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Journal of Endodontics Dec 2023Osteolectin is a secreted glycoprotein of the C-type lectin domain superfamily, expressed in bone tissues and is reported as a novel osteogenic factor that promotes bone...
INTRODUCTION
Osteolectin is a secreted glycoprotein of the C-type lectin domain superfamily, expressed in bone tissues and is reported as a novel osteogenic factor that promotes bone regeneration. However, the effect of osteolectin on human dental pulp cells (hDPCs) has not been reported. Therefore, we aimed to investigate the odontoblastic differentiation of osteolectin in hDPCs and further attempt to reveal its underlying mechanism.
METHODS
Cytotoxicity assays were used to detect the cytotoxicity of osteolectin. The odontoblastic differentiation of hDPCs and its underlying mechanisms were measured by the alkaline phosphatase (ALP) activity, mineralized spots formation, and the gene and protein expression of odontoblastic differentiation through ALP staining, Alizarin red S staining, quantitative real-time polymerase chain reaction, and Western blot analysis, respectively.
RESULTS
WST-1 assay showed osteolectin at concentrations below 300 ng/ml was noncytotoxic and safe for hDPCs. The following experiment demonstrated that osteolectin could increase ALP activity, accelerate the mineralization process, and up-regulate the odontogenic differentiation markers in both gene and protein levels (P < .05). Osteolectin stimulated the phosphorylation of ERK, JNK, and Protein kinase B (AKT) in hDPCs. Extracellular signal-regulated kinase (ERK), Jun N-terminal kinase (JNK), and AKT inhibitors decreased ALP activity and mineralization capacity and suppressed the expression of dentin sialophosphoprotein and dentin matrix protein-1.
CONCLUSION
Osteolectin can promote odontoblastic differentiation of hDPCs, and the whole process may stimulate ERK, JNK, and AKT signaling pathways by increasing p-ERK, p-JNK, and p-AKT signals.
Topics: Humans; Proto-Oncogene Proteins c-akt; Extracellular Matrix Proteins; Dental Pulp; Cell Differentiation; Signal Transduction; Odontoblasts; Alkaline Phosphatase; Cells, Cultured; Cell Proliferation; Phosphoproteins
PubMed: 37774945
DOI: 10.1016/j.joen.2023.09.010 -
Frontiers in Physiology 2023Regenerative dentistry has rapidly progressed since the advancement of stem cell biology and material science. However, more emphasis has been placed on the success of... (Review)
Review
Regenerative dentistry has rapidly progressed since the advancement of stem cell biology and material science. However, more emphasis has been placed on the success of tissue formation than on how well the newly generated tissue retains the original structure and function. Once dentin is lost, tertiary dentinogenesis can be induced by new odontoblastic differentiation or re-activation of existing odontoblasts. The characteristic morphology of odontoblasts generates the tubular nature of dentin, which is a reservoir of fluid, ions, and a number of growth factors, and protects the inner pulp tissue. Therefore, understanding the dynamic but delicate process of new dentin formation by odontoblasts, or odontoblast-like cells, following dentinal defects is crucial. In this regard, various efforts have been conducted to identify novel molecules and materials that can promote the regeneration of dentin with strength and longevity. In this review, we focus on recent progress in dentin regeneration research with biological molecules identified, and discuss its potential in future clinical applications.
PubMed: 38148896
DOI: 10.3389/fphys.2023.1313927 -
Frontiers in Cell and Developmental... 2021Dentin, which composes most of the tooth structure, is formed by odontoblasts, long-lived post-mitotic cells maintained throughout the entire life of the tooth. In...
Dentin, which composes most of the tooth structure, is formed by odontoblasts, long-lived post-mitotic cells maintained throughout the entire life of the tooth. In mature odontoblasts, however, cellular activity is significantly weakened. Therefore, it is important to augment the cellular activity of mature odontoblasts to regenerate physiological dentin; however, no molecule regulating the cellular activity of mature odontoblasts has yet been identified. Here, we suggest that copine-7 (CPNE7) can reactivate the lost functions of mature odontoblasts by inducing autophagy. CPNE7 was observed to elevate the expression of microtubule-associated protein light chain 3-II (LC3-II), an autophagy marker, and autophagosome formation in the pre-odontoblast and mature odontoblast stages of human dental pulp cells. CPNE7-induced autophagy upregulated DSP and DMP-1, odontoblast differentiation and mineralization markers, and augmented dentin formation in mature odontoblasts. Furthermore, CPNE7 also upregulated NESTIN and TAU, which are expressed in the physiological odontoblast process, and stimulated the elongation of the odontoblast process by inducing autophagy. Moreover, lipofuscin, which progressively accumulates in long-lived post-mitotic cells and hinders their proper functions, was observed to be removed in recombinant CPNE7-treated mature odontoblasts. Thus, CPNE7-induced autophagy reactivated the function of mature odontoblasts and promoted the formation of physiological dentin . On the other hand, the well-known autophagy inducer, rapamycin, promoted odontoblast differentiation in pre-odontoblasts but did not properly reactivate the function of mature odontoblasts. These findings provide evidence that CPNE7 functionally reactivates mature odontoblasts and introduce its potential for dentinal loss-targeted clinical applications.
PubMed: 33981704
DOI: 10.3389/fcell.2021.655498 -
International Journal of Molecular... Jun 2023Human dental pulp stem cells (hDPSCs) possess remarkable self-renewal and multilineage differentiation ability. PER2, an essential circadian molecule, regulates various...
Human dental pulp stem cells (hDPSCs) possess remarkable self-renewal and multilineage differentiation ability. PER2, an essential circadian molecule, regulates various physiological processes. Evidence suggests that circadian rhythm and PER2 participate in physiological functions of DPSCs. However, the influence of PER2 on DPSCs' differentiation remains largely unknown. This study aimed to explore the effect and potential mechanism of PER2 on hDPSCs' differentiation. Dental pulp tissues were extracted, and hDPSCs were cultured for and experiments. Dorsal subcutaneous transplantation was performed in 6-week-old male BALB/c mice. The hDPSCs' odontoblastic/osteogenic differentiation was assessed, and mitochondrial metabolism was evaluated. The results indicated PER2 expression increasing during hDPSCs' odontoblastic/osteogenic differentiation. Gain- and loss-of function studies confirmed that PER2 promoted alkaline phosphatase (ALP) activity, mineralized nodules deposition, mRNA expression of , , and protein expression of DSPP and DMP1 in hDPSCs. Furthermore, PER2 enhanced collagen deposition, osteodentine-like tissue formation and DSPP expression . Mitochondrial metabolic evaluation aimed to investigate the mechanism of PER2-mediated hDPSC odontoblastic/osteogenic differentiation, which showed that PER2 increased ATP synthesis, elevated mitochondrial membrane potential and changed expression of proteins regulating mitochondrial dynamics. This study demonstrated that PER2 promoted hDPSCs' odontoblastic/osteogenic differentiation, which involved mitochondrial metabolic change.
Topics: Animals; Mice; Humans; Male; Osteogenesis; Dental Pulp; Odontoblasts; Cell Differentiation; Stem Cells; Cells, Cultured; Cell Proliferation; Period Circadian Proteins
PubMed: 37445839
DOI: 10.3390/ijms241310661 -
Stomatologiia 2023Dentinal fluid is very close in its physical and mechanical properties and composition to blood plasma, which makes it a potentially aggressive biological environment...
BACKGROUND
Dentinal fluid is very close in its physical and mechanical properties and composition to blood plasma, which makes it a potentially aggressive biological environment for modern adhesive systems. An in-depth study of the physiological processes of the functioning of tooth dentin remains relevant in order to solve problems associated with its artificial restoration.
PURPOSE OF THE STUDY
Study using computer simulation speed of movement and pressure distribution of dentinal fluid in the dentinal tubule of the tooth to assess the possibilities of their regulation.
MATERIAL AND METHODS
To model the distribution of flow velocity and pressure of dentinal fluid in the dentinal tubule, the finite element method (Fluent ANSYS computer program) was used.
RESULTS
Immediately behind the spherical tip of the odontoblast, there is a rapid increase in the hydraulic diameter of the flow section of the dentinal tubule, and, accordingly, a decrease in capillary pressure, while the tip of the odontoblast creates a large local hydraulic resistance. The resulting distribution of pressure drop in the damaged dentinal tubule is consistent with the fact that fluid movement is due, to a greater extent, to the capillary effect rather than the inlet pressure into the dentinal tubule.
CONCLUSION
By changing the length of the odontoblast process, it is possible to influence the parameters of the hydrodynamics of dentinal fluid in the space of the dentinal tubule.
Topics: Humans; Dentin; Dentinal Fluid; Computer Simulation; Hydrodynamics
PubMed: 38096387
DOI: 10.17116/stomat20231020625 -
Frontiers in Cell and Developmental... 2021Dental pulp stem cells (DPSCs) are a source of postnatal stem cells essential for maintenance and regeneration of dentin and pulp tissues. Previous transplantation...
Dental pulp stem cells (DPSCs) are a source of postnatal stem cells essential for maintenance and regeneration of dentin and pulp tissues. Previous transplantation studies have shown that DPSCs are able to give rise to odontoblast-like cells, form dentin/pulp-like structures, and induce blood vessel formation. Importantly, dentin formation is closely associated to blood vessels. We have previously demonstrated that DPSC-induced angiogenesis is VEGFR-2-dependent. VEGFR-2 may play an important role in odontoblast differentiation of DPSCs, tooth formation and regeneration. Nevertheless, the role of VEGFR-2 signaling in odontoblast differentiation of DPSCs is still not well understood. Thus, in this study we aimed to determine the role of VEGFR-2 in odontoblast differentiation of DPSCs by knocking down the expression of VEGFR-2 in DPSCs and studying their odontoblast differentiation capacity and . Isolation and characterization of murine DPSCs was performed as previously described. DPSCs were induced by VEGFR-2 shRNA viral vectors transfection (MOI = 10:1) to silence the expression of VEGFR-2. The GFP+ expression in CopGFP DPSCs was used as a surrogate to measure the efficiency of transfection and verification that the viral vector does not affect the expression of VEGFR-2. The efficiency of viral transfection was shown by significant reduction in the levels of VEGFR-2 based on the Q-RT-PCR and immunofluorescence in VEGFR-2 knockdown DPSCs, compared to normal DPSCs. VEGFR-2 shRNA DPSCs expressed not only very low level of VEGFR-2, but also that of its ligand, VEGF-A, compared to CopGFP DPSCs in both transcriptional and translational levels. differentiation of DPSCs in osteo-odontogenic media supplemented with BMP-2 (100 ng/ml) for 21 days demonstrated that CopGFP DPSCs, but not VEGFR-2 shRNA DPSCs, were positive for alkaline phosphatase (ALP) staining and formed mineralized nodules demonstrated by positive Alizarin Red S staining. The expression levels of dentin matrix proteins, dentin matrix protein-1 (), dentin sialoprotein (), and bone sialoprotein (), were also up-regulated in differentiated CopGFP DPSCs, compared to those in VEGFR-2 shRNA DPSCs, suggesting an impairment of odontoblast differentiation in VEGFR-2 shRNA DPSCs. subcutaneous transplantation of DPSCs with hydroxyapatite (HAp/TCP) for 5 weeks demonstrated that CopGFP DPSCs were able to differentiate into elongated and polarized odontoblast-like cells forming loose connective tissue resembling pulp-like structures with abundant blood vessels, as demonstrated by H&E, Alizarin Red S, and dentin matrix staining. On the other hand, in VEGFR-2 shRNA DPSC transplants, odontoblast-like cells were not observed. Collagen fibers were seen in replacement of dentin/pulp-like structures. These results indicate that VEGFR-2 may play an important role in dentin regeneration and highlight the potential of VEGFR-2 modulation to enhance dentin regeneration and tissue engineering as a promising clinical application.
PubMed: 34249919
DOI: 10.3389/fcell.2021.665886 -
Journal of Veterinary Dentistry Dec 2022This review describes the clinical, radiographic and histologic characteristics of dentinogenesis imperfecta diagnosed in two unrelated young dogs without evidence of... (Review)
Review
This review describes the clinical, radiographic and histologic characteristics of dentinogenesis imperfecta diagnosed in two unrelated young dogs without evidence of concurrent osteogenesis imperfecta. The dentition was noted to have generalized coronal discoloration ranging from grey-blue to golden brown. Clinical pulp exposure, coronal wear and fractures were observed as was radiographic evidence of endodontic disease, thin dentin walls or dystrophic obliteration of the pulp canal. The enamel was severely affected by attrition and abrasion despite histologically normal areas; loss was most likely due to poor adherence or support by the underlying abnormal dentin. Histologically, permanent and deciduous teeth examined showed thin, amorphous dentin without organized dentin tubules and odontoblasts had dysplastic cell morphology. Primary dentin disorders, including dentinogenesis imperfecta and dentin dysplasia, have been extensively studied and genetically characterized in humans but infrequently reported in dogs. Treatment in human patients is aimed at early recognition and multi-disciplinary intervention to restore and maintain normal occlusion, aesthetics, mastication and speech. Treatment in both humans and canine patients is discussed as is the documented genetic heritability of primary dentin disorders in humans.
Topics: Humans; Dogs; Animals; Dentinogenesis Imperfecta; Esthetics, Dental; Odontoblasts; Osteogenesis Imperfecta; Dentin; Dog Diseases
PubMed: 36113440
DOI: 10.1177/08987564221123419 -
Frontiers in Physiology 2020Dentin sialoprotein (DSP), the NH2-terminal fragment of dentin sialophosphoprotein (DSPP), is essential for dentin formation and further processed into small fragments...
Dentin sialoprotein (DSP), the NH2-terminal fragment of dentin sialophosphoprotein (DSPP), is essential for dentin formation and further processed into small fragments inside the odontoblasts. Gelatinases, including matrix metalloproteinases 9 (MMP9) and MMP2, were able to cleave DSP(P) in tooth structures. We hypothesized that gelatinases may also cleave DSP intracellularly in the odontoblasts. In this study, the co-expression and physical interaction between DSP and gelatinases were proved by double immunofluorescence and proximity ligation assay (PLA). Intracellular enzymatic activity of gelatinases was verified by gelatin zymography and zymography. To confirm whether DSP was cleaved by active gelatinases intracellularly, lysates of odontoblastic cells treated with a MMP2 inhibitor or a MMP9 inhibitor or a MMP general inhibitor and of odontoblastic cells were analyzed by western blotting. Compared with the odontoblastic cells without inhibitor treatment, all these groups exhibited significantly higher ratios of high molecular weight to low molecular weight band density. FURIN was verified to be co-localized and physically interacted with MMP9 by double immunofluorescence and PLA. The ratio of proMMP9 to activated MMP9 inside the odontoblastic cells were increased when function of endogenous FURIN was inhibited. And overexpressed proMMP9 was intracellularly cleaved by FURIN in the HEK293E cells, which was completely blocked by the mutation of proMMP9 with RTPR substituted by AAAA. Taken together, these results indicate that DSP is intracellularly processed by gelatinases, and FURIN is involved in the intracellular activation of proMMP9 through cleavage of its RTPR motif.
PubMed: 32670089
DOI: 10.3389/fphys.2020.00686 -
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 -
Iranian Endodontic Journal 2020Dental problems are common in human populations. Traditional treatments are focused on managing caries, soft tissue impairments, functional defects, poor aesthetics,... (Review)
Review
Dental problems are common in human populations. Traditional treatments are focused on managing caries, soft tissue impairments, functional defects, poor aesthetics, digestive disorders and alveolar bone resorption. During the last two decades, basic and clinical researches on adult stem cells have established a potential therapeutic concept in tissue regeneration. Among major cells responsible for tooth development, odontoblasts play a key role in the formation of organic and inorganic constituents of dental tissue. A premier stride in the development of novel stem cell-based strategies for the treatment of reversible and irreversible pulpitis is odontoblast regeneration. Among different candidate cell sources for odontoblastic regeneration, use of dental adult stem cells is a preferred option because of their great ability to differentiate into odontoblasts and also their minimally invasive isolation procedure. This review emphasizes on articles that report successful odontoblast-like differentiation of dental mesenchymal stem cells which in turn provide a background for dentin-pulp complex cell therapies, using genetic or chemical manipulation. The series of experiments both and asserted that dental mesenchymal stem cells can efficiently differentiate into functional odontoblast-like cells. However, the review shows there are drawbacks in present methods. Future research should focus on optimizing protocols on odontoblast differentiation of dental stem cells by simultaneously introducing different genes with mutual synergy, combined with chemical or recombinant protein introduction.
PubMed: 36704441
DOI: 10.22037/iej.v15i2.27569