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International Dental Journal Feb 2024The aim of this research was to investigate the functions of Piezo channels in dentin defect, including mechanical signalling and odontoblast responses.
OBJECTIVES
The aim of this research was to investigate the functions of Piezo channels in dentin defect, including mechanical signalling and odontoblast responses.
METHODS
Rat dentin-defect models were constructed, and spatiotemporal expression of Piezo proteins was detected in the pulpo-dentinal complex. Real-time polymerase chain reaction (rtPCR) was used to investigate the functional expression pattern of Piezo channels in odontoblasts. Moreover, RNA interference technology was employed to uncover the underlying mechanisms of the Piezo-driven inflammatory response and repair under fluid shear stress (FSS) conditions in vitro.
RESULTS
Piezo1 and Piezo2 were found to be widely expressed in the odontoblast layer and dental pulp in the rat dentin-defect model during the end stage of reparative dentin formation. The expression levels of the Piezo1 and Piezo2 genes in MDPC-23 cells were high in the initial stage under FSS loading and then decreased over time. Moreover, the expression trends of inflammatory, odontogenic, and mineralisation genes were generally contrary to those of Piezo1 and Piezo2 over time. After silencing of Piezo1/Piezo2, FSS stimulation resulted in significantly higher expression of inflammatory, odontogenesis, and mineralisation genes in MDPC-23 cells. Finally, the expression of genes involved in the integrin β1/ERK1 and Wnt5b/β-catenin signalling pathways was changed in response to RNA silencing of Piezo1 and Piezo2.
CONCLUSIONS
Piezo1 and Piezo2 may be involved in regulating the expression of inflammatory and odontogenic genes in odontoblasts stimulated by FSS.
Topics: Rats; Humans; Animals; Odontoblasts
PubMed: 37833209
DOI: 10.1016/j.identj.2023.07.002 -
Advanced Materials (Deerfield Beach,... May 2024It remains an obstacle to induce the regeneration of hard dentin tissue in clinical settings. To overcome this, a P(VDF-TrFE) piezoelectric film with 2 wt% SrCl addition...
It remains an obstacle to induce the regeneration of hard dentin tissue in clinical settings. To overcome this, a P(VDF-TrFE) piezoelectric film with 2 wt% SrCl addition is designed. The biofilm shows a high flexibility, a harmonious biocompatibility, and a large piezoelectric d coefficient of 14 pC N, all contributing to building an electric microenvironment that favor the recruitment of dental pulp stem cells (DPSCs) and their differentiation into odontoblasts during normal chewing, speaking, etc. On the other hand, the strontium ions can be gradually released from the film, thus promoting DPSC odonto-differentiation. In vivo experiments also demonstrate that the film induces the release of dentin minerals and regeneration of dentin tissue. In the large animal dentin defect models, this piezoelectric film induces in situ dentin tissue formation effectively over a period of three months. This study illustrates a therapeutic potential of the piezoelectric film to improve dentin tissue repair in clinical settings.
Topics: Dentin; Biofilms; Dental Pulp; Strontium; Animals; Humans; Regeneration; Stem Cells; Cell Differentiation; Odontoblasts; Biocompatible Materials
PubMed: 38335452
DOI: 10.1002/adma.202313419 -
Dental Materials : Official Publication... Jun 2024To investigate the transdentinal effects of surface reaction-type pre-reacted glass-ionomer (S-PRG) fillers on odontoblast-like cells.
OBJECTIVES
To investigate the transdentinal effects of surface reaction-type pre-reacted glass-ionomer (S-PRG) fillers on odontoblast-like cells.
METHODS
An eluate of S-PRG fillers was obtained by dissolving the particles in distilled water (1:1 m/v). Dentin discs with similar permeability were mounted into artificial pulp chambers and MDPC-23 cells were seeded on their pulpal surface. The occlusal surface was treated with (n = 10): ultrapure water (negative control - NC), hydrogen peroxide (positive control - PC), S-PRG eluate exposure for 1 min (S-PRG 1 min), or S-PRG filler eluate exposure for 30 min (S-PRG 30 min). After 24 h, cell viability (alamarBlue) and morphology (SEM) were evaluated. The extract obtained from transdentinal diffusion was applied to MDPC-23 pre-cultured in plates for another 24 h to evaluate viability (alamarBlue, 1, 3, and 7 days), gene expression of Col1a1, Alpl, Dspp, and Dmp1 (RT-qPCR, 1 and 7 days), and mineralization (Alizarin Red, 7 days). Data were analyzed with ANOVA (α = 5 %).
RESULTS
While S-PRG 1 min did not differ from NC, S-PRG 30 min reduced 17.9 % viability of cells from discs. S-PRG treatments resulted in low cell detaching from dentin, and the remaining cells exhibited typical morphology or minor cytoplasmic contraction. S-PRG 30 min slightly increased cell viability (6 %) 1 day after contact with the extract. S-PRG treatments upregulated the expression of the investigated genes, especially after 1 day. S-PRG 30 min stimulated mineralization activity by 39.7 %.
CONCLUSIONS
S-PRG filler eluate does not cause transdentinal cytotoxicity on odontoblast-like cells, and long-term exposure can stimulate their dentinogenic-related mineralization activity.
SIGNIFICANCE
The transdentinal elution of ions from S-PRG fillers is not expected to be harmful to the dental pulp and may exert bioactive effects by inducing dentin matrix deposition through the metabolism of underlying odontoblasts.
PubMed: 38871524
DOI: 10.1016/j.dental.2024.06.002 -
Cureus Oct 2023The pathophysiology of calcified dental pulp is considered to be comparable to that of calcified atheroma in the artery. These calcified masses are small nodular which... (Review)
Review
The pathophysiology of calcified dental pulp is considered to be comparable to that of calcified atheroma in the artery. These calcified masses are small nodular which is seen more often in the coronal pulp region than in the radicular pulp. This is generally more common in the elderly population and usually corresponds to increasing age. Calcifications are also found in the brain, breast, arteries, and kidneys. There is a link between pulp calcification and cardiovascular problems. It is commonly assumed that individuals suffering from cardiovascular diseases have a higher risk of calcification in the pulp. The use of radiography as a quick means of detecting cardiovascular disease is possible. The pulp calcification process is usually triggered by the osteoblastic process. The process is identified by the presence of an osteoid matrix, which is built down by odontoblast cells in the pulp's peripheral portions, culminating in the production of tissue that is comparable to dentine. This review will look at pulp calcifications from all angles, including their mechanism, clinical considerations, radiographic features, and management, and also to determine if there is a link between pulp calcification and cardiovascular problems.
PubMed: 38021616
DOI: 10.7759/cureus.47258 -
Journal of Cellular Biochemistry May 2024Odontoblast differentiation is a key process in dentin formation. Mouse dental papilla cells (mDPCs) are pivotal in dentinogenesis through their differentiation into...
Odontoblast differentiation is a key process in dentin formation. Mouse dental papilla cells (mDPCs) are pivotal in dentinogenesis through their differentiation into odontoblasts. Odontoblast differentiation is intricately controlled by transcription factors (TFs) in a spatiotemporal manner. Previous research explored the role of RUNX2 and KLF4 in odontoblast lineage commitment, respectively. Building on bioinformatics analysis of our previous ATAC-seq profiling, we hypothesized that KLF4 potentially collaborates with RUNX2 to exert its biological role. To investigate the synergistic effect of multiple TFs in odontoblastic differentiation, we first examined the spatiotemporal expression patterns of RUNX2 and KLF4 in dental papilla at the bell stage using immunostaining techniques. Notably, RUNX2 and KLF4 demonstrated colocalization in preodontoblast. Further, immunoprecipitation and proximity ligation assays verified the interaction between RUNX2 and KLF4 in vitro. Specifically, the C-terminus of RUNX2 was identified as the interacting domain with KLF4. Functional implications of this interaction were investigated using small hairpin RNA-mediated knockdown of Runx2, Klf4, or both. Western blot analysis revealed a marked decrease in DSPP expression, an odontoblast differentiation marker, particularly in the double knockdown condition. Additionally, alizarin red S staining indicated significantly reduced mineralized nodule formation in this group. Collectively, our findings highlight the synergistic interaction between RUNX2 and KLF4 in promoting odontoblast differentiation from mDPCs. This study contributes to a more comprehensive understanding of the regulatory network of TFs governing odontoblast differentiation.
PubMed: 38720665
DOI: 10.1002/jcb.30577 -
Anatomical Record (Hoboken, N.J. : 2007) May 2024PIEZO1 and PIEZO2 are essential components of mechanogated ion channels, which are required for mechanotransduction and biological processes associated with mechanical...
PIEZO1 and PIEZO2 are essential components of mechanogated ion channels, which are required for mechanotransduction and biological processes associated with mechanical stimuli. There is evidence for the presence of PIEZO1 and PIEZO2 in teeth and periodontal ligaments, especially in cell lines and mice, but human studies are almost nonexistent. Decalcified permanent human teeth and mouse molars were processed for immunohistochemical detection of PIEZO1 and PIEZO2. Confocal laser microscopy was used to examine the co-localization of PIEZO 1 and PIEZO2 with vimentin (a marker of differentiated odontoblasts) in human teeth. In the outer layer of the human dental pulp, abundant PIEZO1- and PIEZO2-positive cells were found that had no odontoblast morphology and were vimentin-negative. Based on their morphology, location, and the absence of vimentin positivity, they were identified as dental pulp stem cells or pre-odontoblasts. However, in mice, PIEZO1 and PIEZO2 were ubiquitously detected and colocalized in odontoblasts. Intense immunoreactivity of PIEZO1 and PIEZO2 has been observed in human and murine periodontal ligaments. Our findings suggest that PIEZO1 and PIEZO2 may be mechanosensors/mechanotransducers in murine odontoblasts, as well as in the transmission of forces by the periodontal ligament in humans and mice.
Topics: Humans; Mice; Animals; Periodontal Ligament; Vimentin; Mechanotransduction, Cellular; Dental Pulp; Ion Channels
PubMed: 37975162
DOI: 10.1002/ar.25351 -
Journal of Biomaterials Applications Aug 2023Pulp-Dentin regeneration is a key aspect of maintain tooth vitality and enabling good oral-systemic health. This study aimed to investigate a nanofibrous scaffold loaded...
Pulp-Dentin regeneration is a key aspect of maintain tooth vitality and enabling good oral-systemic health. This study aimed to investigate a nanofibrous scaffold loaded with a small molecule i.e. Tideglusib to promote odontogenic differentiation. Tideglusib (GSK-3β inhibitor) interaction with GSK-3β was determined using molecular docking and stabilization of β-catenin was examined by confocal microscopy. 3D nanofibrous scaffolds were fabricated through electrospinning and their physicochemical characterizations were performed. Scaffolds were seeded with mesenchymal stem cells or pre-odontoblast cells to determine the cells proliferation and odontogenic differentiation. Our results showed that Tideglusib (TG) binds with GSK-3β at Cys199 residue. Stabilization and nuclear translocation of β-catenin was increased in the odontoblast cells treated with TG. SEM analysis revealed that nanofibers exhibited controlled architectural features that effectively mimicked the natural ECM. UV-Vis spectroscopy demonstrated that TG was incorporated successfully and released in a controlled manner. Both kinds of biomimetic nanofibrous matrices (PCLF-TG100, PCLF-TG1000) significantly stimulated cells proliferation. Furthermore, these scaffolds significantly induced dentinogenic markers (ALP, and DSPP) expression and biomineralization. In contrast to current pulp capping material driving dentin repair, the sophisticated, polymeric scaffold systems with soluble and insoluble spatiotemporal cues described here can direct stem cell differentiation and dentin regeneration. Hence, bioactive small molecule-incorporated nanofibrous scaffold suggests an innovative clinical tool for dentin tissue engineering.
Topics: Tissue Scaffolds; Nanofibers; beta Catenin; Glycogen Synthase Kinase 3 beta; Molecular Docking Simulation; Cells, Cultured; Cell Differentiation; Tissue Engineering; Dental Pulp
PubMed: 37485690
DOI: 10.1177/08853282231190470 -
Journal of Molecular Histology Aug 2023FAM20C phosphorylates secretory proteins at S-x-E/pS motifs, and previous studies of Fam20C-dificient mice revealed that FAM20C played essential roles in bone and tooth...
FAM20C phosphorylates secretory proteins at S-x-E/pS motifs, and previous studies of Fam20C-dificient mice revealed that FAM20C played essential roles in bone and tooth formation. Inactivation of FAM20C in mice led to hypophosphatemia that masks direct effect of FAM20C in these tissues, and consequently the direct role of FAM20C remains unknown. Our previous study reported that osteoblast/odontoblast-specific Fam20C transgenic (Fam20C-Tg) mice had normal serum phosphate levels and that osteoblastic FAM20C-mediated phosphorylation regulated bone formation and resorption. Here, we investigated the direct role of FAM20C in dentin using Fam20C-Tg mice. The tooth of Fam20C-Tg mice contained numerous highly phosphorylated proteins, including SIBLINGs, compared to that of wild-type mice. In Fam20C-Tg mice, coronal dentin volume decreased and mineral density unchanged at early age, while the volume unchanged and the mineral density elevated at maturity. In these mice, radicular dentin volume and mineral density decreased at all ages, and histologically, the radicular dentin had wider predentin and abnormal apical-side dentin with embedded cells and argyrophilic canaliculi. Immunohistochemical analyses revealed that abnormal apical-side dentin had bone and dentin matrix properties accompanied with osteoblast-lineage cells. Further, in Fam20C-Tg mice, DSPP content which is important for dentin formation, was reduced in dentin, especially radicular dentin, which might lead to defects mainly in radicular dentin. Renal subcapsular transplantations of tooth germ revealed that newly formed radicular dentin replicated apical abnormal dentin of Fam20C-Tg mice, corroborating that FAM20C overexpression indeed caused the abnormal dentin. Our findings indicate that odontoblastic FAM20C-mediated phosphorylation in the tooth regulates dentin formation and odontoblast differentiation.
Topics: Mice; Animals; Odontoblasts; Mice, Transgenic; Tooth; Cell Differentiation; Extracellular Matrix Proteins; Dentin; Phosphoproteins; Calcium-Binding Proteins
PubMed: 37357253
DOI: 10.1007/s10735-023-10123-y -
Frontiers in Physiology 2023Mouse and human genetic studies indicate key roles of the ligand in odontogenesis. Previous studies have identified effectors and regulators of the Wnt signaling...
Mouse and human genetic studies indicate key roles of the ligand in odontogenesis. Previous studies have identified effectors and regulators of the Wnt signaling pathway actively expressed during key stages of tooth morphogenesis. However, limitations in multiplexing and spatial resolution hindered a more comprehensive analysis of these signaling molecules. Here, profiling of transcriptomes using fluorescent multiplex hybridization and single-cell RNA-sequencing (scRNA-seq) provide robust insight into the synchronized expression patterns of , , and simultaneously during tooth development. First, we identified transcripts restricted to the epithelium at the stage of tooth bud morphogenesis, contrasting that of and localization to the dental mesenchyme. By embryonic day 15.5 (E15.5), a marked shift of expression from dental epithelium to mesenchyme was noted, while and expression remained enriched in the mesenchyme. By postnatal day 0 (P0), co-localization patterns of , , and were observed in both terminally differentiating and secreting odontoblasts of molars and incisors. Interestingly, exhibited robust expression in fully differentiated ameloblasts at the developing cusp tip of both molars and incisors, an observation not previously noted in prior studies. At P7 and 14, after the mineralization of dentin and enamel, expression was limited to odontoblasts. Meanwhile, Wnt modulators showed reduced or absent signals in molars. In contrast, strong signals persisted in ameloblasts (for ) and odontoblasts (for , , and ) towards the proximal end of incisors, near the cervical loop. Our scRNA-seq analysis used CellChat to further contextualize Wnt pathway-mediated communication between cells by examining ligand-receptor interactions among different clusters. The co-localization pattern of , , and in both terminally differentiating and secreting odontoblasts of molars and incisors potentially signifies the crucial ligand-modulator interaction along the gradient of cytodifferentiation starting from each cusp tip towards the apical region. These data provide cell type-specific insight into the role of Wnt ligands and mediators during epithelial-mesenchymal interactions in odontogenesis.
PubMed: 38274045
DOI: 10.3389/fphys.2023.1316635 -
Odontology Jan 2024Perfect intercellular junctions are key for odontoblast barrier function. However, whether Partitioning defective-3 (Par3) is expressed in odontoblasts and its potential...
Perfect intercellular junctions are key for odontoblast barrier function. However, whether Partitioning defective-3 (Par3) is expressed in odontoblasts and its potential effects on odontoblast junctions are unknown. Herein, we investigated the effect of Par3 on cellular junctions and the biological behavior of odontoblast-lineage cells (OLCs). Whole-transcriptome sequencing was used to analyze the effects of Par3 on OLCs and the underlying molecular mechanism. Par3 was detected under physiological and inflammatory conditions in OLCs. To investigate the regulatory effect of Par3 on junctions between mouse OLCs, the effects of Par3 downregulation on the proliferation, migration, cycle and apoptosis of OLCs were detected by 5-ethyl-2'-deoxyuridine (EdU) and Transwell assays and flow cytometry. Western blotting and alizarin red S and alkaline phosphatase (ALP) staining were used to observe the effect of Par3 downregulation on OLC mineralization. Whole-transcriptome sequencing was used to investigate the biological role of Par3 in OLCs and potential molecular mechanisms. Par3 was located along the odontoblast layer in the rat pulp tissue and in the cytoplasm of OLCs. Par3 expression was downregulated under inflammatory conditions. The OLC junctions were discontinuous, and total Zona occluden-1 (ZO-1) expression and expression of ZO-1 at the membrane in OLCs were reduced after Par3 silencing (P < 0.05). Expression of a junction-related protein (ZO-1) was downregulated after the downregulation of Par3 (P < 0.05), and ZO-1 moved from the cell membrane to the cytoplasm. OLC proliferation and migration were enhanced, but apoptosis and mineralization were inhibited in shPar3-transfected cells (P < 0.05). Sequencing identified 2996 differentially expressed genes (DEGs), which were mainly enriched in the response to stimuli and binding. Downregulation of Par3 could overactivate the PI3k-AKT pathway by promoting AKT phosphorylation (P < 0.05). Downregulation of Par3 may disrupt junctions between OLCs by affecting ZO-1 expression and distribution and promote OLC proliferation and migration but inhibit OLC mineralization. Par3 may interact with 14-3-3 proteins for PI3K-AKT pathway activation to affect OLC junctions and function.
Topics: Mice; Rats; Animals; Odontoblasts; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Cell Line; Intercellular Junctions; Cell Differentiation
PubMed: 37493885
DOI: 10.1007/s10266-023-00838-5