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Journal of Endodontics Nov 2014Dental papilla cells (DPCs) are precursors of odontoblasts and have the potential to differentiate into odontoblasts. Osteoblasts and odontoblasts have many common...
INTRODUCTION
Dental papilla cells (DPCs) are precursors of odontoblasts and have the potential to differentiate into odontoblasts. Osteoblasts and odontoblasts have many common characteristics. Osterix (Osx) is essential for osteoblast differentiation. However, no information is available for the effects of Osx on the odontoblastic differentiation of DPCs. The purpose of this study was to investigate the effects of Osx on the proliferation and odontoblastic differentiation of DPCs.
METHODS
An immortalized human dental papilla cell (hDPC) line was used. Osx was stably overexpressed or knocked down in hDPCs with infection of lentiviral particles to determine its biological effects on hDPCs. The proliferation of cells was measured by the 5-ethynyl-2'-deoxyuridine incorporation assay and direct cell counting. Expressions of dentin sialophosphoprotein, nestin, dentin matrix protein 1, and alkaline phosphatase were detected by real-time polymerase chain reaction to determine the odontoblastic differentiation of cells. The mineralization ability of cells was evaluated by von Kossa staining and alkaline phosphatase activity assay.
RESULTS
Overexpression of Osx retarded the proliferation of hDPCs, whereas knockdown of Osx increased the cell proliferation. Overexpression of Osx promoted the odontoblastic differentiation of hDPCs by up-regulating odontoblastic differentiation genes and increased the mineralization ability of hDPCs. Knockdown of Osx down-regulated odontoblastic differentiation genes and decreased the mineralization ability of hDPCs.
CONCLUSIONS
Osx might function as a potential regulator for the proliferation and odontoblastic differentiation of hDPCs.
Topics: Alkaline Phosphatase; Antimetabolites; Calcification, Physiologic; Cell Count; Cell Culture Techniques; Cell Differentiation; Cell Proliferation; Dental Papilla; Deoxyuridine; Extracellular Matrix Proteins; Gene Expression Regulation; Gene Knockdown Techniques; Genetic Vectors; HEK293 Cells; Humans; Lentivirus; Nestin; Odontoblasts; Phosphoproteins; Sialoglycoproteins; Sp7 Transcription Factor; Transcription Factors
PubMed: 25258338
DOI: 10.1016/j.joen.2014.04.012 -
International Endodontic Journal Dec 2023Fat mass and obesity-associated (FTO) protein, the first discovered N6-methyladenine (m6A) demethylase, played positive roles in bone formation. In this study, the aim...
The N6-methyladenosine demethylase FTO is required for odontoblast differentiation in vitro and dentine formation in mice by promoting RUNX2 exon 5 inclusion through RBM4.
AIM
Fat mass and obesity-associated (FTO) protein, the first discovered N6-methyladenine (m6A) demethylase, played positive roles in bone formation. In this study, the aim was to investigate the function and potential mechanism of Fto in dentine formation.
METHODOLOGY
In vivo model, postnatal 12-day (PN12), 4-week-old (4 wk), 6-week-old (6 wk) healthy male C57BL/6J were randomly divided into Fto knockout (Fto ) mice and wild-type (WT) littermates according to their genotypes, with 3-5 mice in each group. The mandibles of Fto mice and WT control littermates were isolated for analysis by micro-computed tomography (micro-CT), 3-dimensional reconstruction and Haematoxylin-eosin (HE) staining. In vitro, mouse dental papilla cells (mDPCs) and human dental stem pulp cells (hDPSCs) were cultured with odontogenetic medium to evaluate differentiation capacity; expression levels of odontoblastic related genes were evaluated using quantitative real-time polymerase chain reaction (qRT-PCR). The inclusion levels of Runt-related transcription factor 2 (RUNX2) exon 5 in mDPCs and hDPSCs were detected by semiquantitative real-time polymerase chain reaction (RT-PCR). The RNA binding motif protein 4 (RBM4) m6A site was verified through m6A methylated RNA immunoprecipitation (MeRIP) and the stability of RBM4 mRNA influenced by FTO knockdown was measured by mRNA stability assay. Differences with p values < .05 were regarded as statistically significant.
RESULTS
We discovered that Fto mice showed significant dentine formation defects characterized by widened pulp cavity, enlarged pulp-tooth volume ratio, thinned dentine and pre-dentine layer of root (p < .05). Fto mDPCs and FTO-silencing hDPSCs not only exhibited insufficient mineralization ability and decreased expression levels of odontoblastic mineralization related genes (p < .05), but showed significantly reduced Runx2 exon 5 inclusion level (p < .05). FTO knockdown increased the m6A level of RBM4 and destabilized the mRNA of RBM4, thus contributing to the reduced RBM4 expression level. Moreover, Rbm4 overexpression in Fto mDPCs can partly restore Runx2 exon 5 inclusion level and the differentiation ability disrupted by Fto knockout.
CONCLUSION
Thus, within the limitations of this study, the data suggest that FTO promotes odontoblastic differentiation during dentine formation by stabilizing RBM4 mRNA to promote RUNX2 exon 5 inclusion.
Topics: Animals; Humans; Male; Mice; Alpha-Ketoglutarate-Dependent Dioxygenase FTO; Cell Differentiation; Core Binding Factor Alpha 1 Subunit; Dental Pulp; Dentin; Exons; Mice, Inbred C57BL; Odontoblasts; RNA, Messenger; RNA-Binding Proteins; X-Ray Microtomography
PubMed: 37698901
DOI: 10.1111/iej.13975 -
Journal of Endodontics Jul 2015During tooth development, cells originating from the neural crest serve as precursors to the cells in the dental follicle and dental papilla. Therefore, the current... (Comparative Study)
Comparative Study
INTRODUCTION
During tooth development, cells originating from the neural crest serve as precursors to the cells in the dental follicle and dental papilla. Therefore, the current study aimed to understand the associations of cranial neural crest cells (CNCCs), dental follicle cells (DFCs), and dental papilla cells (DPCs) by performing a parallel comparison to evaluate their odontogenic differentiation capacities.
METHODS
In this study, we harvested the 3 cells from C57/green fluorescent protein-positive mice or embryos and compared the cell morphology, surface antigens, microstructures, and gene and protein expression. Under the odontogenic microenvironments provided by treated dentin matrix, the odontogenic differentiations of the 3 cells were further compared in vitro and in vivo.
RESULTS
The gene levels of DFCs in neurofilament, tubulin, and nestin were close to the DPCs, and in alkaline phosphatase, osteopontin, dentin matrix protein 1, and dentin sialophosphoprotein were the lowest in the 3 cells. However, Western blot results showed that DFCs possessed more similar protein profiles to CNCCs than DPCs, including collagen 1, transforming growth factor beta 1, osteopontin, neurofilament, and dentin matrix protein 1. Meanwhile, DFCs as 1 source of dental stem cells possessed high potency in odontogenic differentiation in vitro. Moreover, similar dentinlike tissues were observed in all 3 groups in vivo.
CONCLUSIONS
CNCCs, DFCs, and DPCs possessed different biological characteristics in odontogenic differentiation.
Topics: Animals; Cell Differentiation; Cells, Cultured; Dental Papilla; Dental Sac; Mice, Inbred C57BL; Neural Crest; Odontogenesis
PubMed: 25882137
DOI: 10.1016/j.joen.2015.03.003 -
The Journal of Contemporary Dental... Feb 2020The aim of the study was to quantify the specific spatial displacement of gingival zenith (GZ) and determine a representative value for the interdental papilla height as...
AIM
The aim of the study was to quantify the specific spatial displacement of gingival zenith (GZ) and determine a representative value for the interdental papilla height as a percentage ratio of clinical crown length (CL), as measured from GZ in the maxillary anterior dentition.
MATERIALS AND METHODS
A total of 100 subjects and 1,200 interdental papillae were included. Eighty percent of the population presented with the gingival margin of lateral incisor (LI) teeth positioned coronally to the GZ of the ipsilateral canine (C) and central incisor (CI).
RESULTS
No significant difference was found between mesial and distal papilla proportion (DPP) of maxillary incisor groups. In the C group, numerical values showed higher DPP.
CONCLUSION
The data achieved provide the ideal numerical values for prosthetic, restorative, periodontal, implant, postorthodontic, and esthetic treatment outcomes.
CLINICAL SIGNIFICANCE
The study describes the importance of quantifying the ideal numerical values for prosthetic, restorative, periodontal, implant, postorthodontic, and esthetic treatment outcomes. The readers should understand to quantify the specific spatial displacement of GZ and determine a representative value for the interdental papilla height as a percentage ratio of clinical CL, as measured from GZ in the maxillary anterior dentition.
Topics: Esthetics; Esthetics, Dental; Gingiva; Incisor; Maxilla
PubMed: 32381829
DOI: No ID Found -
Archives of Oral Biology Mar 2022To explore the effects of transforming growth factor-β2 (TGF-β2) and TGF-β1 on the odontogenic and osteogenic differentiation of mesenchymal stem cells (MSCs).
OBJECTIVE
To explore the effects of transforming growth factor-β2 (TGF-β2) and TGF-β1 on the odontogenic and osteogenic differentiation of mesenchymal stem cells (MSCs).
DESIGN
We used lentiviral transduction to knock down TGF-β1 or TGF-β2 in stem cells from dental apical papilla (SCAPs), and to generate bone marrow mesenchymal stem cells (BMSCs) with overexpression of TGF-β1 or TGF-β2. We investigated the odontogenic and osteogenic differentiation abilities of these transductants in vitro and in vivo.
RESULTS
In vitro, TGF-β2 knockdown in SCAPs reduced the expression of odontoblast-related markers DSPP and DMP-1, and increased the expression of osteoblast-related markers OCN and RUNX-2. Conversely, TGF-β1 knockdown had the opposite effects. TGF-β2 overexpression promoted expression of odontoblast-related markers in BMSCs at early differentiation, but inhibited the expression of odontoblast-related markers at later stages. TGF-β2 overexpression attenuated expression of osteogenic-related markers in BMSCs, while TGF-β1 overexpression enhanced odontoblast-related and osteoblast-related markers. SCAP or BMSC transductants were transplanted underneath kidneys in vivo. Masson staining showed that knockdown of TGF-β1, but not TGF-β2 promoted the expression of type I collagen in SCAPs. Immunohistochemical staining showed that TGF-β2 knockdown inhibited DSPP expression in SCAPs, but TGF-β1 knockdown had no obvious effect on DSPP expression. In vivo, TGF-β1 overexpression and TGF-β2 overexpression had no effect on the expression of type I collagen and DSPP in BMSCs.
CONCLUSIONS
TGF-β2 promotes odontogenic differentiation of SCAPs and attenuates osteogenic differentiation of SCAPs and BMSCs. TGF-β1 promotes osteogenic differentiation of BMSCs and plays a complex role in regulating odontogenic differentiation of MSCs.
Topics: Cell Differentiation; Cell Proliferation; Cells, Cultured; Dental Papilla; Mesenchymal Stem Cells; Osteogenesis; Transforming Growth Factor beta1; Transforming Growth Factor beta2
PubMed: 35085927
DOI: 10.1016/j.archoralbio.2022.105357 -
Medicina Oral, Patologia Oral Y Cirugia... May 2017Primordial Odontogenic Tumor (POT) is a recently described odontogenic tumor characterized by a variably cellular loose fibrous tissue with areas similar to the dental...
BACKGROUND
Primordial Odontogenic Tumor (POT) is a recently described odontogenic tumor characterized by a variably cellular loose fibrous tissue with areas similar to the dental papilla, covered by cuboidal to columnar epithelium that resembles the internal epithelium of the enamel organ, surrounded at least partly by a delicate fibrous capsule. The purpose of this study was to investigate the possible histogenesis and biological behavior of this rare tumor by means of a wide immunohistochemical analysis of its epithelial and mesenchymal components.
MATERIAL AND METHODS
The immunoexpression of twenty-three different antibodies were evaluated in four cases of POT.
RESULTS
The epithelial cells that cover the periphery of the tumor showed immunopositivity for Cytokeratins 14 and 19, while Amelogenin, Glut-1, MOC-31, Caveolin-1. Galectin-3, PITX2, p53, Bax, Bcl-2, Survivin and PTEN were variably expressed in focal areas. The mesenchymal component of the tumor was positive for Vimentin, Syndecan-1, PITX2, Endoglin (CD105), CD 34, Cyclin D1, Bax, Bcl-2, Survivin and p53. PTEN and CD 90 showed a moderate positivity. BRAF V600E and Calretinin were negative in all samples. Cell proliferation markers (Ki-67, MCM-7) were expressed in <5% of the tumor cells.
CONCLUSIONS
According to these immunohistochemical findings, we may conclude that POT is a benign odontogenic tumor in which there is both epithelial and mesenchymal activity during its histogenesis, as there is expression of certain components in particular zones in both tissues that suggests this tumor develops during the immature (primordial) stage of tooth development, leading to its inclusion within the group of benign mixed epithelial and mesenchymal odontogenic tumours in the current World Health Organization classification of these lesions.
Topics: Adolescent; Antibodies, Neoplasm; Child, Preschool; Female; Humans; Immunohistochemistry; Jaw Neoplasms; Male; Odontogenic Tumors
PubMed: 28390134
DOI: 10.4317/medoral.21859 -
Stem Cell Research & Therapy Feb 2022Commitment of mouse dental papilla cells (mDPCs) to the odontoblast lineage is critical for dentin formation, and this biological process is regulated by a complex...
BACKGROUND
Commitment of mouse dental papilla cells (mDPCs) to the odontoblast lineage is critical for dentin formation, and this biological process is regulated by a complex transcription factor network. The transcription factor Mycn is a proto-oncogene that plays an important role in tumorigenesis and normal embryonic development. An early study revealed that Mycn is exclusively expressed in dental mesenchymal cells at E15.5, which implies a potential role of Mycn in dentinogenesis. However, the role of Mycn in dentin formation remains elusive. Thus, it is of considerable interest to elucidate the role of Mycn in dentin formation.
METHODS
Mycn; Osr2 (Mycn) and Mycn; K14 (Mycn) transgenic mice were generated, and micro-CT scans were performed to quantitatively analyse the volumetric differences in the molars and incisors of the mutants and their littermates. Mycn was also knocked down in vitro, and alkaline phosphatase (ALP) and alizarin red staining (ARS) were conducted. Cleavage under targets and tagmentation (CUT&Tag) analysis and dual luciferase assays were performed to identify direct downstream targets of Mycn. Immunofluorescence and immunochemistry staining and western blotting (WB) were performed to analyse the expression levels of potential targets. Quantitative PCR, WB, ALP and ARS were performed to test the rescue efficiency.
RESULTS
Mesenchymal ablation of Mycn (Mycn) led to defective dentin formation, while epithelial deletion (Mycn) had no obvious effects on tooth development. ALP and ARS staining revealed that the commitment capacity of mDPCs to the odontoblast lineage was compromised in Mycn mice. CUT&Tag analysis identified Klf4 as a potential direct target of Mycn, and a dual luciferase reporter assay verified that Mycn could bind to the promotor region of Klf4 and directly activate its transcription. Reciprocally, forced expression of Klf4 partially recovered the odontoblastic differentiation capacity of mDPCs with Mycn knockdown.
CONCLUSIONS
Our results elucidated that mesenchymal Mycn modulates the odontoblastic commitment of dental papilla cells by directly regulating Klf4. Our study illustrated the role of Mycn in dentin development and furthers our general comprehension of the transcription factor networks involved in the dentinogenesis process. Thus, these results may provide new insight into dentin hypoplasia and bioengineered dentin regeneration.
Topics: Animals; Cell Differentiation; Kruppel-Like Factor 4; Mice; N-Myc Proto-Oncogene Protein; Odontoblasts; Odontogenesis; Transcription Factors
PubMed: 35193672
DOI: 10.1186/s13287-022-02749-8 -
Scientific Reports May 2022Melatonin plays a critical role in promoting the proliferation of osteoblasts and the growth and development of dental papilla cells. However, the effect and mechanism...
Melatonin plays a critical role in promoting the proliferation of osteoblasts and the growth and development of dental papilla cells. However, the effect and mechanism of melatonin on the growth and development of ALCs still need to be explored. CCK8 assay was used for the evaluation of cell numbers. qRT-PCR was used to identify the differentially expressed genes in ALCs after melatonin treatment. The number and morphology of ALCs were investigated by confocal microscopy. Alkaline phosphatase assay and Alizarin red S staining were used for measuring mineralization. Then, we focused on observing the crucial factors of the signaling pathway by RNA-seq and qRT-PCR. Melatonin limited the cell number of ALCs in a dose-dependent manner and promoted the production of actin fibers. A high concentration of melatonin significantly promoted the mRNA levels of enamel matrix proteins and the formation of mineralized nodules. RNA-seq data showed that Wnt signaling pathway may be involved in the differentiation of ALCs under the influence of melatonin. This study suggests that melatonin plays a regulatory role in the cell number, differentiation, and mineralization of the ALCs, and then shows the relationship between the Wnt signaling pathway with the ALCs under melatonin.
Topics: Ameloblasts; Animals; Cell Differentiation; Cell Line; Cell Proliferation; Melatonin; Mice; Osteoblasts; Osteogenesis; Wnt Signaling Pathway
PubMed: 35581244
DOI: 10.1038/s41598-022-11912-3 -
Journal of Applied Toxicology : JAT Feb 2019Leachables from dental restoratives induce toxicity in gingival and pulp tissues and affect tissue regeneration/healing. Appropriate testing of these materials requires...
Leachables from dental restoratives induce toxicity in gingival and pulp tissues and affect tissue regeneration/healing. Appropriate testing of these materials requires a platform that mimics the in vivo environment and allows the architectural self-assembly of cells into tissue constructs. In this study, we employ a new 3D model to assess the impact of triethyleneglycol dimethacrylate (TEGDMA) on early organization and advanced recruitment/accumulation of immortalized mouse gingival fibroblasts (GFs) and dental papilla mesenchymal cells (DPMCs) in extracellular matrix. We hypothesize that TEGDMA (1) interferes with the developmental architecture of GFs and DPMCs, and (2) inhibits the deposition of mineral. To test these hypotheses, GFs and DPMCs were incubated with the soluble TEGDMA at concentrations (0-2.5) mmol/L. Diameter and thickness of the constructs were determined by microscopic analysis. Cell differentiation was assessed by immunocytochemistry and the secreted mineral detected by alizarin-red staining. TEGDMA interfered with the development of GFs and/or DPMCs microtissues in a dose-dependent manner by inhibiting growth of inter-spherical cell layers and decreasing spheroid size (four to six times). At low/moderate TEGDMA levels, GFs organoids retained their structures while reducing thickness up to 21%. In contrast, at low TEGDMA doses, architecture of DPMC organoids was altered and thickness decreased almost twofold. Overall, developmental ability of TEGDMA-exposed GFs and DPMCs depended on TEGDMA level. GFs constructs were more resistant to structural modifications. The employed 3D platform was proven as an efficient tool for quantifying the effects of leachables on tissue repair capacities of gingiva and dental pulp.
Topics: Animals; Cell Culture Techniques; Cell Differentiation; Cells, Cultured; Composite Resins; Dental Pulp; Extracellular Matrix; Fibroblasts; Gingiva; Mesenchymal Stem Cells; Mice, Inbred C57BL; Organoids; Polyethylene Glycols; Polymethacrylic Acids
PubMed: 30229966
DOI: 10.1002/jat.3714 -
Journal of Dental Research Feb 2020Collagen signaling is critical for proper bone and tooth formation. Discoidin domain receptor 2 (DDR2) is a collagen-activated tyrosine kinase receptor shown to be...
Collagen signaling is critical for proper bone and tooth formation. Discoidin domain receptor 2 (DDR2) is a collagen-activated tyrosine kinase receptor shown to be essential for skeletal development. Patients with loss of function mutations in develop spondylo-meta-epiphyseal dysplasia (SMED), a rare, autosomal recessive disorder characterized by short stature, short limbs, and craniofacial anomalies. A similar phenotype was observed in -deficient mice, which exhibit dwarfism and defective bone formation in the axial, appendicular, and cranial skeletons. However, it is not known if has a role in tooth formation. We first defined the expression pattern of during tooth formation using knock-in mice. expression was detected in the dental follicle/sac and dental papilla mesenchyme of developing teeth and in odontoblasts and the periodontal ligament (PDL) of adults. No LacZ staining was detected in wild-type littermates. This expression pattern suggests a potential role in the tooth and surrounding periodontium. To uncover the function of , we used mice, which contain a spontaneous 150-kb deletion in the locus to produce an effective null. In comparison with wild-type littermates, mice displayed disproportional tooth size (decreased root/crown ratio), delayed tooth root development, widened PDL space, and interradicular alveolar bone defects. mice also had abnormal collagen content associated with upregulation of periostin levels within the PDL. The delayed root formation and periodontal abnormalities may be related to defects in RUNX2-dependent differentiation of odontoblasts and osteoblasts; RUNX2-S319-P was reduced in PDLs from mice, and deletion of in primary cell cultures from dental pulp and PDL inhibited differentiation of cells to odontoblasts or osteoblasts, respectively. Together, our studies demonstrate odontoblast- and PDL-specific expression of in mature and immature teeth, as well as indicate that DDR2 signaling is important for normal tooth formation and maintenance of the surrounding periodontium.
Topics: Animals; Discoidin Domain Receptor 2; Discoidin Domain Receptors; Humans; Mice; Odontogenesis; Receptor Protein-Tyrosine Kinases; Receptors, Mitogen
PubMed: 31869264
DOI: 10.1177/0022034519892563