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Cells Feb 2024Regenerative endodontic procedures (REPs) are promising for dental pulp tissue regeneration; however, their application in permanent teeth remains challenging. We...
Regenerative endodontic procedures (REPs) are promising for dental pulp tissue regeneration; however, their application in permanent teeth remains challenging. We assessed the potential combination of an REP and local dental pulp cell (DPC) transplantation in the mature molars of C57BL/6 mice with (REP + DPC group) or without (REP group) transplantation of DPCs from green fluorescent protein (GFP) transgenic mice. After 4 weeks, the regenerated tissue was evaluated by micro-computed tomography and histological analyses to detect odontoblasts, vasculogenesis, and neurogenesis. DPCs were assessed for mesenchymal and pluripotency markers. Four weeks after the REP, the molars showed no signs of periapical lesions, and both the REP and REP + DPC groups exhibited a pulp-like tissue composed of a cellular matrix with vessels surrounded by an eosin-stained acellular matrix that resembled hard tissue. However, the REP + DPC group had a broader cellular matrix and uniquely contained odontoblast-like cells co-expressing GFP. Vasculogenesis and neurogenesis were detected in both groups, with the former being more prominent in the REP + DPC group. Overall, the REP was achieved in mature mouse molars and DPC transplantation improved the outcomes by inducing the formation of odontoblast-like cells and greater vasculogenesis.
Topics: Mice; Animals; Regenerative Endodontics; Dental Pulp; X-Ray Microtomography; Mice, Inbred C57BL; Dentin; Cell Transplantation
PubMed: 38391961
DOI: 10.3390/cells13040348 -
Journal of Developmental Biology Jan 2024Hyperplastic dental follicles (HDFs) represent odontogenic hamartomatous lesions originating from the pericoronal tissues and are often associated with impacted or...
Hyperplastic dental follicles (HDFs) represent odontogenic hamartomatous lesions originating from the pericoronal tissues and are often associated with impacted or embedded teeth. These lesions may occasionally feature unique calcifying bodies, known as calcifying whorled nodules (CWNs), characterized by stromal cells arranged in a whorled or spiral fashion. CWNs are typically observed in multiple calcifying hyperplastic dental follicles or regional odontodysplasia. In our study, we examined 40 cases of HDFs, including nine instances with characteristics of CWNs, referred to as calcifying hyperplastic dental follicles (CHDFs), which are infrequently accompanied by odontodysplasia. The median ages of the HDFs and CHDFs were 16 (ranging from 3 to 66) and 15 (ranging from 11 to 50) years, respectively. The lower third molars were the most frequently affected by HDSFs and CHDFs, followed by the upper canines. A histological examination was conducted on all 40 cases, with an immunohistochemical analysis performed on 21 of them. Among the cases with CWN, nine affected a single embedded tooth, with one exception. CWNs exhibited diverse calcifications featuring sparse or entirely deposited psammoma bodies, and some displayed dentinoid formation. Immunohistochemically, the stromal cells of HDFs were frequently positive for CD56 and nestin. By contrast, CWNs were negative for CD56 but positive for nestin as well as hairy and enhancer split 1 (HES1), with a few dentin sialoprotein (DSP)-positive calcified bodies. Our results revealed that hamartomatous CHDFs can impact multiple and single-embedded teeth. CWNs composed of nestin and HES1-positive ectomesenchymal cells demonstrated the potential to differentiate into odontoblasts and contribute to dentin matrix formation under the influence of HES1. This study is the first report documenting odontoblastic differentiation in HDFs. The rare occurrence of HDFs and CHDFs contributes to limited comprehension. To prevent misdiagnosis, a better understanding of these conditions is necessary.
PubMed: 38390958
DOI: 10.3390/jdb12010007 -
Alternatives To Laboratory Animals :... Mar 2024models of the dental pulp microenvironment have been proposed for the assessment of biomaterials, to minimise animal use in operative dentistry. In this study, a...
models of the dental pulp microenvironment have been proposed for the assessment of biomaterials, to minimise animal use in operative dentistry. In this study, a scaffold/3-D dental pulp cell culture interface was created in a microchip, under simulated dental pulp pressure, to evaluate the cell-homing potential of a chitosan (CH) scaffold functionalised with calcium aluminate (the 'CHAlCa scaffold'). This microphysiological platform was cultured at a pressure of 15 cm HO for up to 14 days; cell viability, migration and odontoblastic differentiation were then assessed. The CHAlCa scaffold exhibited intense chemotactic potential, causing cells to migrate from the 3-D culture to its surface, followed by infiltration into the macroporous structure of the scaffold. By contrast, the cells in the presence of the non-functionalised chitosan scaffold showed low cell migration and no cell infiltration. CHAlCa scaffold bioactivity was confirmed in dentin sialophosphoprotein-positive migrating cells, and odontoblastic markers were upregulated in 3-D culture. Finally, mineralised matrix deposition by the cells was confirmed in an Alizarin Red-based assay, in which the CHAlCa and CH scaffolds were adapted to fit within dentin discs. More intense deposition of matrix was observed with the CHAlCa scaffold, as compared to the CH scaffold. In summary, we present an platform that provides a simple and reproducible model for selecting and developing innovative biomaterials through the assessment of their cell-homing potential. By using this platform, it was shown that the combination of calcium aluminate and chitosan has potential as an inductive biomaterial that can mediate dentin tissue regeneration during cell-homing therapies.
Topics: Animals; Chitosan; Tissue Scaffolds; Dental Pulp; Biocompatible Materials; Cell Differentiation; Cells, Cultured; Tissue Engineering; Aluminum Compounds; Calcium Compounds
PubMed: 38351650
DOI: 10.1177/02611929241232558 -
Annals of Anatomy = Anatomischer... Apr 2024Vascularization is an essential event for both embryonic organ development and tissue repair in adults. During mouse tooth development, endothelial cells migrate into...
BACKGROUND
Vascularization is an essential event for both embryonic organ development and tissue repair in adults. During mouse tooth development, endothelial cells migrate into dental papilla during the cap stage, and form blood vessels through angiogenesis. Megakaryocytes and/or platelets, as other hematopoietic cells, express angiogenic molecules and can promote angiogenesis in adult tissues. However, it remains unknown which cells are responsible for attracting and leading blood vessels through the dental papilla during tooth development.
METHODS
Here we analyzed the spatiotemporal expression of c-Mpl mRNA in developing molar teeth of fetal mice. Expression patterns were then compared with those of several markers of hematopoietic cells as well as of angiogenic elements including CD41, erythropoietin receptor, CD34, angiopoietin-1 (Ang-1), Tie-2, and vascular endothelial growth factor receptor2 (VEGFR2) through in situ hybridization or immunohistochemistry.
RESULTS
Cells expressing c-Mpl mRNA was found in several parts of the developing tooth germ, including the peridental mesenchyme, dental papilla, enamel organ, and dental lamina. This expression occurred in a spatiotemporally controlled fashion. CD41-expressing cells were not detected during tooth development. The spatiotemporal expression pattern of c-Mpl mRNA in the dental papilla was similar to that of Ang-1, which preceded invasion of endothelial cells. Eventually, at the early bell stage, the c-Mpl mRNA signal was detected in morphologically differentiating odontoblasts that accumulated in the periphery of the dental papilla along the inner enamel epithelium layer of the future cusp region.
CONCLUSION
During tooth development, several kinds of cells express c-Mpl mRNA in a spatiotemporally controlled fashion, including differentiating odontoblasts. We hypothesize that c-Mpl-expressing cells appearing in the forming dental papilla at the cap stage are odontoblast progenitor cells that migrate to the site of odontoblast differentiation. There they attract vascular endothelial cells into the forming dental papilla and lead cells toward the inner enamel epithelium layer through production of angiogenic molecules (e.g., Ang-1) during migration to the site of differentiation. C-Mpl may regulate apoptosis and/or proliferation of expressing cells in order to execute normal development of the tooth.
Topics: Animals; Mice; Endothelial Cells; RNA, Messenger; Tooth; Tooth Germ; Vascular Endothelial Growth Factor A
PubMed: 38336176
DOI: 10.1016/j.aanat.2024.152227 -
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 -
Frontiers in Cell and Developmental... 2024Dental caries is one of the most common human pathological conditions resulting from the invasion of bacteria into the dentin. Current treatment options are limited. In...
Dental caries is one of the most common human pathological conditions resulting from the invasion of bacteria into the dentin. Current treatment options are limited. In many cases, endodontic therapy leads to permanent pulp tissue loss. Dentin-pulp complex regeneration involves dental pulp stem cells (DPSCs) that differentiate into odontoblast-like cells under an inflammatory context. However, limited information is available on how DPSC differentiation processes are affected under inflammatory environments. We identified the crucial role of complement C5a and its receptor C5aR in the inflammation-induced odontoblastic DPSC differentiation. Here, we further investigated the role of a second and controversial C5a receptor, C5L2, in this process and explored the underlying mechanism. Human DPSCs were examined during 7-, 10-, and 14-day odontogenic differentiation treated with TNFα, C5L2 CRISPR, and tyrosine receptor kinase B (TrkB) antagonist [cyclotraxin-B (CTX-B)]. Our data demonstrate that C5L2 CRISPR knockout (KO) enhances mineralization in TNFα-stimulated differentiating DPSCs. We further confirmed that C5L2 CRISPR KO significantly enhances dentin sialophosphoprotein (DSPP) and dentin matrix protein-1 (DMP-1) expression after 14-day odontoblastic DPSC differentiation, and treatment with CTX-B abolished the TNFα/C5L2 CRISPR KO-induced DSPP and DMP-1 increase, suggesting TrkB's critical role in this process. Our data suggest a regulatory role of C5L2 and TrkB in the TNFα-induced odontogenic DPSC differentiation. This study may provide a useful tool to understand the mechanisms of the role of inflammation in dentinogenesis that is required for successful DPSC engineering strategies.
PubMed: 38318114
DOI: 10.3389/fcell.2024.1338419 -
Revista Cientifica Odontologica... 2023Identify the histological characteristics of bovine dentin using histological technique by decalcification with Masson's Trichrome staining.
OBJECTIVES
Identify the histological characteristics of bovine dentin using histological technique by decalcification with Masson's Trichrome staining.
MATERIALS AND METHODS
This was an observational and descriptive study using units of analysis. Bovine teeth were used, which were subjected to a decalcification technique and Masson's trichrome staining. Microscopic observation and registration of the structures was carried out.
RESULTS
In the microscopic observation of the teeth subjected to decalcification, dentin, predentin and the structural organization of the tubular and peritubular dentin were observed. The odontoblasts arranged in a palisade with the odontoblastic process included within the dentinal tubule and the periprocedural space. The presence of interglobular dentin is not observed in areas of coronary dentin. The histomorphological pattern corresponding to dentin-predentin-pulp is arranged in a similar way to human teeth.
CONCLUSIONS
Bovine dentin constitutes an ideal substrate for in vitro research work with dental biomaterials, since it presents histological characteristics similar to bovine teeth; the absence of interglobular dentin would not represent a significant difference according to what was observed in this study.
PubMed: 38312466
DOI: 10.21142/2523-2754-1104-2023-176 -
Archives of Oral Biology Apr 2024To evaluate the regulatory effect of transcription factor EB (TFEB) on the odontoblastic differentiation of dental pulp stem cells(DPSCs) in vivo and in vitro.
OBJECTIVE
To evaluate the regulatory effect of transcription factor EB (TFEB) on the odontoblastic differentiation of dental pulp stem cells(DPSCs) in vivo and in vitro.
DESIGNS
RNA-seq was used to detect differentially expressed genes in differentiated DPSCs. Lysosomes and the expression of the related gene TFEB were examined in DPSCs. DPSCs were then transfected with lentivirus for TFEB-overexpression. Cell proliferation was detected using CCK-8 and EdU assays, while cell differentiation was detected using ALP and ARS detection kits. Subsequently, mitophagy and cell metabolism were examined using TEM and Seahorse. An odontoblastic differentiation model was constructed subcutaneously in nude mice. Finally, the effects of glycolysis and mitophagy inhibitors were evaluated on odontoblastic differentiation and the associated mechanisms were explored.
RESULTS
TFEB overexpression promoted a significant increase in ALP activity and the expression of differentiation-related genes in DPSCs, while it inhibited cell proliferation. In vivo, TFEB overexpression caused higher bone volume/trabecular volume(BV/TV), and an increase in collagen formation and heightened DMP-1 expression. Furthermore, Seahorse flux analysis demonstrated that TFEB promoted metabolic reprogramming. Transmission electron microscope(TEM) results indicated an increase in mitochondrial autophagosomes after TFEB overexpression, and the expression of mitophagy-related genes was also elevated. The odontoblastic differentiation of DPSCs promoted by TFEB overexpression was suppressed after the addition of 2-DG and Midiv-1. Addition of Midiv-1 reduced the glycolytic rate of DPSCs, while addition of 2-DG also decreased the mitophagy level of the cells.
CONCLUSIONS
Our results showed that TFEB promoted the odontoblastic differentiation of DPSCs and identified mitophagy and metabolic reprogramming as a positive feedback loop.
Topics: Animals; Mice; Cell Differentiation; Cell Proliferation; Cells, Cultured; Dental Pulp; Feedback; Mice, Nude; Mitophagy; Odontoblasts; Stem Cells; Humans
PubMed: 38309196
DOI: 10.1016/j.archoralbio.2024.105909 -
International Journal of Biological... Mar 2024Incorporation of growth factors, signaling molecules and drugs can be vital for the success of tissue engineering in complex structures such as the dentoalveolar region....
Development and characterization of colloidal pNIPAM-methylcellulose microgels with potential application for drug delivery in dentoalveolar tissue engineering strategies.
Incorporation of growth factors, signaling molecules and drugs can be vital for the success of tissue engineering in complex structures such as the dentoalveolar region. This has led to the development of a variety of drug release systems. This study aimed to develop pNIPAM-methylcellulose microgels with different synthesis parameters based on a 2 full factorial design of experiments for this application. Microgel properties, including volume phase transition temperature (VPTT), hydrodynamic size, drug loading and release, and cytocompatibility were systematically evaluated. The results demonstrated successful copolymerization and development of the microgels, a hydrodynamic size ranging from ∼200 to ∼500 nm, and VPTT in the range of 34-39 °C. Furthermore, loading of genipin, capable of inducing odontoblastic differentiation, and its sustained release over a week was shown in all formulations. Together, this can serve as a solid basis for the development of tunable drug-delivering pNIPAM-methylcellulose microgels for specific tissue engineering applications.
Topics: Microgels; Gels; Methylcellulose; Tissue Engineering; Transition Temperature
PubMed: 38307741
DOI: 10.1016/j.ijbiomac.2024.129684 -
Journal of Dental Sciences Jan 2024High glucose (HG)-induced aberrant proliferation, apoptosis and odontoblastic differentiation of dental pulp cells (DPCs) have been implicated in the pathogenesis of...
BACKGROUND/PURPOSE
High glucose (HG)-induced aberrant proliferation, apoptosis and odontoblastic differentiation of dental pulp cells (DPCs) have been implicated in the pathogenesis of impaired diabetic pulp healing; however, the underlying mechanism remains unclear. This study aimed to investigate the role of mitochondrial reactive oxygen species (mtROS) and mitochondria in HG-induced dysfunction and apoptosis of DPCs.
MATERIALS AND METHODS
Human DPCs (hDPCs) were cultured in a low-glucose, high-glucose, mannitol, and MitoTEMPO medium in vitro. Methylthiazol tetrazolium assay, Annexin V-FITC/PI staining and scratch-wound assay were used to analyze cell proliferation, apoptosis and migration, respectively. Alkaline phosphatase staining and alizarin red S staining were used to evaluate cell differentiation. DCF-DA staining, MitoSOX staining, MitoTracker Red staining, JC-1 staining, and adenosine triphosphate (ATP) kit assay were performed to investigate total ROS and mtROS generation, mitochondrial density, mitochondrial membrane potential (MMP), and ATP synthesis, respectively. Quantitative PCR assay was performed to detect the mRNA expression of mitochondrial biogenesis- and dynamics-related markers. Transmission electron microscopy was used to observe the mitochondrial ultrastructure.
RESULTS
HG augmented the production of total ROS and mtROS, and triggered mitochondrial damage in hDPCs, as reflected by decreased mitochondrial density, depolarized MMP, reduced ATP synthesis, altered mRNA expression of mitochondrial biogenesis- and dynamics-related markers, and abnormal mitochondrial ultrastructure. Supplementation of MitoTEMPO alleviated the mitochondrial damage and reversed the aberrant proliferation, apoptosis, migration and odontoblastic differentiation of HG-stimulated hDPCs.
CONCLUSION
HG triggers mitochondrial damage via augmenting mtROS generation, resulting in the inhibited proliferation, migration, and odontoblastic differentiation of hDPCs and enhanced their apoptosis.
PubMed: 38303803
DOI: 10.1016/j.jds.2023.04.008