Differentiation Of Dental Stem Cells Research Articles

Effects of Growth Factors on the Differentiation of Dental Stem Cells: A Systematic Review and Meta-analysis (Part I).

To evaluate the biological interaction between dental stem cells (DSCs) and different growth factors in the field of regenerative endodontics. A systematic search was conducted in the electronic databases up to October 2021. This study followed the preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines. Ex vivo studies evaluating the biological interactions of DSCs and growth factors were included. The meta-analysis was performed according to the type of growth factor. The outcomes were cell viability/ proliferation and mineralization. Standardized mean differences (SMDs) were estimated using the random-effect maximum-likelihood method (P < .05). Additional analysis was performed to find any potential source of heterogeneity. Twenty articles were included in the systematic review; meta-analysis was performed for fibroblast growth factor-2 (FGF-2) and Transforming growth factor-ß1 (TGF-β1) (n = 5). Results showed that use of FGF-2 significantly increased cell proliferation on day 1-(SMD = 3.56, P = 0.00), 3-(SMD = 9.04, P = 0.00), 5-(SMD = 8.37, P = 0.01), and 7 (SMD=8.51, P=0.00) than the control group. TGF-ß1 increased alkaline phosphatase (ALP) activity more than control only on day 3 (SMD = 3.68, P = 0.02). TGF-β1 had no significant effect on cell proliferation on days 1 and 3 (P > 0.05) and on ALP activity on days 5 and 7 (P > 0.05). Meta-regression analysis showed that different covariates (i.e., cell type, passage number, and growth factors' concentration) could significantly influence the effect sizes at different follow- ups (P < 0.05). Specific growth factors might enhance the proliferation and mineralization of DSCs; however, the obtained evidence was weak. Due to the high heterogeneity among the included studies, other growth factors' inhibitory/stimulatory effects on DSCs could not be evaluated.

An overview of autophagy in the differentiation of dental stem cells

An overview of autophagy in the differentiation of dental stem cells

Dental Tissue Engineering by Neural Differentiation of Dental Stem Cells and Nano-systems: A Review

Background: Pulpitis is a pulpal inflammation. It generally occurs when there is inflammation within a tooth as a result of anything like grinding or decay. After dental inflammation, microcirculation and sensory nerve activity seem to play the most critical role in reducing inflammation. Therefore, researchers emphasize the study of dental nerve activity, especially in acute clinical problems in inflamed teeth and pulp regeneration. This review aims to investigate the possibility of using dental stem cells to regenerate dental nerves in order to repair dentin-pulp complexes for maintaining and restoring tooth structure and function, which nanosystems can help in this matter. Materials and Methods: In this paper, we review the literature regarding the theory of dental tissue engineering by neural differentiation of dental stem cells and nano-systems, and the comprehensive search on PubMed, Scopus, and Web of Science was conducted up to July 2022. Results: According to recent studies, dental soft and hard tissue healing also includes nerve fibers. A deeper understanding of how dental nerves are implicated in pulpitis may assist endodontic treatment. Stem cell-based treatments may be used to regenerate dental nerves to repair dentin-pulp complexes to maintain and restore tooth structure and function. Conclusion: The emphasis on dental nerve regeneration appears to be a critical stage in fostering spontaneous tooth regeneration as well as a sustainable tooth regeneration method. It is essential to further investigate dental tissue engineering by neural differentiation of dental stem cells.

MicroRNAs Function in Dental Stem Cells as a Promising Biomarker and Therapeutic Target for Dental Diseases.

Undifferentiated, highly proliferative, clonogenic, and self-renewing dental stem cells have paved the way for novel approaches to mending cleft palates, rebuilding lost jawbone and periodontal tissue, and, most significantly, recreating lost teeth. New treatment techniques may be guided by a better understanding of these cells and their potential in terms of the specificity of theregenerative response. MicroRNAs have been recognized as an essential component in stem cell biology due to their role as epigenetic regulators of the processes that determine stem cell destiny. MicroRNAs have been proven to be crucial in a wide variety of molecular and biological processes, including apoptosis, cell proliferation, migration, and necrocytosis. MicroRNAs have been recognized to control protein translation, messenger RNA stability, and transcription and have been reported to play essential roles in dental stem cell biology, including the differentiation of dental stem cells, the immunological response, apoptosis, and the inflammation of the dental pulp. Because microRNAs increase dental stem cell differentiation, they may be used in regenerative medicine to either preserve the stem cell phenotype or to aid in the development of tooth tissue. The development of novel biomarkers and therapies for dental illnesses relies heavily on progress made in our knowledge of the roles played by microRNAs in regulating dental stem cells. In this article, we discuss how dental stem cells and their associated microRNAs may be used to cure dental illness.

KLF9 Promotes Osteogenic Differentiation of Dental Stem Cells by Negatively Regulating Notch1 Mediated Signaling Pathway.

Human dental stem cells (DSCs) are excellent sources of cells for treating dental and craniofacial diseases. However, the mechanisms regulating DSC osteogenic differentiation are still unclear. In this study, we aimed to determine the role of Krüppel-like factor 9 (KLF9) in regulating the biological functions of DSCs and explore the underlying molecular mechanisms. Bioinformatic analyses, quantitative real-time polymerase chain reaction (qRT‒PCR) and Western blotting were performed to determine the KLF9 level during osteogenic differentiation of DSCs. The effects of KLF9 depletion or overexpression on DSC osteogenic differentiation were then evaluated. The osteogenic potential and associated mineralized nodule-forming activities of DSCs were monitored via Alizarin red S staining and quantitative analyses of osteogenic markers. The regulatory effect of KLF9 on the Notch1 signaling pathway was analyzed by luciferase reporter assays. KLF9 mRNA expression was consistently increased during mesenchymal stem cell osteogenic differentiation in multiple public datasets, and our qRT‒PCR and Western blotting data further validated this finding. In addition, KLF9 depletion promoted proliferation and suppressed osteogenic differentiation of DSCs, while enforced expression of KLF9 promoted the DSC osteogenic potential. Mechanistically, KLF9 negatively regulated the Notch1-mediated signaling pathway by directly binding to the Notch1 promoter. More importantly, Notch1 inhibition/overexpression partially rescued the suppressive/enhancing effects of KLF9 depletion/overexpression on the osteogenic differentiation of DSCs, indicating that Notch1 is a functional downstream target of KLF9. In summary, our results strongly demonstrate that KLF9 is a crucial transcription factor that controls the osteogenic differentiation of DSCs by negatively regulating the Notch1 signaling pathway.

Galloylated proanthocyanidins in dentin matrix exhibit biocompatibility and induce differentiation in dental stem cells.

Grape seed extract contains a complex mixture of proanthocyanidins (PACs), a plant biopolymer used as a biomaterial to improve reparative and preventive dental therapies. Co-polymerization of PACs with type I collagen mechanically reinforces the dentin extracellular matrix. This study assessed the biocompatibility of PACs from grape seed extract on dental pulp stem cells (DPSCs) in a model simulating leaching through dentin to the pulp cavity. The aim was to determine the type of PACs (galloylated vs. non-galloylated) within grape seed extract that are most compatible with dental pulp tissue. Human demineralized dentin was treated with selectively-enriched dimeric PACs prepared from grape seed extract using liquid-liquid chromatography. DPSCs were cultured within a 2D matrix and exposed to PAC-treated dentin extracellular matrix. Cell proliferation was measured using the MTS assay and expression of odontoblastic genes was analyzed by qRT-PCR. Categorization of PACs leaching from dentin was performed using HPLC-MS. Enriched dimeric fractions containing galloylated PACs increased the expression of certain odontoblastic genes in DPSCs, including Runt-related transcription factor 2 (RUNX2), vascular endothelial growth factor (VEGF), bone morphogenetic protein 2 (BMP2), basic fibroblast growth factor (FGF2), dentin sialophosphoprotein (DSPP) and collagen, type I, alpha 1 (COLI). Galloylated dimeric PACs also exhibited minor effects on DPSC proliferation, resulting in a decrease compared to control after five days of treatment. The non-galloylated dimer fraction had no effect on these genes or on DPSC proliferation. Galloylated PACs are biocompatible with DPSCs and may exert a beneficial effect on cells within dental pulp tissue. The observed increase in odontoblastic genes induced by galloylated PACs together with a decrease in DPSC proliferation is suggestive of a shift toward cell differentiation. This data supports the use of dimeric PACs as a safe biomaterial, with galloylated dimeric PACs exhibiting potential benefits to odontoblasts supporting dentin regeneration.

Precision-engineered niche for directed differentiation of MSCs to lineage-restricted mineralized tissues

The major difference between tissue healing and regeneration is the extent of instructional cues available to precisely direct the biological response. A classic example is reparative or osteodentin that is seen in response to physicochemical injury to the pulp-dentin complex. Dentin regeneration can direct the differentiation of dental stem cells using concerted actions of both soluble (biomolecules, agonists, and antagonists) and insoluble (matrix topology) cues. The major purpose of this study was to examine the synergistic combination of two discrete biomaterial approaches by utilizing nanofiber scaffolds in discrete configurations (aligned or random) with incorporated polymeric microspheres capable of controlled release of growth factors. Further, to ensure appropriate disinfection for clinical use, Radio-Frequency Glow Discharge (RFGD) treatments were utilized, followed by seeding with a mesenchymal stem cell (MSC) line. SEM analysis revealed electrospinning generated controlled architectural features that significantly improved MSC adhesion and proliferation on the aligned nanofiber scaffolds compared to randomly oriented scaffolds. These responses were further enhanced by RFGD pre-treatments. These enhanced cell adhesion and proliferative responses could be attributed to matrix-induced Wnt signaling that was abrogated by pre-treatments with anti-Wnt3a neutralizing antibodies. Next, we incorporated controlled-release microspheres within these electrospun scaffolds with either TGF-β1 or BMP4. We observed that these scaffolds could selectively induce dentinogenic or osteogenic markers (DSPP, Runx2, and BSP) and mineralization. This work demonstrates the utility of a novel, modular combinatorial scaffold system capable of lineage-restricted differentiation into bone or dentin. Future validation of this scaffold system in vivo as a pulp capping agent represents an innovative dentin regenerative approach capable of preserving tooth pulp vitality.

Enhancement of Odontoblastic Differentiation of Stem Cells from Exfoliated Deciduous Tooth Using N-acetylcysteine—An In Vitro Study

The study was conducted to evaluate the effects of N-acetylcysteine (NAC) on the propagation and differentiation of stem cells from human exfoliated deciduous teeth(SHED). SHEDs were isolated by explant culture method and characterized for stem cell properties using flow cytometry method. MTT assay and Cell Counting Kit-8 (CCK-8) assay were used to examine the viability and proliferation of the SHEDs. The effects of NAC-induced osteo/odontoblastic differentiation of SHEDs were determined by functional staining for mineralization, and the gene expression of osteo/odontoblastic transcription factors and proteins was evaluated by real-time quantitative reverse transcription-polymerase chain reaction(qRT-PCR) analyses. Protein levels of collagen type 1 (COL1), dentin sialophosphoprotein (DSPP), and dentin matrix acidic phosphoprotein 1(DMP-1) were calculated by the Western blot method to assess the osteo/odontogenic differentiation. SHEDs presented mesenchymal stem cell (MSC)-like characteristics on flow cytometric analysis. The cell viability and metabolic activity of SHEDs were increased with an increase in the concentrations of NAC from 0.5 to 10 nM. However, the concentrations of NAC from 0.5 to 2.5 mM did not affect cell proliferation. NAC incorporated at a concentration of 2.5 mM showed higher mineralization and considerably increased gene expression levels of runt-related transcription factor 2 (RUNX2), COL1A1, DSPP, and DMP-1. It significantly increased the protein expression of odontoblast-related matrix proteins like COL1, DSPP, and DMP-1. NAC regulates the healthy propagation of dental stem cells in vitro. Its effects on the differentiation of dental pulp SHEDs remain unidentified. This study explores that NAC can encourage the mineralization of SHEDs and differentiate them into the odontoblastic lineage. The results propose that NAC could have a significant pharmacological role in activating and enhancing odontogenic differentiation of dental stem cells and possibly a prospect in regenerative dentistry.

Insulin-like growth factor 1 promotes neural differentiation of human stem cells from the apical papilla

ObjectiveInsulin-like growth factor 1 (IGF1) is one of the vital factors in regenerative endodontics. Previous studies have focused on the role of IGF1 in the mineralization of dental tissues. However, the role of IGF1 in the neural differentiation of dental stem cells was little discussed. DesignIGF1 was overexpressed in human stem cells from the apical papilla (hSCAPs) by lentivirus and knocked down in hSCAPs by small interfering RNA. The neural differentiation level of hSCAPs was investigated histologically by HE staining and Nissl staining after neural induction for 3 days. The expression of proteins was examined by western blot and immunofluorescence. ResultsIGF1 promoted neural differentiation of hSCAPs, more cell processes and Nissl-positive body stained cells. IGF1 overexpression could both promote glial differentiation in hSCAPs, characterized by the increase of S100β and GFAP proteins, and neuronal differentiation, characterized by the increase of βIII-tubulin and functional GAD67/vGLUT1 proteins. Conversely, IGF1 knockdown suppressed both glial and neuronal differentiation. IGF1 activated AKT to regulate the early neural differentiation of hSCAPs. ConclusionsThe results indicate IGF1 could promote neural differentiation of hSCAPs by activating AKT signaling and provide a cue for the candidate of induced neural seeding cells in regenerative endodontics.

Effects of iRoot SP on osteogenic differentiation of human stem cells from apical papilla

BackgroundResearch shows that nano-bioceramics can modulate the differentiation of dental stem cells. The novel ready-to-use calcium-silicate-based root-canal sealer iRoot SP is widely used in root filling. Accordingly, the aim of this study was to evaluate the effects of iRoot SP on proliferation and osteogenic differentiation in human stem cells from the apical papilla (hSCAPs).MethodshSCAPs were isolated and characterized in vitro, then cultured with various concentrations of iRoot SP extract. Cell proliferation was assessed by CCK-8 assay, and scratch-wound-healing assays were performed to evaluate cell-migration capacity. hSCAPs were then cultured in osteogenic medium supplemented with iRoot SP extracts. Alkaline phosphatase (ALP) activity assay was used to evaluate ALP enzyme levels. Alizarin red staining and cetylpyridinium chloride (CPC) assays were performed to assess calcified-nodule formation and matrix-calcium accumulation of hSCAPs. The mRNA and protein expression levels of the osteogenic markers OCN, OSX, Runx2, and DSPP were determined by qRT-PCR and Western blotting. The data were analyzed using one-way ANOVA and LSD-t tests.ResultsiRoot SP at low concentrations (2, 0.2, and 0.02 mg/mL) is nontoxic to hSCAPs. iRoot SP at concentrations of 0.02 and 0.2 mg/mL significantly increases cell-migration capacity. In terms of osteogenic differentiation, 0.2 mg/mL iRoot SP promotes intracellular ALP activity and the formation of mineralized nodules. Moreover, the expression of osteogenic markers at the mRNA and protein levels are upregulated by iRoot SP.ConclusioniRoot SP is an effective filling material for periapical bone regeneration.

Odontoblast-like Cytodifferentiation of Dental Stem Cells: A 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 in vitro and in vivo 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.

Micropatterned hydrogels and cell alignment enhance the odontogenic potential of stem cells from apical papilla in-vitro

IntroductionAn understanding of the extracellular matrix characteristics which stimulate and guide stem cell differentiation in the dental pulp is fundamental for the development of enhanced dental regenerative therapies. Our objectives, in this study, were to determine whether stem cells from the apical papilla (SCAP) responded to substrate stiffness, whether hydrogels providing micropatterned topographical cues stimulate SCAP self-alignment, and whether the resulting alignment could influence their differentiation towards an odontogenic lineage in-vitro. MethodsExperiments utilized gelatin methacryloyl (GelMA) hydrogels of increasing concentrations (5, 10 and 15%). We determined their compressive modulus via unconfined compression and analyzed cell spreading via F-actin/DAPI immunostaining. GelMA hydrogels were micropatterned using photolithography, in order to generate microgrooves and ridges of 60 and 120μm, onto which SCAP were seeded and analyzed for self-alignment via fluorescence microscopy. Lastly, we analyzed the odontogenic differentiation of SCAP using alkaline phosphatase protein expression (ANOVA/Tukey α=0.05). ResultsSCAP appeared to proliferate better on stiffer hydrogels. Both 60 and 120μm micropatterned hydrogels guided the self-alignment of SCAP with no significant difference between them. Similarly, both 60 and 120μm micropattern aligned cells promoted higher odontogenic differentiation than non-patterned controls. SignificanceIn summary, both substrate mechanics and geometry have a statistically significant influence on SCAP response, and may assist in the odontogenic differentiation of dental stem cells. These results may point toward the fabrication of cell-guiding scaffolds for regenerative endodontics, and may provide cues regarding the development of the pulp-dentin interface during tooth formation.

IGFBP5 promotes angiogenic and neurogenic differentiation potential of dental pulp stem cells.

Dental stem cells for dental pulp regeneration have become a new strategy for pulpitis treatment. Angiogenesis and neurogenesis play a vital role in the pulp-dentin complex regeneration, and appropriate growth factors will promote the process of angiogenesis and neurogenesis. Insulin-like growth factor-binding protein 5 (IGFBP5) is involved in the regulation of tooth growth and development. A previous study showed that IGFBP5 enhanced osteo/odontogenic differentiation of dental stem cells. Our research intends to reveal the function of IGFBP5 in the angiogenic and neurogenic differentiation of human dental stem cells. Human dental pulp stem cells (DPSCs) were used in the present study. Lentiviral IGFBP5 shRNA was used to silence the IGFBP5. Retroviruses expressing Wild-type IGFBP5 were used to over-express IGFBP5. Angiogenic and neurogenic differentiation were carried out by in vitro study. Real-time RT-PCR and western blot results showed that over-expression of IGFBP5 upregulated the expressions of angiogenic markers, including VEGF, PDGFA and ANG-1, and neurogenic markers, including NCAM, TH, Nestin, βIII-tubulin, and TH, in DPSCs. Moreover, microscope observation confirmed that over-expression of IGFBP5 enhanced neurosphere formation in DPSCs in size and amount. Immunofluorescence staining results showed that over-expression of IGFBP5 also prompted the percentage of Nestin and βIII-tubulin positive neurospheres in DPSCs. While depletion of IGFBP5 downregulated the expressions of VEGF, PDGFA, ANG-1, NCAM, TH, Nestin, βIII-tubulin, and TH, it decreased the neurosphere formation and percentage of Nestin and βIII-tubulin positive neurospheres in DPSCs. In conclusion, our results revealed that IGFBP5 promoted angiogenic and neurogenic differentiation potential of DPSCs in vitro and provided the possible potential target for enhancing directed differentiation of dental stem cells and dental pulp-dentin functional regeneration.

Potassium dihydrogen phosphate promotes the proliferation and differentiation of human periodontal ligament stem cells via nuclear factor kappa B pathway

IntroductionPeriodontal ligament stem cells (PDLSCs) are vital for the regeneration of periodontal tissues. Potassium dihydrogen phosphate (KH2PO4) has recently been applied as a component of the mineralization inducing medium (MM), which can be used to induce osteogenic differentiation of dental stem cells. However, whether KH2PO4 has effects on PDLSCs has not been studied. Materials and methodsPDLSCs were isolated by magnetic activated cell sorting and cultured. Alkaline phosphatase (ALP) activity and ALP protein expression of PDLSCs treated with different concentrations of KH2PO4 were examined to make sure the optimal concentration of KH2PO4 for the following experiments. The effects of KH2PO4 on the proliferation and differentiation of PDLSCs were investigated by flow cytometry, cell counting kit-8 assay, alizarin red staining, real-time RT-PCR, and Western blot. The involvement of nuclear factor kappa B (NF-κB) pathway in KH2PO4-treated PDLSCs was analyzed by Western blot and alizarin red staining. ResultsALP activity assay and ALP protein expression examination revealed that 1.8 mmol/L KH2PO4 was the optimal concentration for the induction of hPDLSCs by KH2PO4. The proliferation and mineralization capacity of PDLSCs treated with KH2PO4 were enhanced as compared with the control group. PDLSCs treated with KH2PO4 showed an improved proliferation capacity in logarithmic growth phase at day 7. As PDLSCs were treated with KH2PO4, the expression of odonto/osteogenic markers (OCN/OCN, DSP/DSPP, OSX/OSX, RUNX2/RUNX2, and ALP/ALP) in cells were up-regulated at day 3 or 7. Moreover, the expression of IκBα in cytoplasm was down-regulated, along with an increased expression of p-P65 in cytoplasm and an up-regulated expression of P65 in nucleus. When treated with BMS345541 (the specific NF-κB inhibitor), the odonto/osteogenic differentiation of KH2PO4-treated PDLSCs was significantly attenuated. ConclusionKH2PO4 can improve the proliferation and odonto/osteogenic differentiation capacity of PDLSCs via NF-κB pathway, and thus represents a potential target involved in the regeneration of periodontium for clinical treatments.

HPL promotes osteogenic differentiation of stem cells in 3D scaffolds.

Human platelet lysate (hPL) has been considered as the preferred supplement for the xeno-free stem cell culture for many years. However, the biological effect of hPL on the proliferation and differentiation of dental stem cells combined with the use of medical grade synthetic biomaterial is still under investigation. Thus, the optimal scaffold composition, cell type and specific growth conditions, yet need to be formulated. In this study, we aimed to investigate the regenerative potential of dental stem cells seeded on synthetic scaffolds and maintained in osteogenic media supplemented with either hPL or xeno-derived fetal bovine serum (FBS). Two types of dental stem cells were isolated from human impacted third molars and intact teeth; stem cells of apical papilla (SCAP) and periodontal ligament stem cells (PDLSCs). Cells were expanded in cell culture media supplemented with either hPL or FBS. Consequently, proliferative capacity, immunophenotypic characteristics and multilineage differentiation potential of the derived cells were evaluated on monolayer culture (2D) and on synthetic scaffolds fabricated from poly ’lactic-co-glycolic’ acid (PLGA) (3D). The functionality of the induced cells was examined by measuring the concentration of osteogenic markers ALP, OCN and OPN at different time points. Our results indicate that the isolated dental stem cells showed similar mesenchymal characteristics when cultured on hPL or FBS-containing culture media. Scanning electron microscopy (SEM) and H&E staining revealed the proper adherence of the derived cells on the 3D scaffold cultures. Moreover, the increase in the concentration of osteogenic markers proved that hPL was able to produce functional osteoblasts in both culture conditions (2D and 3D), in a way similar to FBS culture. These results reveal that hPL provides a suitable substitute to the animal-derived serum, for the growth and functionality of both SCAP and PDLSCs. Thus the use of hPL, in combination with PLGA scaffolds, can be useful in future clinical trials for dental regeneration.

Regulation of Wnt signaling pathway on calcium hydroxide-promoted osteogenic differentiation of hDPSCs

Objective To investigate the role of Wnt/β-catenin signaling pathway in the osteogenic differentiation of human dental pulp stem cells (hDPSCs). Methods After 14 days of the calcium hydroxide training, the cytoskeletal changes of hDPSCs, the expression of β-catenin, i.e. the key promoter of in the Wnt signaling pathway, and the cell localization were detected by laser scanning confocal technique. The Wnt signaling pathways were up-regulated and inhibited, and the osteogenic differentiation and mineralization of hDPSCs were detected by Western Blot and alizarin red staining after 14 days of the training. Results The cytoskeleton of hDPSCs was rearranged by the effect of calcium hydroxide, and the β-catenin migration from nucleus to cytoplasm were observed. The number of calcium nodules in hDPSCs was decreased after blocking Wnt signaling pathway by Dickkopf-related protein 1 (DKK-1). The calcium hydroxide treatment can promoted dentin sialophosphoprotein (DSPP) expression in hDPSCs. Conclusions Calcium hydroxide can down-regulate the expression of canonical Wnt signaling pathway and promote osteogenic differentiation and mineralization and odontogenetic differentiation of hDPSCs. Key words: Calcium hydroxide; Osteogenic differentiation; Human dental pulp stem; Wnt pathway

Anti-inflammatory effect of date seeds (Phoenix dactylifera L) on carrageenan-induced edema in rats

Purpose: To investigate the influence of sodium hypochlorite (NaOCl) on human dental stem cell proliferation and differentiation.Method: Dental pulp stem cells (DPSCs), periodontal ligament stem cell (PDLSCs), and gingival mesenchymal stem cells (GMSCs) were treated with NaOCl. Cell viability was evaluated with cellular counting kit-8 (CCK8), and cellular adenosine triphosphate (ATP) levels were analyzed by bromodeoxyuridine (BrdU) incorporation and subsequent flow cytometry. Quantitative polymerase chain reaction (qPCR) and western blotting were performed to detect the expressions of differentiation markers.Results: The viability and ATP levels of all three stem cells types were impaired by NaOCl in a concentration- and time-dependent manners. However, the decrease ATP in GMSCs was less than the other two stem cell population (p < 0.05). NaOCl treatment significantly suppressed the proliferation of dental stem cells (p < 0.05). With regard to differentiation marker expression levels, the decrease in Stro-1 was greater in treatment groups when compared to control on Day 7, while increase in levels of dentin sialophosphoprotein (DSPP), bone sialoprotein (BSP), and osteocalcin (OC) was smaller (p < 0.05). The expressional changes of Stro-1, DSPP, BSP, and OC were more prominent in DPSMs and PDLSCs than in GMSCs.Conclusion: NaOCl dose-dependently impairs the viability, proliferation and differentiation of dental stem cells. Thus, its toxicity to dental stem cells needs to be considered in clinical application.Keywords: Dental stem cells, Sodium hypochlorite, Viability, Proliferation, Differentiation

Evaluation of the cytotoxic effects of sodium hypochlorite on human dental stem cells

Purpose: To investigate the influence of sodium hypochlorite (NaOCl) on human dental stem cell proliferation and differentiation. Method: Dental pulp stem cells (DPSCs), periodontal ligament stem cell (PDLSCs), and gingival mesenchymal stem cells (GMSCs) were treated with NaOCl. Cell viability was evaluated with cellular counting kit-8 (CCK8), and cellular adenosine triphosphate (ATP) levels were analyzed by bromodeoxyuridine (BrdU) incorporation and subsequent flow cytometry. Quantitative polymerase chain reaction (qPCR) and western blotting were performed to detect the expressions of differentiation markers. Results: The viability and ATP levels of all three stem cells types were impaired by NaOCl in a concentration- and time-dependent manners. However, the decrease ATP in GMSCs was less than the other two stem cell population (p < 0.05). NaOCl treatment significantly suppressed the proliferation of dental stem cells (p < 0.05). With regard to differentiation marker expression levels, the decrease in Stro-1 was greater in treatment groups when compared to control on Day 7, while increase in levels of dentin sialophosphoprotein (DSPP), bone sialoprotein (BSP), and osteocalcin (OC) was smaller (p < 0.05). The expressional changes of Stro-1, DSPP, BSP, and OC were more prominent in DPSMs and PDLSCs than in GMSCs. Conclusion: NaOCl dose-dependently impairs the viability, proliferation and differentiation of dental stem cells. Thus, its toxicity to dental stem cells needs to be considered in clinical application. Keywords: Dental stem cells, Sodium hypochlorite, Viability, Proliferation, Differentiation

Sodium fluoride regulates the osteo/odontogenic differentiation of stem cells from apical papilla by modulating autophagy.

Fluoride (sodium fluoride) is thought to be essential in the development of tooth, and research shows that fluoride can modulate the differentiation of dental stem cells. However, the effects of fluoride on the committed differentiation of stem cells from apical papilla (SCAPs) and the underlying mechanisms remain unclear. Here, SCAPs were isolated from healthy extracted human third molars with immature roots and then were cultured with NaF conditioned media. Cell Counting Kit-8, EdU staining, and flow cytometry were performed to detected the proliferation activity. Alkaline phosphatase (ALP) activity, Alizarin Red staining, Western blot assay, and real-time reverse-transcription polymerase chain reaction were applied to assess the osteo/odontogenic differentiation NaF-treated SCAPs. Western blot assay and transmission electron microscope were used to evaluate the autophagy involved in the differentiation of SCAPs. ALP activity, ALP protein, and messenger RNA (mRNA) expression showed that 0.5 mM was the optimal concentration for the induction of SCAPs by NaF. 0.5 mM NaF-treated SCAPs induced more mineralized nodules as compared with untreated cells. Moreover, the osteo/odontogenic markers (RUNX2, OSX, DSP, and OCN) in mRNA levels were upregulated while the protein levels of these markers increased considerably in 0.5 mM NaF-treated SCAPs. Furthermore, the autophagy-related proteins (LC3, ATG5, and Beclin1) increased in NaF-treated SCAPs, and the osteo/odontogenic makers significantly decreased while silencing ATG5 to block autophagy. In all, sodium fluoride can regulate the osteo/odontogenic differentiation of SCAPs by modulating autophagy.

Hypothesis: A Challenge of Overexpression Zfp521 in Neural Tendency of Derived Dental Pulp Stem Cells.

Neurodegenerative diseases have now become a major challenge, especially in aged societies. Most of the traditional strategies used for treatment of these diseases are untargeted and have little efficiency. Developments in stem cell investigations have given much attention to cell therapy as an alternative concept in the regeneration of neural tissues. Dental pulp stem cells (DPSCs) can be readily obtained by noninvasive procedures and have been shown to possess properties similar to well-known mesenchymal stem cells. Furthermore, based on their neural crest origin, DPSCs are considered to have a good potential to differentiate into neural cells. Zfp521 is a transcription factor that regulates expression of many genes, including ones involved in the neural differentiation process. Therefor based on neural crest origin of the cell and high expression of neural progenitor markers, we speculate that sole overexpression of Zfp521 protein can facilitate differentiation of dental stem cells to neural cells and researchers may find these cells suitable for therapeutic treatment of neurodegenerative diseases.