EA) and chronological age(CA) was assessed by analysing orthopantomograms and hand-wrist radiographs. At the onset of the PGS, the Willems method showed the closest value to the CA for both sexes(girls = 0.17 ± 0.78, boys = 0.006 ± 0.96 years). At the peak, the Willems method showed the closest value to the CA for girls(0.005 ± 1.09 years), and the GPA method for boys(0.05 ± 0.68 years). At the end of the PGS, the Willems method showed the closest value to the CA for girls(0.09 ± 1.23 years), and the CBM for boys(0.24 ± 0.74 years). It was observed that all methods concluded with the closest result to the CA at the onset period of the PGS. Our study demonstrates that sex and PGS are effective in over- or under- estimations that are obtained according to the CA. [ABSTRACT FROM AUTHOR]

Yes associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) are ubiquitous transcriptional co-activators that control organ development, homeostasis, and tissue regeneration. Current in vivo evidence suggests that YAP/TAZ regulates enamel knot formation during murine tooth development, and is indispensable for dental progenitor cell renewal to support constant incisor growth. Being a critical sensor for cellular mechano-transduction, YAP/TAZ lays at the center of the complex molecular network that integrates mechanical cues from the dental pulp chamber and surrounding periodontal tissue into biochemical signals, dictating in vitro cell proliferation, differentiation, stemness maintenance, and migration of dental stem cells. Moreover, YAP/TAZ-mediated cell-microenvironment interactions also display essential regulatory roles during biomaterial-guided dental tissue repair and engineering in some animal models. Here, we review recent advances in YAP/TAZ functions in tooth development, dental pulp, and periodontal physiology, as well as dental tissue regeneration. We also highlight several promising strategies that harness YAP/TAZ activation for promoting dental tissue regeneration. [ABSTRACT FROM AUTHOR]

Tooth enamel is generated by ameloblasts. Any failure in amelogenesis results in defects in the enamel, a condition known as amelogenesis imperfecta. Here, we report that mice with deficient autophagy in epithelial-derived tissues (K14-Cre;Atg7 F/F and K14-Cre;Atg3 F/F conditional knockout mice) exhibit amelogenesis imperfecta. Micro–computed tomography imaging confirmed that enamel density and thickness were significantly reduced in the teeth of these mice. At the molecular level, ameloblast differentiation was compromised through ectopic accumulation and activation of NRF2, a specific substrate of autophagy. Through bioinformatic analyses, we identified Bcl11b, Dlx3, Klk4, Ltbp3, Nectin1, and Pax9 as candidate genes related to amelogenesis imperfecta and the NRF2-mediated pathway. To investigate the effects of the ectopic NRF2 pathway activation caused by the autophagy deficiency, we analyzed target gene expression and NRF2 binding to the promoter region of candidate target genes and found suppressed gene expression of Bcl11b, Dlx3, Klk4, and Nectin1 but not of Ltbp3 and Pax9. Taken together, our findings indicate that autophagy plays a crucial role in ameloblast differentiation and that its failure results in amelogenesis imperfecta through ectopic NRF2 activation. [ABSTRACT FROM AUTHOR]

Purpose: The aim of this cross-sectional study was to evaluate the impact of different body mass index status on dental caries, tooth development, and tooth eruption regarding the development of dental tissues and dental health problems. Materials amp; Methods: One-hundred-seventy-six children, aged between 3-12 years old, were included in the study and distributed into 4 groups (n:44) according to their body mass index as underweight, healthy, overweight, and obesity. Data on tooth development, tooth eruption, and dental caries were recorded by using modified Demirjian, Carvalho, and decayed-missing-filled-tooth (DMFT), respectively. According to the obtained data, p lt;0.05 was accepted as significant. Results: According to the results, DMFT was significantly higher in children with underweight than overweight and obesity whereas, whereas earlier tooth eruption was seen in children with healthy and obesity than underweight (p lt;0.05). Also, there was a significant difference between the groups in terms of DMFT (p lt;0.05). Conclusion: Dental caries was higher in children with malnourishment while tooth eruption was earlier in children with obesity. Therefore, general practitioners or pediatricians should be aware of the oral effects of obesity or malnutrition and consult a pediatric dentist.

Abstract Background Dental age (DA) estimation using two convolutional neural networks (CNNs), VGG16 and ResNet101, remains unexplored. In this study, we aimed to investigate the possibility of using artificial intelligence-based methods in an eastern Chinese population. Methods A total of 9586 orthopantomograms (OPGs) (4054 boys and 5532 girls) of the Chinese Han population aged from 6 to 20 years were collected. DAs were automatically calculated using the two CNN model strategies. Accuracy, recall, precision, and F1 score of the models were used to evaluate VGG16 and ResNet101 for age estimation. An age threshold was also employed to evaluate the two CNN models. Results The VGG16 network outperformed the ResNet101 network in terms of prediction performance. However, the model effect of VGG16 was less favorable than that in other age ranges in the 15–17 age group. The VGG16 network model prediction results for the younger age groups were acceptable. In the 6-to 8-year-old group, the accuracy of the VGG16 model can reach up to 93.63%, which was higher than the 88.73% accuracy of the ResNet101 network. The age threshold also implies that VGG16 has a smaller age-difference error. Conclusions This study demonstrated that VGG16 performed better when dealing with DA estimation via OPGs than the ResNet101 network on a wholescale. CNNs such as VGG16 hold great promise for future use in clinical practice and forensic sciences.

Background: Tooth development, as one of the major mineralized tissues in the body, require fine-tuning of mineralization micro-environment. The interaction between dental epithelium and mesenchyme plays a decisive role in this process. With epithelium–mesenchyme dissociation study, we found interesting expression pattern of insulin-like growth factor binding protein 3 (IGFBP3) in response to disruption of dental epithelium–mesenchyme interaction. Its action and related mechanisms as regulator of mineralization micro-environment during tooth development are investigated. Results: Expressions of osteogenic markers at early stage of tooth development are significantly lower than those at later stage. BMP2 treatment further confirmed a high mineralization micro-environment is disruptive at early stage, but beneficial at later stage of tooth development. In contrast, IGFBP3's expression increased gradually from E14.5, peaked at P5, and decreased afterwards, demonstrating an inverse correlation with osteogenic markers. RNA-Seq and Co-immunoprecipitation showed that IGFBP3 regulates the Wnt/beta-catenin signaling pathway activity by enhancing DKK1 expression and direct protein–protein interaction. The suppression of the mineralization microenvironment effectuated by IGFBP3 could be reversed by the DKK1 inhibitor WAY-262611, further demonstrating that IGFBP3 exerted its influence via DKK1. Conclusion: A deeper understanding of tooth development mechanisms is essential for tooth regeneration, which have great implications for dental care. The current study demonstrated that the IGFBP3 expression is regulated in accordance with the needs of the mineralization microenvironment during tooth development, and IGFBP3 exerts its modulating action on osteogenic/odontogenic differentiation of hDPSCs by DKK1-Wnt/ beta-catenin axis. [ABSTRACT FROM AUTHOR]

The Notch pathway is an ancient, evolutionary conserved intercellular signaling mechanism that is involved in cell fate specification and proper embryonic development. The Jagged2 gene, which encodes a ligand for the Notch family of receptors, is expressed from the earliest stages of odontogenesis in epithelial cells that will later generate the enamel-producing ameloblasts. Homozygous Jagged2 mutant mice exhibit abnormal tooth morphology and impaired enamel deposition. Enamel composition and structure in mammals are tightly linked to the enamel organ that represents an evolutionary unit formed by distinct dental epithelial cell types. The physical cooperativity between Notch ligands and receptors suggests that Jagged2 deletion could alter the expression profile of Notch receptors, thus modifying the whole Notch signaling cascade in cells within the enamel organ. Indeed, both Notch1 and Notch2 expression are severely disturbed in the enamel organ of Jagged2 mutant teeth. It appears that the deregulation of the Notch signaling cascade reverts the evolutionary path generating dental structures more reminiscent of the enameloid of fishes rather than of mammalian enamel. Loss of interactions between Notch and Jagged proteins may initiate the suppression of complementary dental epithelial cell fates acquired during evolution. We propose that the increased number of Notch homologues in metazoa enabled incipient sister cell types to form and maintain distinctive cell fates within organs and tissues along evolution. [ABSTRACT FROM AUTHOR]

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Dentin is a major type of hard tissue of teeth and plays essential roles for normal tooth function. Odontoblasts are responsible for dentin formation. Mutations or deficiency in various genes affect the differentiation of odontoblasts, leading to irreversible dentin developmental defects in animals and humans. Whether such dentin defects can be reversed by gene therapy for odontoblasts remains unknown. In this study, we compare the infection efficiencies of six commonly used adeno-associated virus (AAV) serotypes (AAV1, AAV5, AAV6, AAV8, AAV9, and AAVDJ) in cultured mouse odontoblast-like cells (OLCs). We show that AAV6 serotype infects OLCs with the highest efficiency among the six AAVs. Two cellular receptors, which are able to recognize AAV6, AAV receptor (AAVR), and epidermal growth factor receptor (EGFR), are strongly expressed in the odontoblast layer of mouse teeth. After local administration to mouse molars, AAV6 infects the odontoblast layer with high efficiency. Furthermore, AAV6-Mdm2 was successfully delivered to teeth and prevents the defects in odontoblast differentiation and dentin formation in Mdm2 conditional knockout mice (a mouse model of dentinogenesis imperfecta type Ⅲ). These results suggest that AAV6 can serve as a reliable and efficient vehicle for gene delivery to odontoblasts through local injection. In addition, human OLCs were also successfully infected by AAV6 with high efficiency, and both AAVR and EGFR are strongly expressed in the odontoblast layer of extracted human developing teeth. These findings suggest that AAV6-mediated gene therapy through local injection may be a promising treatment approach for hereditary dentin disorders in humans. [ABSTRACT FROM AUTHOR]

Recent years have improved our understanding of the plasticity of cell types behind inducing, building, and maintaining different types of teeth. The latest efforts were aided by progress in single-cell transcriptomics, which helped to define not only cell states with mathematical precision but also transitions between them. This includes new aspects of dental epithelial and mesenchymal stem cell niches and beyond. These recent efforts revealed continuous and fluid trajectories connecting cell states during dental development and exposed the natural plasticity of tooth-building progenitors. Such "developmental" plasticity seems to be employed for organizing stem cell niches in adult continuously growing teeth. Furthermore, transitions between mature cell types elicited by trauma might represent a replay of embryonic continuous cell states. Alternatively, they could constitute transitions that evolved de novo, not known from the developmental paradigm. In this review, we discuss and exemplify how dental cell types exhibit plasticity during dynamic processes such as development, self-renewal, repair, and dental replacement. Hypothetically, minor plasticity of cell phenotypes and greater plasticity of transitions between cell subtypes might provide a better response to lifetime challenges, such as damage or dental loss. This plasticity might be additionally harnessed by the evolutionary process during the elaboration of dental cell subtypes in different animal lineages. In turn, the diversification of cell subtypes building teeth brings a diversity of their shape, structural properties, and functions. [ABSTRACT FROM AUTHOR]

CCN proteins are matricellular proteins and are important modulators of development and function of adult organs. However, there is no literature reporting the localization of CCN proteins during postnatal tooth development and the formation of periodontium. Therefore, the aim of our study was to investigate the expression of CCN1, CCN3, CCN4, CCN5 and CCN6 in the developing postnatal teeth. Wistar rats were used at postnatal (PN) 3.5, 7, 16 and 21 days and maxillas were processed for immunohistochemistry. At PN3.5 and PN7, preameloblasts (PA), secretory ameloblasts (SA), odontoblasts (OD) and dental pulp (DP) showed moderate to strong staining for CCN1, CCN4 and CCN6 respectively. CCN5 was intensely expressed in predentin, whereas CCN5 was undetectable in PA, SA, OD and DP. At PN16 and PN21, moderate to strong reaction with CCN1, CCN4 and CCN6 was evident in OD, DP, reduced enamel epithelium (REE), osteoblasts (OB) and periodontal ligament (PDL) respectively, while CCN5 was negative to weakly expressed in REE, OD, DP, OB, PDL and osteocytes (OC). Interestingly, the expression of CCN1, CCN4 and CCN6 was initially negative at PN16 but strong at PN21 in OC. Furthermore, there was no staining for CCN3 in the tissues studied. These results demonstrated that the expression pattern of CCN1, CCN4 and CCN6 is similar and inversely correlated with that of CCN3. CCN5 exhibits a unique distribution pattern. These data indicate that CCN proteins might play regulatory roles in amelogenesis, dentinogenesis, osteogenesis and PDL homeostasis. Li et al. (2022) investigated the localization of CCN1, CCN3, CCN4, CCN5 and CCN6 in the postnatal developing teeth. The results demonstrate that the expression of CCN1, CCN4 and CCN6 is similar and inversely correlated with CCN3. CCN5 exhibits a unique expression pattern. These data indicate that CCN proteins might play regulatory roles in amelogenesis, dentinogenesis, osteogenesis and periodontal ligament (PDL) homeostasis. [ABSTRACT FROM AUTHOR]

Dentin is the major hard tissue of teeth formed by differentiated odontoblasts. How odontoblast differentiation is regulated remains enigmatic. Here, we report that the E3 ubiquitin ligase CHIP is highly expressed in undifferentiated dental mesenchymal cells and downregulated after differentiation of odontoblasts. Ectopic expression of CHIP inhibits odontoblastic differentiation of mouse dental papilla cells, whereas knockdown of endogenous CHIP has opposite effects. Chip (Stub1) knockout mice display increased formation of dentin and enhanced expression of odontoblast differentiation markers. Mechanistically, CHIP interacts with and induces K63 polyubiquitylation of the transcription factor DLX3, leading to its proteasomal degradation. Knockdown of DLX3 reverses the enhanced odontoblastic differentiation caused by knockdown of CHIP. These results suggest that CHIP inhibits odontoblast differentiation by targeting its tooth-specific substrate DLX3. Furthermore, our results indicate that CHIP competes with another E3 ubiquitin ligase, MDM2, that promotes odontoblast differentiation by monoubiquitylating DLX3. Our findings suggest that the two E3 ubiquitin ligases CHIP and MDM2 reciprocally regulate DLX3 activity by catalyzing distinct types of ubiquitylation, and reveal an important mechanism by which differentiation of odontoblasts is delicately regulated by divergent post-translational modifications. [ABSTRACT FROM AUTHOR]

Objectives: This study aimed to identify formation of tubular dentin induced by transforming growth factor‐β (TGF‐β) and bone morphogenic protein (BMP) signaling pathway in dental epithelial cells. Methods: We collected conditioned medium (CM) of rTGF‐β1/rBMP‐2‐treated HAT‐7 and treated to MDPC‐23 cells. The expression levels of odontoblast differentiation markers, KLF4, DMP1, and DSP were evaluated by real‐time PCR and Western blot analysis. To evaluate whether CM of rTGF‐β1/rBMP‐2 induces tubular dentin formation, we made a beagle dog tooth defect model. Results: Here, we show that Cpne7 is regulated by Smad4‐dependent TGF‐β1/BMP2 signaling pathway in dental epithelial cells. CM of rTGF‐β1/rBMP‐2 treated HAT‐7 or rCPNE7 raises the expression levels of KLF4, DMP1, and DSP in MDPC‐23 cells. When rTGF‐β1 or rBMP‐2 is directly treated to MDPC‐23 cells, however, expression levels of Cpne7‐regulated genes remain unchanged. In a beagle dog defect model, application of rTGF‐β1/BMP2‐treated CM resulted in tubular tertiary dentin mixed with osteodentin at cavity‐prepared sites, while rTGF‐β1 group exhibited homogenous osteodentin. Conclusions: Taken together, Smad4‐dependent TGF‐β1/BMP2 signaling regulates Cpne7 in dental epithelial cells, and CPNE7 protein secreted from pre‐ameloblasts mediates odontoblast differentiation via epithelial–mesenchymal interaction. [ABSTRACT FROM AUTHOR]

Dental and craniofacial bone development is a highly coordinated process that is tightly controlled by genetics and influenced by complex environments. The abnormal regulation of many development-related signaling molecules may lead to abnormal tooth development, severe craniofacial bone formation disorders, and developmental deformities. Transforming growth factor-β (TGF-β) is widely expressed in vivo and participates in many cellular biological processes, showing complex regulatory roles in mammalian craniofacial bone growth and tooth development. In tooth development, abnormal TGF-β signaling can lead to the failure of tooth germ formation, and its deletion mutation can directly affect odontoblast differentiation and enamel formation defects. However, the current research on TGF-β mainly focuses on the early stage of tooth development, and a comprehensive and systematic study of TGF-β-related tooth development is lacking. TGF-β signal transduction mainly controls the development of teeth and craniofacial bone by regulating the expression of development-related molecules via the classical Smad-dependent signaling pathway. In addition, the nonclassical mitogen-activated protein kinase (MAPK) pathway also participates in this process. Abnormal TGF-β signaling may cause jaw development disorders, temporomandibular joint dysplasia and inflammation, and cleft palate. Because the specific regulatory mechanism of TGF-β in craniofacial bone development has not been fully elucidated, its specific application in the treatment of related diseases is also greatly limited. This paper describes the new research progress of TGF-β in the development of teeth, jaws, temporomandibular joints and palate as well as related diseases.

Abstract Background The comparison of the two Willems dental age estimation methods (gender-specific (Willems I) and non-gender-specific (Willems II)) has not been fully investigated. Here we aimed to explore the applicability of the Willems dental age estimation in an Eastern Chinese population, which may cast light on the field of dental age estimation. Methods A total of 1211 oral panoramic radiographs (582 boys and 629 girls) of the Chinese Han population aged 11–16 years old were collected. Dental ages (DAs) were calculated using the Willems method. Statistical significance was set at a p-value

Heat shock proteins (HSPs) are a class of molecular chaperones with expression increased in response to heat or other stresses. HSPs regulate cell homeostasis by modulating the folding and maturation of intracellular proteins. Tooth development is a complex process that involves many cell activities. During tooth preparation or trauma, teeth can be damaged. The damaged teeth start their repair process by remineralizing and regenerating tissue. During tooth development and injury repair, different HSPs have different expression patterns and play a special role in odontoblast differentiation and ameloblast secretion by mediating signaling pathways or participating in protein transport. This review explores the expression patterns and potential mechanisms of HSPs, particularly HSP25, HSP60 and HSP70, in tooth development and injury repair. [ABSTRACT FROM AUTHOR]

X-linked hypohidrotic ectodermal dysplasia (XLHED), caused by a genetic deficiency of ectodysplasin A1 (EDA1), is a rare developmental disorder of ectodermal derivatives such as hair, sweat glands, and teeth. The absence of sweat glands and perspiration can evoke life-threatening hyperthermia. As molecular genetic findings are not always conclusive, the concentrations of circulating EDA1 may help to distinguish between total and partial EDA1 deficiencies. We previously treated nine male patients with obvious signs of XLHED with a recombinant EDA1 replacement protein, Fc-EDA, either shortly after birth (n = 3) or by prenatal administration in gestational week 26 and beyond (n = 6). Here, we present the long-term follow-up for up to six years. In patients who had received Fc-EDA after birth, neither sweat glands nor sweating ability were detected at the age of 12–60 months. In contrast, prenatal EDA1 replacement resulted in ample sweat gland development and pilocarpine-inducible sweating in all treated subjects, who also attained more permanent teeth than their untreated affected relatives. Normal perspiration has persisted for six years in the two oldest boys treated repeatedly with Fc-EDA in utero. When they had a sauna, adequate thermoregulation was evidenced. Lower sweat production after single prenatal dosing may indicate a dose–response relationship. The absence of circulating EDA1 in five prenatally treated subjects proved that these children would have been unable to perspire if they had been left untreated. The sixth infant was shown to produce an EDA1 molecule that, albeit interacting with its cognate receptor, cannot activate EDA1 signaling. In conclusion, a causal treatment of XLHED before birth is feasible. [ABSTRACT FROM AUTHOR]