Differentiation Of Dental Pulp Cells Research Articles

DLX6-AS1 regulates odonto/osteogenic differentiation in dental pulp cells under the control of BMP9 via the miR-128-3p/MAPK14 axis: A laboratory investigation.

The regenerative capacity of dental pulp relies on the odonto/osteogenic differentiation of dental pulp cells (DPCs), but dynamic microenvironmental changes hinder the process. Bone morphogenetic protein 9 (BMP9) promotes differentiation of DPCs towards an odonto/osteogenic lineage, forming dentinal-like tissue. However, the molecular mechanism underlying its action remains unclear. This study investigates the role of DLX6 antisense RNA 1 (DLX6-AS1) in odonto/osteogenic differentiation induced by BMP9. Custom RT2 profiler PCR array, quantitative Real-Time PCR (qRT-PCR) and western blots were used to investigate the expression pattern of DLX6-AS1 and its potential signal axis. Osteogenic ability was evaluated using alkaline phosphatase and alizarin red S staining. Interactions between lncRNA and miRNA, as well as miRNA and mRNA, were predicted through bioinformatic assays, which were subsequently validated via RNA immunoprecipitation and dual luciferase reporter assays. Student's t-test or one-way ANOVA with post hoc Tukey HSD tests were employed for data analysis, with a p-value of less than .05 considered statistically significant. DLX6-AS1 was upregulated upon BMP9 overexpression in DPCs, thereby promoting odonto/osteogenic differentiation. Additionally, miR-128-3p participated in BMP9-induced odonto/osteogenic differentiation by interacting with the downstream signal MAPK14. Modifying the expression of miR-128-3p and transfecting pcMAPK14/siMAPK14 had a rescue impact on odonto/osteogenic differentiation downstream of DLX6-AS1. Lastly, miR-128-3p directly interacted with both MAPK14 and DLX6-AS1. DLX6-AS1 could regulate the odonto/osteogenic differentiation of DPCs under the control of BMP9 through the miR-128-3p/MAPK14 axis.

DLX6-AS1 regulates odonto/osteogenic differentiation in dental pulp cells under the control of BMP9 via the miR-128-3p/MAPK14 axis: A laboratory investigation.

The regenerative capacity of dental pulp relies on the odonto/osteogenic differentiation of dental pulp cells (DPCs), but dynamic microenvironmental changes hinder the process. Bone morphogenetic protein 9 (BMP9) promotes differentiation of DPCs towards an odonto/osteogenic lineage, forming dentinal-like tissue. However, the molecular mechanism underlying its action remains unclear. This study investigates the role of DLX6 antisense RNA 1 (DLX6-AS1) in odonto/osteogenic differentiation induced by BMP9. Custom RT2 profiler PCR array, quantitative Real-Time PCR (qRT-PCR) and western blots were used to investigate the expression pattern of DLX6-AS1 and its potential signal axis. Osteogenic ability was evaluated using alkaline phosphatase and alizarin red S staining. Interactions between lncRNA and miRNA, as well as miRNA and mRNA, were predicted through bioinformatic assays, which were subsequently validated via RNA immunoprecipitation and dual luciferase reporter assays. Student's t-test or one-way ANOVA with post hoc Tukey HSD tests were employed for data analysis, with a p-value of less than .05 considered statistically significant. DLX6-AS1 was upregulated upon BMP9 overexpression in DPCs, thereby promoting odonto/osteogenic differentiation. Additionally, miR-128-3p participated in BMP9-induced odonto/osteogenic differentiation by interacting with the downstream signal MAPK14. Modifying the expression of miR-128-3p and transfecting pcMAPK14/siMAPK14 had a rescue impact on odonto/osteogenic differentiation downstream of DLX6-AS1. Lastly, miR-128-3p directly interacted with both MAPK14 and DLX6-AS1. DLX6-AS1 could regulate the odonto/osteogenic differentiation of DPCs under the control of BMP9 through the miR-128-3p/MAPK14 axis.

Anti-inflammatory cerium-containing nano-scaled mesoporous bioactive glass for promoting regenerative capability of dental pulp cells.

This study aimed to investigate the anti-inflammatory and odontoblastic effects of cerium-containing mesoporous bioactive glass nanoparticles (Ce-MBGNs) on dental pulp cells as novel pulp-capping agents. Ce-MBGNs were synthesized using a post-impregnation strategy based on the antioxidant properties of Ce ions and proposed the first use of Ce-MBGNs for pulp-capping application. The biocompatibility of Ce-MBGNs was analysed using the CCK-8 assay and apoptosis detection. Additionally, the reactive oxygen species (ROS) scavenging ability of Ce-MBGNs was measured using the 2,7-Dichlorofuorescin Diacetate (DCFH-DA) probe. The anti-inflammatory effect of Ce-MBGNs on THP-1 cells was further investigated using flow cytometry and quantitative real-time polymerase chain reaction (RT-qPCR). Moreover, the effect of Ce-MBGNs on the odontoblastic differentiation of the dental pulp cells (DPCs) was assessed by combined scratch assays, RT-qPCR, western blotting, immunocytochemistry, Alizarin Red S staining and tissue-nonspecific alkaline phosphatase staining. Analytically, the secretions of tumour necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) were detected with enzyme-linked immunosorbent assay (ELISA). Ce-MBGNs were confirmed to effectively scavenge ROS in THP-1-derived macrophages and DPCs. Flow cytometry and RT-qPCR assays revealed that Ce-MBGNs significantly inhibited the M1 polarization of macrophages (Mφ). Furthermore, the protein levels of TNF-α and IL-1β were downregulated in THP-1-derived macrophages after stimulation with Ce-MBGNs. With a step-forward virtue of promoting the odontoblastic differentiation of DPCs, we further confirmed that Ce-MBGNs could regulate the formation of a conductive immune microenvironment with respect to tissue repair in DPCs, which was mediated by macrophages. Ce-MBGNs protected cells from self-produced oxidative damage and exhibited excellent immunomodulatory and odontoblastic differentiation effects on DPCs. As a pulp-capping agent, this novel biomaterial can exert anti-inflammatory effects and promote restorative dentine regeneration in clinical treatment. We believe that this study will stimulate further correlative research on the development of advanced pulp-capping agents.

LncRNA CARMN facilitates odontogenic differentiation of dental pulp cells by impairing EZH2.

This study aimed to reveal the potential role of CARMN in odontogenic differentiation of dental pulp cells (DPCs). Laser capture microdissection was used to detect Carmn in DPCs and odontoblasts in P0 mice. After manipulating CARMN expression in odontogenic differentiation induced hDPCs, the state of odontogenic differentiation was evaluated by ALP staining, ARS, and related marker expression in qRT-PCR and western blotting. The subcutaneous transplantation of HA/β-TCP loaded with hDPCs was performed to verify CARMN's role in promoting odontogenic differentiation in vivo. RNAplex and RIP were employed to reveal potential mechanism of CARMN in hDPCs. CARMN expressed more abundantly in odontoblasts than DPCs in P0 mice. CARMN expression boosted during in vitro odontogenic differentiation of hDPCs. CARMN overexpression enhanced odontogenic differentiation of hDPCs in vitro, while inhibition impaired the process. CARMN overexpression in HA/β-TCP composites promoted more mineralized nodule formation in vivo. CARMN knockdown led to soared EZH2, while CARMN overexpression brought about EZH2 inhibition. CARMN functioned via direct interaction with EZH2. The results uncovered CARMN as a modulator during the odontogenic differentiation of DPCs. CARMN promoted odontogenic differentiation of DPCs by impairing EZH2.

Mitochondrial reactive oxygen species promote mitochondrial damage in high glucose-induced dysfunction and apoptosis of human dental pulp cells

Background/purposeHigh 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 methodsHuman 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. ResultsHG 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. ConclusionHG triggers mitochondrial damage via augmenting mtROS generation, resulting in the inhibited proliferation, migration, and odontoblastic differentiation of hDPCs and enhanced their apoptosis.

Concentration-Dependent Cellular Uptake of Graphene Oxide Quantum Dots Promotes the Odontoblastic Differentiation of Dental Pulp Cells via the AMPK/mTOR Pathway.

As zero-dimension nanoparticles, graphene oxide quantum dots (GOQDs) have broad potential for regulating cell proliferation and differentiation. However, such regulation of dental pulp cells (DPSCs) with different concentrations of GOQDs is insufficiently investigated, especially on the molecular mechanism. The purpose of this study was to explore the effect and molecular mechanism of GOQDs on the odontoblastic differentiation of DPSCs and to provide a theoretical basis for the repair of pulp vitality by pulp capping. CCK-8, immunofluorescence staining, alkaline phosphatase activity assay and staining, alizarin red staining, qRT-PCR, and western blotting were used to detect the proliferation and odontoblastic differentiation of DPSC coculturing with different concentrations of GOQDs. The results indicate that the cellular uptake of low concentration of GOQDs (0.1, 1, and 10 μg/mL) could promote the proliferation and odontoblastic differentiation of DPCSs. Compared with other concentration groups, 1 μg/mL GOQDs show better ability in such promotion. In addition, with the activation of the AMPK signaling pathway, the mTOR signaling pathway was inhibited in DPSCs after coculturing with GOQDs, which indicates that low concentrations of GOQDs could regulate the odontoblastic differentiation of DPSCs by the AMPK/mTOR signaling pathway.

The Wisdom in Teeth: Neuronal Differentiation of Dental Pulp Cells.

Mesenchymal stem/stromal cells (MSCs) are found in almost all postnatal organs. Under appropriate environmental cues, multipotency enables MSCs to serve as progenitors for several lineage-specific, differentiated cell types. In vitro expansion and differentiation of MSCs give the opportunity to obtain hardly available somatic cells, such as neurons. The neurogenic potential of MSCs makes them a promising, autologous source to restore damaged tissue and as such, they have received much attention in the field of regenerative medicine. Several stem cell pool candidates have been studied thus far, but only a few of them showed neurogenic differentiation potential. Due to their embryonic ontology, stem cells residing in the stroma of the dental pulp chamber are an exciting source for in vitro neural cell differentiation. In this study, we review the key properties of dental pulp stem cells (DPSCs), with a particular focus on their neurogenic potential. Moreover, we summarize the various presently available methods used for neural differentiation of human DPSCs also emphasizing the difficulties in reproducibly high production of such cells. We postulate that because DPSCs are stem cells with very close ontology to neurogenic lineages, they may serve as excellent targets for neuronal differentiation in vitro and even for direct reprogramming.

Effects of inflammation in dental pulp cell differentiation and reparative response

The responsiveness of the dentin-pulp complex is possible due to the stimulation of dental pulp cells, which begin to synthesize and secrete dentin matrix. The inflammatory process generated by harmful stimuli should be understood as a natural event of the immune response, resulting in the recruitment of hematopoietic cells, which cross the endothelial barrier and reach the site affected by the injury in order to eliminate the damage and provide an appropriate environment for the restoration of homeostasis. The repair process occurs in the presence of adequate blood supply, absence of infection, and with the participation of pro-inflammatory cytokines, growth factors, extracellular matrix components, and other biologically active molecules. Prostaglandins and leukotrienes are bioactive molecules derived from the metabolism of arachidonic acid, as a result of a variable range of cellular stimuli. The aim of this review is to describe the process of formation and biomineralization of the dentin-pulp complex and how pro-inflammatory events can modify this response, with emphasis on the lipid mediators prostaglandins and leukotrienes derived from arachidonic acid metabolism.

Calcium Phosphate Cement Promotes Odontoblastic Differentiation of Dental Pulp Cells In Vitro and In Vivo

In the case of pulp injury, odontoblastic differentiation of dental pulp cells (DPCs) at the site of the exposed pulp is necessary for a successful direct pulp capping treatment. Calcium phosphate cement (CPC), a kind of hydroxyapatite-like bone cement, exhibits therapeutic potential in osteogenesis by regulating cell cycle progression and promoting osteoblastic differentiation. Based on the similar biological process of osteo/odontoblastic differentiation, the present study evaluated the effects of CPC on odontoblastic differentiation of DPCs in vitro and in vivo, respectively. The morphology of CPC was observed by scanning electron microscopy. Colony-forming units were used to assess the antibacterial activity. The effects of CPC on cell proliferation and odontoblastic differentiation of human dental pulp cells (hDPCs) were also measured. Histological staining was performed to observe the reparative dentin formation in rat molars. In vitro, results of antibacterial studies showed that CPC significantly inhibited the growth of Streptococcus mutans. The appropriate concentration of CPC extract showed low cytotoxicity on hDPCs. Furthermore, CPC extract also promoted odontoblastic differentiation and mineralization compared with the control group, as shown by a dynamic increase in the expression of odontogenic marker genes and the increased number of mineralized nodules at 21 days. The pulpotomy models with CPC facilitated the formation of dentin bridge with the highly expressed dentin matrix protein 1 (DMP1) in odontoblast-like cells. In conclusion, the favorable biocompatibility, antibacterial property and bio-inductivity of CPC suggest that CPC can be used as a promising direct pulp capping material.

Leukotriene B4 loaded in microspheres regulate the expression of genes related to odontoblastic differentiation and biomineralization by dental pulp stem cells

BackgroundLeukotriene B4 (LTB4) is a potent lipid mediator that stimulate the immune response. Because dental pulp inflammation and dentin repair are intrinsically related responses, the aim of this research was to investigate the potential of LTB4 in inducing differentiation of dental pulp stem cells.MethodsMicrospheres (MS) loaded with LTB4 were prepared using an oil emulsion solvent extraction evaporation process and sterility, characterization, efficiency of LTB4 encapsulation and in vitro LTB4 release assay were investigated. Mouse dental pulp stem cells (OD-21) were stimulated with soluble LTB4 or MS loaded with LTB4 (0.01 and 0.1 μM). Cytotoxicity and cell viability was determined by lactate dehydrogenase and methylthiazol tetrazolium assays. Gene expression were measured by quantitative reverse transcription polymerase chain reaction after 3, 6, 24, 48 and 72 h. Mineralized nodule formation was assessed after 28 days of OD-21 cell stimulation with LTB4 in mineralized media or not. Groups were compared using one-way ANOVA test followed by Dunnett’s post-test (α = 0.05).ResultsTreatment with LTB4 or MS loaded with LTB4 (0.01 and 0.1 µm-μM) were not cytotoxic to OD-21 cells. Treatment with LTB4 modulated the expression of the Ibsp (integrin binding sialoprotein) and Runx2 (runt-related transcription factor 2) genes differently depending on the experimental period analyzed. Interestingly LTB4 loaded in microspheres (0.1 μM) allowed long term dental pulp cell differentiation and biomineralization.ConclusionLTB4, soluble or loaded in MS, were not cytotoxic and modulated the expression of the Ibsp and Runx2 genes in cultured OD-21 cells. When LTB4 was incorporated into MS, odontoblast differentiation and mineralization was induced in long term culture.

Effect of mesoporous bioactive glass on odontogenic differentiation of human dental pulp stem cells.

Healthy pulp tissue plays an important role in normal function and long-term retention of teeth. Mesoporous bioactive glass (MBG) as a kind of regenerative biomaterials shows the potential in preserving the vital pulp. In this study, MBG prepared by organic template method combined with sol-gel method were used in human dental pulp cell culture and ectopic mineralization experiment. Real-Time PCR was used to explore its ability to induce odontogenic differentiation of dental pulp cells. MBG and rat crowns were implanted under the skin of nude mice for 4 weeks to observe the formation of pulp dentin complex. We found that MBG can release Si and Ca ions and has a strong mineralization activity in vitro. The co-culture of MBG with human dental pulp cells promoted the expression of DMP-1 (dentin matrix protein-1) and ALP (alkalinephosphatase) without affecting cell proliferation. After 4 weeks of subcutaneous implantation in nude mice, the formation of hard tissue with regular structure under the material could be seen, and the structure was similar to dentin tubules. These results indicate that MBG can induce the differentiation of dental pulp cells and the formation of dental pulp-dentin complex and has the potential to promote the repair and regeneration of dental pulp injuries.

Endoplasmic reticulum stress response mediated by the PERK-eIF2α-ATF4 pathway is involved in odontoblastic differentiation of human dental pulp cells

ObjectiveRNA-activated protein kinase-like ER-resident kinase (PERK) was a major transducer of Endoplasmic reticulum (ER) stress response and it directly phosphorylated α-subunit of eukaryotic initiation factor 2 (eIF2α), which specifically promoted the translation of activating transcription factor 4 (ATF4), an important transcription factor in cells’ differentiation. The purpose of this study was to establish whether ER stress mediated by PERK-eIF2α-ATF4 pathway was involved in odontoblastic differentiation of human dental pulp cells (DPCs). MethodsDPCs were isolated from extracted teeth and cultured in odontogenic medium. A recombinant lentiviral vector was constructed to transfect DPCs for PERK knockdown. Alkaline phosphatase (ALP) and Alizarin red S staining were used to characterize the odontoblastic differentiation. Real-time polymerase chain reactions (RT-PCR) were performed to analyze the genes’ expressions in DPCs’ odontoblastic differentiation. The mRNA and protein levels of ER stress markers were examined by RT-PCR and western blot. ResultsDPCs cultured in odontogenic media showed increased ALP activity and mineralized nodule formation. Notably, treatment with differentiation medium resulted in the up-regulation of genes, such as osteocalcin (OCN), bone sialoprotein (BSP), dentin sialophosphoprotein (DSPP), splicing x-box binding protein-1 (sXBP1), ATF4 and glucose-regulated protein 78 (GRP78). Meanwhile, the expressions of PERK-eIF2α-ATF4 pathway proteins, phosphorylated PERK, phosphorylated eIF2α and ATF4, increased in odontoblastic induction cells compared with controls. Furthermore, inhibition of PERK (PERK knockdown) decreased ALP activity and matrix mineralization in DPCs accompanied by the decrease expression of phosphorylated eIF2α and ATF4. ConclusionThese results suggested that PERK-eIF2α-ATF4 pathway was involved in the odontoblastic differentiation of DPCs.

Photobiomodulation therapy does not depend on the differentiation of dental pulp cells to enhance functional activity associated with angiogenesis and mineralization.

The purpose of this study is to analyze the influence of InGaAlP diode laser (660nm) with or without an odontogenic medium (OM) in the functional activity of OD-21 cells. Undifferentiated OD-21 pulp cells were cultivated with or without OM and divided into four groups (n = 5): nonirradiated control (C -), nonirradiated + OM (C +), irradiated (L -), and irradiated + OM (L +). Laser application was performed in two sessions of a 24-h interval with an irradiance of 11.3 mW/cm2, energy density of 1J/cm2, and total cumulative energy/well of 4.6J. Cell proliferation, VEGF-164 expression, mineralization, and expression of Alp, Runx2, and Dmp1 genes, as well as immunolocalization of RUNX2 and MEPE proteins, were evaluated. Data were analyzed by statistical tests (α = 0.05). All studied groups showed a similar increase in cell proliferation with or without OM. After 7 and 10days, a significatively higher concentration of VEGF-164 in L - group when compared to C - group was observed. A significant increase in mineralized nodules in the L + was noted when compared to C + in the same conditions. Photobiomodulation upregulated significantly Runx2 and Dmp1 expression after 10days in L - and after 7days in L + , with downregulation of Dmp1 after 10days in L + group. Immunolocalization of RUNX2 and MEPE was expressive after 7days of culture in the cytoplasm adjacent to the nucleus with a decrease after 10days, regardless of the presence of OM. Photobiomodulation enhances metabolism associated with angiogenesis, gene expression, and mineralization regardless of the odontogenic medium in OD-21 cells.

Chitosan-Calcium-Simvastatin Scaffold as an Inductive Cell-Free Platform

The development of biomaterials based on the combination of biopolymers with bioactive compounds to develop delivery systems capable of modulating dentin regeneration mediated by resident cells is the goal of current biology-based strategies for regenerative dentistry. In this article, the bioactive potential of a simvastatin (SV)–releasing chitosan-calcium-hydroxide (CH-Ca) scaffold was assessed. After the incorporation of SV into CH-Ca, characterization of the scaffold was performed. Dental pulp cells (DPCs) were seeded onto scaffolds for the assessment of cytocompatibility, and odontoblastic differentiation was evaluated in a microenvironment surrounded by dentin. Thereafter, the cell-free scaffold was adapted to dentin discs positioned in artificial pulp chambers in direct contact with a 3-dimensional (3D) culture of DPCs, and the system was sealed to simulate internal pressure at 20 cm/H2O. In vivo experiments with cell-free scaffolds were performed in rats’ calvaria defects. Fourier-transform infrared spectroscopy spectra proved incorporation of Ca and SV into the scaffold structure. Ca and SV were released upon immersion in a neutral environment. Viable DPCs were able to spread and proliferate on the scaffold over 14 d. Odontoblastic differentiation occurred in the DPC/scaffold constructs in contact with dentin, in which SV supplementation promoted odontoblastic marker overexpression and enhanced mineralized matrix deposition. The chemoattractant potential of the CH-Ca scaffold was improved by SV, with numerous viable and dentin sialoprotein–positive cells from the 3D culture being observed on its surface. Cells at 3D culture featured increased gene expression of odontoblastic markers in contact with the SV-enriched CH-Ca scaffold. CH-Ca-SV led to intense mineralization in vivo, presenting mineralization foci inside its structure. In conclusion, the CH-Ca-SV scaffold induces differentiation of DPCs into a highly mineralizing phenotype in the presence of dentin, creating a microenvironment capable of attracting pulp cells to its surface and inducing the overexpression of odontoblastic markers in a cell-homing strategy.

Inactivation of PI3K/Akt promotes the odontoblastic differentiation and suppresses the stemness with autophagic flux in dental pulp cells

Background/purposeAutophagy is involved in controlling differentiation of various cell types. The present study aimed to investigate the mechanism related to autophagy in regulating odontogenic differentiation of dental pulp cells. Materials and methodsHuman dental pulp cells (HDPCs) were cultured in differentiation inductive medium (DM) and odontoblastic differentiation and mineralization were evaluated by alkaline phosphatase (ALP) staining and Alizarin red S staining, respectively. Tooth cavity preparation was made on the mesial surface of lower first molars in rat. The expression of autophagy-related signal molecules was detected using Western blot analysis and Immunohistochemistry. ResultsHDPCs cultured in DM showed increased autophagic flux and declined phosphorylation of phosphoinositide 3-kinases (PI3K), protein kinase B (Akt), and mTOR. Dentin matrix protein-1 (DMP-1) and dentin sialoprotein (DSP), markers of odontoblastic differentiation, were upregulated and autophagic activation showing increased LC3-II and decreased p62 levels was observed during odontogenic differentiation of HDPCs. However, PI3K blocker 3-methyladenine (3MA), lentiviral shLC3 and Akt activator SC79 attenuated the expression of LC3II as well as DMP-1, ALP activity and mineralization enhanced in HDPCs under DM condition. In addition, 3MA, shLC3 and SC79 recovered the expression of pluripotency factor CD146, Oct4 and Nanog downregulated in DM condition. In rat tooth cavity preparation model, the expression of LC3B and DMP-1 was elevated near odontoblast-dentin layer during reparative dentin formation, whereas 3MA significantly reduced the expression of LC3B and DMP-1. ConclusionThese findings indicated autophagy promotes the odontogenic differentiation of dental pulp cells modulating stemness via PI3K/Akt inactivation and the repair of pulp.

Hybrid porous zirconia scaffolds fabricated using additive manufacturing for bone tissue engineering applications

For the formation of new bone in critical-sized bone defects, bioactive scaffolds with an interconnected porous network are necessary. Herein, we fabricated three-dimensional (3D) porous hybrid zirconia scaffolds to promote hybrid functionality, i.e., excellent mechanical properties and bioactive performance. Specifically, the 3D printed scaffolds were subjected to Zn-HA/glass composite coating on glass-infiltrated zirconia (ZC). In addition, to pertain the extracellular matrix of bone, biopolymer (alginate/gelatine) was embedded in a developed 3D construct (ZB and ZCB). A zirconia-printed scaffold (Z) group served as a control. The structural and mechanical properties of the constructed scaffolds were studied using essential characterization techniques. Furthermore, the biological performance of the designed scaffolds was tested by a sequence of in vitro cell tests, including the attachment, proliferation, and osteogenic differentiation of dental pulp cells (DPCs). The ZC and ZCB scaffolds exhibited 20% higher compression strength than the zirconia (Z) scaffolds. More importantly, the ZC constructs exhibited superior cell-adhesion, distribution, and osteogenic differentiation ability due to the synergistic effects of the composite coating. In addition, the biopolymer-embedded scaffolds (ZB, ZCB) showed an excellent biological and mechanical performance. Thus, our results suggest that the Zn-HA/glass composite-coated glass-infiltrated zirconia (ZC, ZCB) scaffolds are a dynamic approach to designing bioactive 3D scaffolds for the load-bearing bone regeneration applications.

The histone deacetylase inhibitor, entinostat (MS-275), induces the odontogenic differentiation of an odontoblast-like cell line in the absence of an osteoblast mineralization medium.

The aim of this study was to determine whether histone deacetylase inhibitors (HDACi), including entinostat (MS-275), valproic acid (VPA), trichostatin A (TSA), and sodium butyrate (NaB), promoted the odontogenic differentiation of the odontoblast-like cell line, MDPC-23 in the absence of an osteoblast mineralization medium. The cells were cultured in basal medium (Dulbecco's modified Eagle medium) with and without (controls) the inhibitors. The cell viability and migration were assessed using the cell proliferation reagent WST-1 and a scratch wound healing assay, respectively. The mRNA expression levels of bone morphogenetic protein (Bmp)-2 and -4, collagen 1 alpha 1 (Col1α1), osteocalcin (Oc), dentin matrix protein1 (Dmp1), dentin sialophosphoprotein (Dspp), runt-related transcription factor 2 (Runx2), Krueppel-like factor 5 (Klf5), and Msh homeobox 1(Msx1) were evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). Alizarin red and alkaline phosphatase assays were performed to determine the extent of mineralization in the culture systems. No significant differences in cell numbers were observed between the controls and the MS-275-, VPA-, and NaB-treated cells; however, a significant difference was observed with TSA (concentration, 1000nM). The scratch wound healing assay showed no effect of cell migration in the MS-275 (1.0µM)-treated cells when compared with the controls at 24h. Furthermore, MS-275, VPA, and NaB increased the mRNA expression levels of Bmp-2 and -4, Oc, and Runx2 followed by the mineralization of the cells. Only MS-275 significantly increased the expression levels of Dmp1, Dspp, Klf5, and Msx1 in the cells. These findings indicated that MS-275 may be considered as a reliable candidate for the odontogenic differentiation of dental pulp cells.

SHED, PRF, and Chitosan as Three-Dimensional of Tissue Engineering for Dental Pulp Regeneration

The gold standard for pulpitis irreversible treatment is root canal treatment. However, it caused the loss of tooth vitality. To restore tooth vitality, materials that have regenerative ability in the pulp is needed. The exfoliated deciduous teeth stem cells (SHED) not only expressed specific markers for mesenchymal stem cells (MSC) but also induced the odontoblastic differentiation, and stimulating the formation of endothelium and fibroblast. The combination SHED with platelet rich fibrin (PRF) and chitosan were able to facilitate and increase the migration, proliferation and odontoblastic differentiation of dental pulp cells. Based on that fact, the combination of SHED, PRF and chitosan as three-dimensional tissue engineering is promising as new modality to pulp regeneration in the clinical setting. The purpose of this review is describing the potential combination of SHED, PRF, and chitosan scaffold as three-dimensional tissue engineering for pulp regeneration.

Drug Repurposing for Tooth Regeneration: The Promising Premises.

Drug repurposing which identifies new therapeutic use(s) for drugs currently in use is a brand-new avenue of research interest worldwide. It circumvents the high-end monetary and time investment usually associated with contemporary drug discoveries. In the field of dentistry, recent studies in drug repurposing focuses in attaining dentin repair or reduction of bone resorption associated with apical periodontitis. Metformin, an anti-diabetic drug has shown pro-osteogenic properties. Aspirin a known anti-inflammatory agent with anticoagulant action is found to modulate the differentiation of dental pulp cells. The significant role of glycogen synthase kinase-3 inhibitors in activating the Wnt/-beta cat signaling pathway of mesenchymal pulp stem cells may pave the way to the pharmacological treatment of dental caries in near future. It is to be noted here that further preclinical and clinical studies are warranted for the regular therapeutic use of these potential drugs in clinical dentistry.

EDTA Promotes the Mineralization of Dental Pulp In Vitro and In Vivo

IntroductionDentin regeneration is one of the main goals of vital pulp treatment in which the biological properties of dental pulp cells (DPCs) need to be considered. In our previous study, we showed that EDTA could enhance the stromal cell–derived factor 1 alpha–induced migration of DPCs. The purpose of this study was to explore the effects of EDTA on the mineralization of dental pulp in vitro and in vivo. MethodsDPCs were obtained from human premolars or third molars. Alkaline phosphatase assays and alizarin red S staining were used to examine the degree of differentiation and mineralized nodule formation of DPCs. Real-time polymerase chain reaction and Western blot analysis were performed to detect the messenger RNA and protein expressions of mineralization-related markers in DPCs. Extracellular-regulated protein kinase and Smad inhibitors were used to study the roles of these 2 signaling pathways in this process. In addition, pulp exposures were created on 18 premolars of 2 beagle dogs (>12 months) using a high-speed dental handpiece. The experimental group (n = 9) was treated with 12% EDTA for 5 minutes, and the control group (n = 9) was treated with sterile saline for the same duration. Mineral trioxide aggregate was used for direct pulp capping followed by glass ionomer cement sealing. Samples were collected 3 months later, and the regenerated dentin was assessed by micro–computed tomographic and histologic analyses. ResultsExposure to 12% EDTA promoted the activity of alkaline phosphatase, the formation of mineralized nodules, and the messenger RNA and protein expressions of mineralization-related markers in DPCs. Furthermore, the process of 12% EDTA enhancing the differentiation of DPCs was mediated by the extracellular-regulated protein kinase 1/2 signaling pathway and inhibited by the Smad2/3 signaling pathway. In vivo, compared with the control group, more regenerated dentin that had fewer tunnel defects was formed in the 12% EDTA-treated group. ConclusionsOur results showed that 12% EDTA could promote the mineralization of dental pulp in vitro and in vivo.