Journal of dental research最新文献

Extracellular DNA (eDNA) is one of the core components of the extracellular matrix (ECM) in biofilms and provides attachment sites for microbes and other ECM components. However, little is known about the functions and underlying mechanisms of eDNA in the cariogenicity of dental plaque biofilms. A recent study demonstrated that conformational diversity of eDNA exists in biofilms, and the transition of eDNA from right-handed (B-DNA) to left-handed (Z-DNA) is associated with the structural stability and pathogenicity of biofilms. Caries-related biofilm is a complex multispecies microenvironment. The presence and biological function of the conformational transition of eDNA within this biofilm have not been previously reported. In this study, we found that extracellular Z-DNA is widely present in carious tissues and cariogenic biofilm, especially Streptococcus mutans, indicating its possible role in the occurrence and activity of dental caries. The content of extracellular Z-DNA showed species heterogeneity. The modulation of Z-DNA formation affected the level of extracellular polysaccharide. Increased formation of Z-DNA substantially strengthened the cariogenicity of the biofilm by increasing DNase resistance, structural density, and acid production. These insights provide a new perspective to understand the underlying function of the conformation transition of eDNA in promoting carious lesions, as well as a possible anti-biofilm strategy targeting extracellular Z-DNA.

Periodontitis has recently been recognized as an inflammatory disease caused by oxidative stress, with mitochondrial dysfunction being a key factor leading to oxidative stress. PTEN-induced kinase 1 (PINK1) is an essential protein for mitochondrial quality control, which protects cells from oxidative stress by inducing mitophagy to degrade damaged mitochondria, but its role in periodontitis has not been elucidated. This study aimed to explore the contribution and underlying mechanisms of Pink1 in regulating the differentiation and function of osteoclasts during periodontitis. Here we observed a significant downregulation of PINK1 expression in periodontitis-affected tissues. Then we constructed a periodontitis model in mice with fluorescently labeled mononuclear/macrophages, and the results showed that as the modeling time extended, the alveolar bone destruction gradually worsened and was accompanied by gradually decreased Pink1 expression in osteoclasts and a significantly increased osteoclast number. In vitro experiments further demonstrated a negative correlation between Pink1 and osteoclast differentiation. In addition, alveolar bone destruction in the Pink1 knockout mice was significantly more advanced than that in the littermate wild type mice after ligature-induced periodontitis and enhanced osteoclastogenesis and bone-resorptive capacity in vitro. RNA-sequencing analysis and in vitro validation revealed that the absence of Pink1 led to a decrease in oxidative phosphorylation levels and an enhancement of calcium-mediated signaling, specifically the calcineurin-NFATc1 pathway, via an intracellular calcium source. Further mechanistic studies found that the deficiency of Pink1 inhibited mitophagy but strengthened mitochondrial-endoplasmic reticulum coupling, which, by promoting the interaction of Mfn2-IP3R-VDAC1 proteins, increased the concentration of mitochondrial calcium ions, thereby triggering more active osteoclast differentiation. The aforementioned process can be reversed by the IP3R channel inhibitor Bcl-XL. These findings unveiled that Pink1 was involved in osteoclast differentiation by regulating mitochondrial calcium transport mediated by mitochondria-associated endoplasmic reticulum membranes, providing a new theoretical basis for the pathogenesis and treatment of periodontitis.

Periodontal and peri-implant diseases are a significant public health problem worldwide, resulting in the destruction of the supporting bone. These bone defects can cause esthetic problems, increased relapse rate, and eventually tooth loss. The etiology of periodontal disease involves an influx of innate immune cells (neutrophils and monocytes) and upregulation of local inflammatory cytokines in the gingiva. Biodegradable polymeric nanoparticles are an inexpensive, safe, and effective means of preventing innate immune activation by bacterial biofilms. We therefore hypothesize that this technology is a potential means of managing periodontal disease. Polylactic acid (PLA) particles were fabricated using an oil-in-water emulsion and used as a therapy in ligature-induced periodontitis and peri-implantitis. Mice were treated daily with nanoparticles or saline control through intravenous injection for 5 or 7 d. Bone loss and quality were characterized using micro-computed tomography and histology, and immune cell infiltrate was characterized by flow cytometry and enzyme-linked immunosorbent assay. PLA particle therapy prevented bone loss in both periodontitis and peri-implantitis. Particle treatment was associated with decreased osteoclast activation. Flow cytometry showed particles were mainly taken up by macrophages and limited inflammatory monocyte recruitment to the ligature site. In vitro evaluation of particle therapy demonstrated the inhibition of toll-like receptor activation during particle treatment. These results extended to monocytes that had been presensitized by titania nanoparticles. Taken together, the results of these experiments demonstrated that cargo-less PLA particle therapy may be a safe, cost-effective therapy to manage inflammatory bone loss in periodontal disease.

Multiple sensory afferents, including mechanosensitive and nociceptive nerves, are projected to the periodontium. Peptidergic afferents expressing transient receptor potential vanilloid 1 (TRPV1), a receptor for capsaicin, mediate pain caused by orthodontic forces. However, their role in orthodontic force-induced alveolar bone remodeling is poorly understood as is the contribution of mechanosensitive ion channels such as Piezo2 in nociceptive nerves. To investigate this role, we studied orthodontic tooth movement and alveolar bone remodeling using neural manipulations and genetic mouse models. Chemical ablation of TRPV1-expressing afferents localized to the trigeminal ganglia decreased orthodontic force-induced tooth movement and the number of osteoclasts in alveolar bone on the compression side. The extent of the force-induced increase in the ratio of receptor activator of nuclear factor kappa-B ligand/osteoprotegerin in the periodontium was modestly decreased in the chemical ablation group. Furthermore, chemogenetic silencing of TRPV1-lineage afferents reduced orthodontic tooth movement and the number of osteoclasts. Piezo2 was expressed in most periodontal afferents, and chemogenetic inhibition of Piezo2-expressing neurons decreased orthodontic tooth movement and the number of osteoclasts. In addition, the conditional knockout of Piezo2 in TRPV1-lineage afferents decreased orthodontic tooth movement and the number of osteoclasts. Overall, these results suggest that nociceptor neurons play critical roles in orthodontic force-induced alveolar bone remodeling and that the mechanical activation of neuronal Piezo2 in nociceptive nerves facilitates orthodontic tooth movement and associated alveolar bone remodeling.

Amelogenesis, the process of enamel formation, is tightly regulated and essential for producing the tooth enamel that protects teeth from decay and wear. Disruptions in amelogenesis can result in amelogenesis imperfecta, a group of genetic conditions characterized by defective enamel, including enamel hypoplasia, marked by thin or underdeveloped enamel. Mutations in the KMT2D (MLL4) gene, which encodes histone H3 lysine 4 methyltransferase, are associated with Kabuki syndrome, a developmental disorder that can involve dental anomalies such as enamel hypoplasia. However, the specific role of KMT2D in amelogenesis remains poorly understood. To address this gap, we generated a conditional knockout (cKO) mouse model with ectoderm-specific deletion of Kmt2d (Krt14-Cre;Kmt2d^fl/fl, or Kmt2d-cKO) and characterized the resulting enamel defects using gross, radiographic, histologic, cellular, and molecular analyses. Micro-computed tomography and scanning electron microscopy revealed that adult Kmt2d-cKO mice exhibited 100% penetrant amelogenesis imperfecta, characterized by hypoplastic and hypomineralized enamel, partially phenocopying human Kabuki syndrome. Additionally, Kmt2d-cKO neonates developed molar tooth germs with subtle cusp shape alterations and mild delays in ameloblast differentiation at birth. RNA sequencing analysis of the first molar tooth germ at birth revealed that 33.7% of known amelogenesis-related genes were significantly downregulated in the Kmt2d-cKO teeth. Integration with KMT2D CUT&RUN sequencing results identified 8 overlapping genes directly targeted by KMT2D. Reanalysis of a single-cell RNA sequencing data set in the developing mouse incisors revealed distinct roles for these genes in KMT2D-regulated differentiation across various cell subtypes within the dental epithelium. Among these genes, Satb1 and Sp6 are likely direct targets involved in the differentiation of preameloblasts into ameloblasts. Taken together, we propose that KMT2D plays a crucial role in amelogenesis by directly activating key genes involved in ameloblast differentiation, offering insights into the molecular basis of enamel development and related dental pathologies.

Dental caries, as one of the prevalent oral infectious diseases worldwide, constitutes a considerable disease burden. Fluoride has been widely used to prevent dental caries for decades. However, fluoride alone may not always be sufficient. The major cariogenic bacterial species, Streptococcus mutans, has not been effectively controlled by daily fluoride exposure, possibly because it has a detoxification mechanism. Studies have shown that most microorganisms have fluoride exporters dedicated to exporting fluoride ions (F^-). S. mutans possesses 2 homologous genes, eriC^F1 and eriC^F2, which encode fluoride exporters, but their function has not been fully clarified. In this work, we constructed the markerless gene deletion mutants, overexpression, and complemented strains of S. mutans UA159. Assessing fluoride sensitivity, intracellular F^- levels, and cell membrane permeability revealed that EriC^F1 was the major functional unit of the fluoride exporter in S. mutans. To further enhance the antibacterial efficiency of fluoride, we identified 3 diphenylurea derivatives that might target EriC^F1 by molecular docking, which significantly enhanced the antibacterial effect of sodium fluoride (NaF) by synergistically impeding fluoride efflux, as demonstrated by chequerboard broth microdilution assays. Moreover, these compounds combined with 1 mM NaF impaired the cariogenicity of S. mutans significantly in vivo and with good biocompatibility, especially compounds 9 and 15. Collectively, these findings suggest that fluoride exporters in S. mutans could serve as a potential target for caries prevention, and the diphenylurea derivatives identified for targeting EriC^F1 could be a valuable therapeutic approach when combined with fluoride, providing promising measures for dental caries prevention.

The stability of messenger RNA (mRNA) is controlled by proteins that bind to adenosine-uridine-rich sequences (AREs) in their 3' untranslated regions (3'UTR), known as AU-binding proteins. One of these proteins is tristetraprolin (TTP; encoded by Zfp36), which promotes degradation of mRNAs with AREs in their 3'UTR. TTP accelerates the decay of its target transcripts, many of which encode proinflammatory mediators that promote tumorigenesis. TTP underexpression has been reported in multiple cancer types. Oral squamous cell carcinoma is an aggressive disease characterized by high morbidity and few therapeutic options. The role of TTP has not been studied in oral epithelium homeostasis nor in its carcinogenesis. Herein, using tissue-specific TTP knockout mice (TTP-KO), we show that TTP expression is relevant for oral epithelium homeostasis. TTP-KO mice developed dysplastic lesions in the tongue along with inflammatory infiltrates in the connective tissue. Analysis of the inflammatory infiltrate revealed the presence of mast cells (MCs), CD45+ cells, and CD11b+ cells, with the MCs being the most abundant cell type and associated with cyclooxygenase-2 expression. Recruitment of MCs was dependent on tumor necrosis factor-α (TNFα) upon TTP ablation in the tongue. Although the infiltration of MCs was dependent on TNFα activity, this did not affect the development of tongue dysplasia. We analyzed the status of the NF-κB pathway, finding its activation. In addition, we demonstrate that K-ras activation combined with Zfp36 deletion leads to the rapid onset of the oral tongue phenotype and significantly reduces mouse survival. Our results support the notion that TTP expression protects against oral carcinogenesis, regulates the inflammatory infiltrate, and maintains the epithelial microenvironment, potentially serving as a barrier to tumorigenesis.

Odontoblasts are terminally differentiated cells that exhibit mechanosensitivity and mineralization capacity. Mechanosensitive ion channels such as Piezo1 are present in odontoblasts and are associated with their physiological functions via Ca²⁺ signaling. Both Ca²⁺ signals via Ca²⁺ influx from mechanosensitive ion channels and Ca²⁺ release from Ca²⁺ stores function as secondary messenger systems for various biological phenomena. The endoplasmic reticulum (ER) serves as an intracellular Ca²⁺ store that mobilizes intracellular Ca²⁺. Changes in Ca²⁺ concentration inside the ER are among the factors that cause ER stress. Perivascular cells are located around odontoblasts in the dental pulp. Although such formation indicates that perivascular cells interact with odontoblasts, their detailed profiles under developmental and pathological conditions remain unclear. In this study, we revealed that pericyte marker, neural/glial antigen 2 (NG2)-positive cells, in cell-rich zones (CZs) can differentiate into Piezo1-positive odontoblasts following genetic odontoblast depletion in mice, and modeled as odontoblast death after severe dentin injury and as reparative dentin formation. NG2-positive pericytes differentiated into odontoblasts faster than glial cells. To determine how NG2-positive cells differentiate into Piezo1-positive odontoblasts, we focused on the ER-stress sensor protein, activating transcription factor 6a (ATF6a). After genetic odontoblast depletion, NG2-positive cells regenerated in the odontoblast layer and were capable of acting as functional odontoblasts. In the presence of extracellular Ca²⁺, the application of a sarco/ER Ca²⁺-ATPase (SERCA) inhibitor, thapsigargin, known as an ER-stress inducer, increased the intracellular Ca²⁺ concentration in the odontoblast lineage cells (OLCs). The increase was significantly inhibited by the application of a pharmacologic Piezo1 inhibitor, indicating that ER stress by SERCA inhibition augmented Piezo1-induced responses in odontoblast progenitor cells. However, the physiological activation of G_q-coupled receptors by adenosine diphosphate did not induce Piezo1 activation. Gene silencing of ATF6a and/or NG2 impaired the mineralization of OLCs. Overall, ATF6a orchestrates the differentiation of NG2-positive pericytes into functional odontoblasts that act as sensory receptor cells and dentin-forming cells.

It has been questioned whether Streptococcus mutans can still be considered the major etiological agent for caries. The main argument is that most evidence has been based on single-species identification. The composition of the oral microbiome was not analyzed. This systemic review aims to assess the prevalence and abundance of S. mutans in caries-active (CA) and caries-free (CF) subjects based on clinical studies in which the microbiome was investigated. Three databases (PubMed, Cochrane, Embase) were searched until May 22, 2023, for eligible publications that included CA and CF subjects and reported the detection of both S. mutans and the oral microbial community, using DNA-based methods. The clinical and microbial outcomes were summarized and further analyzed using a random-effects model. Of 22 eligible studies, 3 were excluded due to the high risk of bias. In the remaining 19 studies, 16 reported the prevalence of S. mutans, 11 reported its relative abundance, and 8 reported both parameters. The prevalence of S. mutans in CA was either similar to (n = 4) or higher than (n = 12) the CF group. The reported relative abundance in CA was higher than CF in all 11 studies, although the values varied from 0.001% to 5%. Meta-analysis confirmed the significance of these findings. The summary of microbial community data did not reveal other caries-associated bacterial genera/species than S. mutans. In conclusion, the collected evidence based on microbiome studies suggests a strong association between the prevalence and abundance of S. mutans and caries experience. While the cariogenic role of S. mutans in the oral ecosystem should be recognized, its actual function warrants further exploration.