Comprehensive analysis of transcription factors involved in odontoblast differentiation mechanism.

Primary cultured odontoblasts rapidly lose their tissue-specific phenotype. To identify transcription factors (TF) that are important for the maintenance of the odontoblast phenotype, primary cultures of C57BL/6 J mouse dental mesenchymal cells (DMC) were isolated, and expression of TF and odontoblast marker genes in cells immediately after isolation and 2 days after culture were comprehensively evaluated and compared using RNA-sequencing (RNA-seq). The expression of odontoblast markers in mouse dental mesenchymal cells decreased rapidly after isolation. In addition, the expression of Hedgehog-related, Notch-related, and immediate- early gene (IEG)-related transcription factors significantly decreased. Forced expression of these genes in lentiviral vectors, together with fibroblast growth factor 4 (FGF4), fibroblast growth factor 9 (FGF9), and the Wnt pathway activator CHIR99021, significantly induced the expression of odontogenic marker genes. These results indicate, for the first time, that Notch signaling and early genes may be important for maintaining odontoblast cultures. Furthermore, simultaneous stimulation of FGF, Wnt, Hedgehog, Notch pathways, and IEG transcription factors cooperatively promoted the maintenance of the odontoblast phenotype. These results suggest that the Hedgehog and Notch signaling pathways may play an important role in maintaining odontoblast phenotypes, in addition to FGF and Wnt signaling.

Current Status of Next-Generation Sequencing in Bone Genetic Diseases.

The development of next-generation sequencing (NGS) has dramatically increased the speed and volume of genetic analysis. Furthermore, the range of applications of NGS is rapidly expanding to include genome, epigenome (such as DNA methylation), metagenome, and transcriptome analyses (such as RNA sequencing and single-cell RNA sequencing). NGS enables genetic research by offering various sequencing methods as well as combinations of methods. Bone tissue is the most important unit supporting the body and is a reservoir of calcium and phosphate ions, which are important for physical activity. Many genetic diseases affect bone tissues, possibly because metabolic mechanisms in bone tissue are complex. For instance, the presence of specialized immune cells called osteoclasts in the bone tissue, which absorb bone tissue and interact with osteoblasts in complex ways to support normal vital functions. Moreover, the many cell types in bones exhibit cell-specific proteins for their respective activities. Mutations in the genes encoding these proteins cause a variety of genetic disorders. The relationship between age-related bone tissue fragility (also called frailty) and genetic factors has recently attracted attention. Herein, we discuss the use of genomic, epigenomic, transcriptomic, and metagenomic analyses in bone genetic disorders.

IL-33 deficiency suppresses alveolar bone loss in a ligature-induced periodontitis model.

Interleukin-33 (IL-33) is a member of the IL-1 cytokine family that has been studied primarily in the context of type 2 immune responses. Recent reports suggest that IL-33 also enhances the func- tions of various immune cells and contributes to the development of different inflammatory diseas- es. Interestingly, IL-33 and its receptor ST2 axis exerted either inhibitory or promotional effects on alveolar bone loss in various periodontitis models. Using a mouse model of ligature-induced periodontitis, we found that the levels of mRNAs encoding IL-33 and other inflammatory cyto- kines (IL-1α, IL-1ß, IL-6, and TNFα) were augmented in gingival tissues of wild-type (WT) mice, and that the alveolar bone loss amount was lower in IL-33-deficient than WT mice. The numbers and proportions of IFN-γ-producing CD8+ T and regulatory T cells were decreased while those of Th17 cells were increased in the draining lymph nodes of IL-33-deficient mice compared to WT mice. Additionally, the level of RNA encoding an osteoclastogenic molecule, i.e., receptor activa- tor of nuclear factor kappa-B ligand (RANKL), in ligated gingival tissue was higher in IL-33-defi- cient than WT mice. These results suggest that IL-33 is involved in alveolar bone loss in the ligature-induced periodontitis model, although IL-33 may inhibit osteoclast differentiation.

Bond Strength of Two Resin Cements with Leucite-reinforced Ceramic Using Different Bonding Agents.

AIM: To compare the bond strength of two resin cements to leucite-reinforced ceramics using three different boding agents and evaluate the compatibility of bonding agents. MATERIALS AND METHODS: Twenty extracted sound human molars were sectioned horizontally 2-3 mm above the cementoenamel junction (CEJ). CAD/CAM ceramic blocks for inLab were also sectioned to create 4 mm thick and bonded to the dentin. The adhesive groups assigned were divided into four adhesive groups: Group I: Variolink II dual-cure resin cement and Scotchbond Multi-Purpose Plus adhesive, group II: Multilink Automix dual-cure resin cement and Multilink primers, group III: Multilink Automix and Clearfil SE bond 2 (CSE2) adhesive, group IV: Multilink Automix and CSE2 with light curing after adhesive application. Five specimens of each group were sectioned perpendicular to obtain six microsticks of 1 × 1 mm width from each sample. Microtensile bond strength data were expressed in MPa. Fracture modes (FrMs) analyzed for the surfaces were divided into six patterns. Microtensile bond strength data were statistically analyzed with one-way ANOVA and Tukey post hoc tests (α = 0.05). T-test was performed at the 5% significance level to analyze groups III and IV with and without light curing. RESULTS: Group I showed the highest µTBS average of 13.67 MPa, group IV showed 12.26 MPa, group III showed 12.15 MPa, and group II showed the lowest average of 10.84 MPa. No significant differences were found between the bonding agents. However, the six types of failure modes, although all observed, were characterized by the adhesive system: Type I: adhesive failure of laminated dentin and ceramic; type II: adhesive failure of laminated ceramic; type III: adhesive failure of laminated dentin; type IV: cohesive failure of luting agent; type V: cohesive failure of dentin, and type VI: mixed failure of adhesion and cohesion. As a result, the FrM most commonly observed was the adhesive failure at the luting cement-ceramic block interface. CONCLUSION: The combination of resin cements and bonding agents did not significantly affect the bond strength of CAD/CAM ceramic restorations and dentin. CLINICAL SIGNIFICANCE: Several universal bonding agents are currently available for direct and indirect bonding, and using the same bonding agent for direct and indirect restorations could simplify inventory and benefit routine clinical practice. How to cite this article: Aida N, Koi K, Amaya-Pajares SP, et al. Bond Strength of Two Resin Cements with Leucite-reinforced Ceramic Using Different Bonding Agents. J Contemp Dent Pract 2023;24(11):859-863.

Progress and Current Status in Hajdu-Cheney Syndrome with Focus on Novel Genetic Research.

Hajdu-Cheney syndrome (HCS) is a rare autosomal dominant manifestation of a congenital genetic disorder caused by a mutation in the NOTCH2 gene. NOTCH signaling has variations from NOTCH 1 to 4 and maintains homeostasis by determining and regulating the proliferation and differentiation of various cells. In HCS, the over-accumulated NOTCH2 causes abnormal bone resorption due to its continuous excessive signaling. HCS is characterized by progressive bone destruction, has complex wide-range clinical manifestations, and significantly impacts the patient's quality of life. However, no effective treatment has been established for HCS to date. There are genetic variants of NOTCH2 that have been reported in the ClinVar database of the U.S. National Institutes of Health. In total, 26 mutant variants were detected based on the American College of Medical Genetics and Genomics (ACMC). To date, there has been no comprehensive compilation of HCS mutations. In this review, we provide the most comprehensive list possible of HCS variants, nucleotide changes, amino acid definitions, and molecular consequences reported to date, following the ACMC guidelines.

The effect of BMP4, FGF8 and WNT3a on mouse iPS cells differentiating to odontoblast-like cells.

We investigated whether BMP4, FGF8, and/or WNT3a on neural crest-like cells (NCLC) derived from mouse induced pluripotent stem (miPS) cells will promote differentiation of odontoblasts-like cells. After the miPS cells matured into embryonic body (EB) cells, they were cultured in a neural induction medium to produce NCLC. As the differentiation of NCLC were confirmed by RT-qPCR, they were then disassociated and cultured with a medium containing, BMP4, FGF8, and/or WNT3a for 7 and 14 days. The effect of these stimuli on NCLC were assessed by RT-qPCR, ALP staining, and immunocytochemistry. The cultured EB cells presented a significant increase of Snai1, Slug, and Sox 10 substantiating the differentiation of NCLC. NCLC stimulated with more than two stimuli significantly increased the odontoblast markers Dmp-1, Dspp, Nestin, Alp, and Runx2 expression compared to control with no stimulus. The expression of Dmp-1 and Dspp upregulated more when FGF8 was combined with WNT3a. ALP staining was positive in groups containing BMP4 and fluorescence was observed in immunocytochemistry of the common significant groups between Dmp-1 and Dspp. After stimulation, the cell morphology demonstrated a spindle-shaped cells with long projections resembling odontoblasts. Simultaneous BMP4, FGF8, and WNT3a stimuli significantly differentiated NCLC into odontoblast-like cells.

Successful development and implementation of a digital dentistry curriculum at a US dental school.

Introducing new technology in the dental school curriculum may face some uphill challenges navigating through university-level bureaucracy, financial ramifications, and influencing the mindset of certain faculty. A dental school that relies heavily on teaching traditional restorative procedures may particularly face many challenges to embrace new technology. The restorative department at a US dental school created a division of computer-aided design and computer-aided manufacturing (CAD/CAM) dentistry to introduce the concept of digital dentistry to dental students and faculty. This article describes methods that were adopted to improve acceptance and increase the productivity of chairside CAD/CAM restorations for both students and faculty at the dental school in US.

Intentional Replantation to Treat Apical Periodontitis of Maxillary First Molar with Foreign Body Located Outside Apical Foramen Using CBCT: A Case Report.

The recent use of cone beam computed tomography (CBCT) in the field of dentistry to obtain 3-dimensional (3D) images has enabled more effective examination and diagnosis in endodontic treatment. Such information has also been reported to be of benefit in surgical endodontic treatment such as intentional replantation. Here, we report a case of intentional replantation with the assistance of CBCT in which a good therapeutic outcome was achieved. The patient was a 30-year-old woman who visited our hospital with the chief complaint of spontaneous pain in the right maxillary first molar. Dental radiographs revealed a radiolucent area in the apical portion of the distal root, and a radiopaque area thought to be a broken shard from a small surgical instrument measuring approximately 1.5 mm in length. The shard was located outside the distal apical foramen. Based on these findings, the diagnosis was acute suppurative apical periodontitis of the right maxillary first molar. Infected root canal therapy was subsequently commenced. The patient's symptoms showed no improvement, however, and the pain persisted. Therefore, dental CBCT was performed to obtain 3D images, which confirmed a radiopaque area thought to be a broken shard from a small surgical instrument located outside the apical foramen of the distal root and facing in a direction that made it impossible to remove from the root canal. Because the patient's symptoms had shown no improvement and a foreign body was observed outside the apical foramen, intentional replantation combined with root resection was performed with informed consent. At a 1-year follow-up visit, progress was good, and there were no reported symptoms or signs. When reaching a diagnosis is difficult based on clinical findings and dental radiographs alone, the 3D images provided by CBCT offer a means of securing a more reliable diagnosis, allowing planning of treatment to be more effective.

Microscopic Endodontics in Infected Root Canal with Calcified Structure: A Case Report.

Calcium deposited within a root canal due to exogenous stimuli may hamper root canal treatment. In endodontic treatment, an operating microscope allows the conditions within the root canal to be directly viewed and evaluated. This report describes a case in which an operating microscope was used to facilitate the excision of a calcified structure from within a root canal at an early stage in the treatment of an infection. An 18-year-old man was referred to our clinic due to suspected chronic suppurative apical periodontitis of the right maxillary central incisor. Periapical radiography confirmed the presence of a radioopaque structure inside the root canal that was likely to pose an obstacle to endodontic treatment. After opening the pulp chamber, an operating microscope was used to directly confirm the presence of the calcified structure in the root canal, which was removed using an ultrasonic tip. The infected root canal was treated using calcium hydroxide. Two months later, closure of the apical foramen as a result of calcification of the apical foramen was confirmed and the root canal filled. Using an operating microscope to directly view a structure posing an obstacle to root canal treatment made it possible to perform an excision while avoiding risks such as canal perforation.

An endodontic-periodontal lesion with primary periodontal disease: a case report on its bacterial profile.

The microflora in the periodontal pockets can affect the dental pulp and cause endodontic-periodontal lesions or retrograde pulpitis. Here we report an endodontic-periodontal lesion together with its bacterial profile. The lesion occurred in the maxillary right first molar of a 40-year-old woman who presented at our hospital complaining of a violent toothache since the previous night. Clinically, the tooth was caries-free and an electric pulp test showed it to be vital. The tooth showed signs of advanced periodontitis and the periodontal pocket was deep, reaching the apex of the palatal root. The clinical diagnosis was an endodontic-periodontal lesion with primary periodontal disease. Subsequent endodontic treatment comprised pulp extirpation and root canal filling, followed by periodontal treatment consisting of scaling and root planing. The tooth was finally restored with a full metal crown. No further signs of periodontal disease or periapical lesions have been observed to date. Bacteria were sampled from the root canal and periodontal pocket for a microbiological assessment using 16S rRNA gene-based PCR. Microbiologically, the profile of the bacterial species from the palatal root canal was similar to that from the periodontal pocket of the palatal root. Porphyromonas gingivalis, Fusobacterium nucleatum and Eikenella corrodens were detected in both samples. The occurrence of bacteria common to both sites in this patient further supports the proposition that periodontal disease is the definitive source of root canal infections. The present results suggest that a bacterial examination would be helpful in confirming and supporting the clinical diagnosis in such lesions.

Digital video image processing from dental operating microscope in endodontic treatment.

Recently, optical microscopes have been used in endodontic treatment, as they offer advantages in terms of magnification, illumination, and documentation. Documentation is particularly important in presenting images to patients, and can take the form of both still images and motion video. Although high-quality still images can be obtained using a 35-mm film or CCD camera, the quality of still images produced by a video camera is significantly lower. The purpose of this study was to determine the potential of RegiStax in obtaining high-quality still images from a continuous video stream from an optical microscope. Video was captured continuously and sections with the highest luminosity chosen for frame alignment and stacking using the RegiStax program. The resulting stacked images were subjected to wavelet transformation. The results indicate that high-quality images with a large depth of field could be obtained using this method.

Dens evaginatus on occlusal surface of maxillary second molar: a case report.

Dens evaginatus is a rare dental anomaly characterized by the development of a tubercle on the occlusal surface of the tooth and can cause pulpitis, pulp necrosis, and periapical periodontitis due to tubercular fracture or attrition. Unlike with caries, pain caused by dens evaginatus may manifest itself in a distant location. Therefore, diagnosing the cause of that pain may prove problematic. Dens evaginatus usually occurs in the mandibular premolars. We report a successfully treated case in which dens evaginatus was difficult to diagnose due to distant radiation of pulpitis-induced pain. This pain occurred as a result of fracture of a tubercle located on the occlusal surface of the maxillary second molar, which is very rare.

Immunohistochemical characterization of elastic system fibers in rat molar periodontal ligament.

Among elastic system fibers, oxytalan fibers are known as a ubiquitous component of the periodontal ligament, but the localization and role of elastin-containing fibers, i.e., elastic and elaunin fibers, has yet to be clarified. In this study, we immunohistochemically investigated the localization of elastin and fibrillin, major proteins of elastin-containing fibers in the periodontal ligament of rat lower first molars. At the light microscope level, distribution of elastin-positive fibers was not uniform but often concentrated in the vicinity of blood vessels in the apical region of the ligament. In contrast, fibrillin-positive fibers were more widely distributed throughout the ligament, and the pattern of their distribution was comparable to the reported distribution of oxytalan fibers. At the ultrastructural level, assemblies or bundles of abundant fibrillin-containing microfibrils were intermingled with a small amount of elastin. This observation indicated that elastin-positive fibers observed under the light microscope were elaunin fibers. No mature elastic fibers, however, were found in the ligament. These results show that the major components of elastic system fibers in the periodontal ligament of the rat mandibular first molar were oxytalan and elaunin fibers, suggesting that the elastic system fibers play a role in the mechanical protection of the vascular system.