Odontogenic Differentiation Of Dental Pulp Stem Cells Research Articles

The Role of ORAI1 in the Odontogenic Differentiation of Human Dental Pulp Stem Cells

Pulp capping, or placing dental materials directly onto the vital pulp tissues of affected teeth, is a dental procedure that aims to regenerate reparative dentin. Several pulp capping materials are clinically being used, and calcium ion (Ca2+) released from these materials is known to mediate reparative dentin formation. ORAI1 is an essential pore subunit of store-operated Ca2+ entry (SOCE), which is a major Ca2+ influx pathway in most nonexcitable cells. Here, we evaluated the role of ORAI1 in mediating the odontogenic differentiation and mineralization of dental pulp stem cells (DPSCs). During the odontogenic differentiation of DPSCs, the expression of ORAI1 increased in a time-dependent manner. DPSCs knocked down with ORAI1 shRNA (DPSC/ORAI1sh) or overexpressed with dominant negative mutant ORAI1E106Q (DPSC/E106Q) exhibited the inhibition of Ca2+ influx and suppression of odontogenic differentiation and mineralization as demonstrated by alkaline phosphatase (ALP) activity/staining as well as alizarin red S staining when compared with DPSCs of their respective control groups (DPSC/CTLsh and DPSC/CTL). The gene expression for odontogenic differentiation markers such as osteocalcin, bone sialoprotein, and dentin matrix protein 1 (DMP1) was also suppressed. When DPSC/CTL or DPSC/E106Q cells were subcutaneously transplanted into nude mice, DPSC/CTL cells induced mineralized tissue formation with significant increases in ALP and DMP1 staining in vivo, whereas DPSC/E106Q cells did not. Collectively, our data showed that ORAI1 plays critical roles in the odontogenic differentiation and mineralization of DPSCs by regulating Ca2+ influx and that ORAI1 may be a therapeutic target to enhance reparative dentin formation.

Effects of adrenomedullin on proliferation and differentiation of dental pulp stem cells

Objective To explore the effects of adrenomedullin( ADM) on the proliferation and odontogenic differentiation potential of dental pulp stem cells( DPSCs). Methods The 3rdgeneration of cultured human DPSCs were treated with ADM of different concentrations( 10- 6mol / L,10- 7mol / L and 10- 8mol / L) for 24 h,and a blank control group was also set up. Cell apoptosis and cell cycle were measured by flow cytometry; the mRNA and protein expressions of alkaline phosphatase( ALP),an odontogenic differentiation marker,were analyzed by qPCR and Western blotting analysis,respectively. Results Cell cycle analysis showed that the proportion of G2/ S / M phase cells in the three ADM treatment groups were significantly higher than that in the blank control group( P 0. 05),but there were no significant differences between the three ADM groups. Cell apoptosis analysis showed that the proportion of cells labelled by Annexin Ⅴ+/ PI-was significantly lower in the three ADM groups than that in the blank control group( P 0. 05), but there were no signficant differences between the three ADM groups. The results of qPCR and Western blotting analysis showed that the ALP expression in DPSCs was increased significantly compared with that in the blank control group( P 0. 01),but there were no significant differences between the three ADM groups. Conclusion ADM can promote the proliferation and inhibit the apoptosis of DPSCs,and it can also promote odontogenic differentiation of DPSCs.

Odontogenic Differentiation of Dental Pulp Derived Stem Cells on Tricalcium Phosphate Scaffolds

Abstract Background/purpose The regeneration of dental-related tissue is a major problem in dentistry. Thus, it is beneficial to develop dental constructs that are fabricated with dental pulp stem cells (DPSCs) and an appropriate scaffold. The present study investigates the level of odontogenic differentiation of human DPSCs on tricalcium phosphate (TCP) scaffolds. Materials and methods We isolated pulp stem cells from human third molars and culture-expanded them through several successive subcultures. The cells from passage 3 were then loaded onto TCP scaffolds and treated with odontogenic supplements (OSs) that included vitamin D3 for a period of 21 days. DPSCs cultivated on TCP without OS, a monolayer culture treated with OS, and normal pulp tissue were the controls. We compared the groups in terms of odontogenic differentiation markers. Results Alkaline phosphatase (ALP) activity and the amount of culture mineralization, as well as the expression levels of dentin sialophosphoprotein ( DSPP ) and dentin matrix acidic phosphoprotein 1 ( DMP1 ) genes tended to be significantly higher in the three-dimensional (3D) cultures treated with OS compared to those 3D cultures without OS and the monolayer culture with OS (P Conclusion The 3D culture system improves odontogenic differentiation of DPSCs. The differentiation level of the cells in 3D culture is significantly lower than that of odontoblasts present in pulp tissue. TCP biomaterial possesses an odontogenic-inducing property.

Effects of interleukin-1β on mineralization potential of dental pulp stem cells

To investigate the effects of pro-inflammatory cytokine interleukin-1β (IL-1β) on mineralization potential of dental pulp stem cells (DPSC). Rat DPSC were cultured in vitro and randomly divided into three groups, IL-1β (10 µg/L), osteogenic inductive medium and non-osteogenic inductive medium. After 3, 7, and 12 days of treatment, the cultures were evaluated for cell proliferation and calcium deposit. Real-time polymerase chain reaction was used to detect the gene expression levels of osteocalcin (OC), bone sialoprotein (BSP), dentin sialophosphoprotein (DSPP) and dentin matrix protein 1 (DMP-1). In vivo test, after 3 day's treatment with IL-1β, the cell-scaffold complexes were implanted subcutaneously in mice for 8 weeks. Histological analysis was performed to evaluate hard tissue formation. In vitro test, after 3-day's treatment, IL-1β improved cell proliferation to 137.22 DNA µg/L and cell viability becomes (97.12 ± 7.18)% of control. The gene expression levels of OC, BSP, DSPP and DMP-1 are (378.19 ± 16.22)%, (427.12 ± 18.22)%, (247.19 ± 10.11)% and (198.29 ± 10.23)% respectively. The results of IL-1β's group was notable increased compared with non-osteogenic induction medium and the statistical differences are significant. IL-1β induced the odontogenic differentiation of DPSC. However, these effects tended to continuously decrease with treatment time. Histological analysis demonstrated that in the group treated with IL-1β hard tissue was markedly formed in vivo. IL-1β may induce the mineralization of DPSC and play an important role in host defenses and tissue repair.

The odontogenic differentiation of human dental pulp stem cells on nanofibrous poly(l-lactic acid) scaffolds in vitro and in vivo

The aim of this study was to investigate the odontogenic differentiation of human dental pulp stem cells (DPSCs) on nanofibrous (NF)-poly(l-lactic acid) (PLLA) scaffolds in vitro and in vivo. Highly porous NF-PLLA scaffolds which mimic the architecture of collagen type I fibers were fabricated by the combination of a phase-separation technique and a porogen-leaching method. The human DPSCs were then seeded onto the scaffolds and cultured in different media for odontogenic differentiation: "Control" medium without supplements; "DXM" medium containing 10(-8)M dexamethasone (DXM), 50 microgml(-1) ascorbic acid and 5mM beta-glycerophosphate; "BMP-7+DXM" medium containing 10(-8)M DXM, 50 microgml(-1) ascorbic acid, 5mM beta-glycerophosphate plus 50 ngml(-1) bone morphogenetic protein 7 (BMP-7). For odontogenic differentiation study in vitro, alkaline phosphatase activity quantification, reverse transcription polymerase chain reaction, scanning electron microscopy, von Kossa staining and calcium content quantification were carried out. While both "DXM" medium and "BMP-7+DXM" medium induced the DPSCs to odontoblast-like cells, the "BMP-7+DXM" medium had greater inducing capacity than the "DXM" medium. Consistent with the in vitro studies, the "BMP-7+DXM" group presented more extracellular matrix and hard tissue formation than the "DXM" group after 8 weeks of ectopic implantation in nude mice. Differentiation of DPSCs into odontoblast-like cells was identified by the positive immunohistochemical staining for dentin sialoprotein. In conclusion, odontogenic differentiation of DPSCs can be achieved on NF-PLLA scaffolds both in vitro and in vivo; the combination of BMP-7 and DXM induced the odontogenic differentiation more effectively than DXM alone. The NF-PLLA scaffold and the combined odontogenic inductive factors provide excellent environment for DPSCs to regenerate dental pulp and dentin.