Gene expression changes in bioceramic paste-treated human dental pulp cells (original) (raw)
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Biomaterials, 2010
Biocompatibility of dentin bonding agents (DBAs) and resin composite is important to preserve the pulp vitality after operative restoration. Bisphenol-glycidyl-methacrylate (BisGMA) is one common monomer adding into DBAs and resin. In this study, we found that exposure of human dental pulp cells to BisGMA (>0.1 mM) led to cytotoxicity, G2/M cell cycle arrest and apoptosis as analyzed by propidium iodide (PI) and PI/annexin V dual fluorescent flow cytometry. These events were associated with a decline of cdc2, cdc25C and cyclinB1 expression at both mRNA and protein levels. BisGMA also induced the expression of hemeoxygenase-1 (HO-1), an oxidative stress responsive gene, in pulp cells. Catalase could prevent the BisGMA-induced alteration of cell cycle-related genes (cdc2, cdc25C, cyclinB1) and HO-1 expression in dental pulp cells. Interestingly, Zn-protoporphyrin (2.5e5 mM), a HO inhibitor, enhanced the BisGMA-induced reactive oxygen species (ROS) production and cytotoxicity. These results suggest that exposure to higher concentrations of BisGMA may stimulate ROS production, cell cycle arrest, apoptosis and cell death. Inducing the expression of HO-1 in dental pulp cells by BisGMA is mediated by ROS production and important to protect dental pulp against the toxicity by monomers present in composite resin and DBAs.
The Journal of Contemporary Dental Practice
Aim: The aim of this study was to compare the effects of white MTA-Angelus (wMTA), Biodentine® (Biodentine) and TotalFill® BC Root Repair Material TM putty (TotalFill) on human dental pulp stromal cells (hDPSCs) in vitro. Materials and methods: hDPSCs were isolated from third molars of healthy young adults. Material elutes at different concentrations were prepared. Cells were exposed to the eluates for 1, 3, and 7 days. Cell proliferation was evaluated using 3-(4,5-dimethyl-thiazoyl)-2, 5-diphenyltetrazolium bromide assay. The expression of alkaline phosphatase (ALP), osteoprotegerin (OPG), osteocalcin (OC), collagen1A (Col1A), runt-related transcription factor 2 (RUNX2), vascular endothelial growth factor-A (VEGF-A), fibroblast growth factor-1 (FGF-1), interleukin 6 (IL6), tumor necrosis factor alpha (TNFα), and interleukin-1-beta (IL1β) was determined by reverse transcription-polymerase chain reaction (RT-PCR). VEGF-A protein levels and ALP activity were quantified in the culture supernatant. Data were analyzed by two-way analysis of variance (ANOVA). p values <0.05 were considered statistically significant. Results: hDPSC proliferation was decreased in a dose-related manner for all materials on day 3. The same effect was observed with wMTA and TotalFill on day 7. RT-PCR showed that Biodentine increased the expression of the osteogenic markers ALP, OPG, and OC. TotalFill decreased the ALP expression and activity, enhanced the production of angiogenic VEGF-A, and downregulated the inflammatory IL6 on day 7. Conclusion: Although the tested materials are used interchangeably in vital pulp therapy, the findings showed varied hDPSC responses. Biodentine did not affect cell proliferation and increased the expression of osteo-/odontogenic markers compared to wMTA and TotalFill, whereas TotalFill decreased cell proliferation and exhibited enhanced angiogenic and anti-inflammatory effects over time. Clinical significance: The clinical significance of the results needs further investigation in an attempt to provide recommendations on the selection of bioceramic pulp capping material under different scenarios of pulpal pathosis.
Microarray expression profiling of human dental pulp from single subject
Clinical and investigative medicine. Médecine clinique et experimentale, 2008
Microarray is a recently developed simultaneous analysis of expression patterns of thousand of genes. The aim of this research was to evaluate the expression profile of human healthy dental pulp in order to find the presence of genes activated and encoding for proteins involved in the physiological process of human dental pulp. We report data obtained by analyzing expression profiles of human tooth pulp from single subjects, using an approach based on the amplification of the total RNA. Experiments were performed on a high-density array able to analyse about 21,000 oligonucleotide sequences of about 70 bases in duplicate, using an approach based on the amplification of the total RNA from the pulp of a single tooth. Obtained data were analyzed using the S.A.M. system (Significance Analysis of Microarray) and genes were merged according to their molecular functions and biological process by the Onto-Express software. The microarray analysis revealed 362 genes with specific pulp expres...
The Effect of Tooth Bleaching on Substance P Expression in Human Dental Pulp
Journal of Endodontics, 2008
The purpose of this study was to quantify the effect of tooth bleaching on substance P (SP) expression in healthy human dental pulp. Forty pulp samples were obtained from healthy premolars in which extraction was indicated for orthodontic reasons. Thirty of these premolars were assigned into three different tooth-bleaching protocols: group 1 (n ϭ 10): Opalescence Xtra Boost (Ultradent Products, South Jordan, UT) (38% H 2 O 2) for 15 minutes; group 2 (n ϭ 10): Lase Peroxide (DMC, Brazil) (35% H 2 O 2) activated with infrared laser diode (Biolux; BioArt, Brazil) for 3 minutes, and group 3 (n ϭ 10): Zoom! Whitening System (Discuss Dental, Culver City, CA) (25% H 2 O 2) light activated for 20 minutes. The remaining 10 healthy premolars serve as a control group. Teeth were anesthetized immediately after bleaching and were extracted 10 minutes later. All pulp samples were processed and SP was measured by radioimmunoassay. Greater SP expression was found in the Zoom! Whitening System, followed by the Lase Peroxide group, Opalescence Xtra Boost, and the lower SP values were for the control group. Analysis of variance showed statistically significant differences between groups (p ϭ 0.0001). Tukey HSD post hoc tests showed significant differences in the light (p Ͻ 0.01) and laser (p Ͻ 0.05) activated bleaching systems when compared with control values. It can be concluded that lightand laser-activated tooth-bleaching systems increase SP expression in human dental pulp significantly higher than normal values.
Materials, 2018
The purpose of this study was to evaluate the diffusion capacity and the biological effects of different bleaching products on human dental pulp stem cells (hDPSCs). The bleaching gel was applied for 90, 30 or 15 min to enamel/dentine discs that adapted in an artificial chamber. The diffusion of hydrogen peroxide (HP) was analysed by fluorometry and the diffusion products were applied to hDPSCs. Cell viability, cell migration and cell morphology assays were performed using the eluates of diffusion products. Finally, cell apoptosis and the expression of mesenchymal stem cell markers were analysed by flow cytometry. Statistical analysis was performed using analysis of variance and Kruskal-Wallis or Mann-Whitney tests (α < 0.05). Significant reductions of approximately 95% in cell viability were observed for the 3 × 15 min groups (p < 0.001), while 1 × 30 min of PerfectBleach and 1 × 90 min of PolaNight resulted in reductions of 50% and 60% in cell viability, respectively (p < 0.001). Similar results were obtained in the migration assay. Moreover, the 3 × 15 min group was associated with cell morphology alterations and reductions of >70% in cell live. Finally, hDPSCs maintained their mesenchymal phenotype in all conditions. Similar concentrations of carbamide peroxide (CP) and HP in different commercial products exhibited different biological effects on hDPSCs.
Physiological and molecular responses in the pulp associated with early inflammatory process
2016
Therapies to promote pulp repair and regeneration after injury should be underpinned by a deep understanding of normal tissue behaviour, cellular cross-talk and regulation. The objectives of this study were to investigate the expression of cyclooxygenases (COX1 and COX2), prostanoid receptors (EP1 and EP2) and nitric oxide synthase 1 (NOS1) within the normal dental pulp. The effect of experimental inflammatory conditions on these elements at mRNA level (both normal and experimentally inflamed) were investigated to explore the possibility of a nitric oxide (NO)/prostaglandin (PG) signalling pathway interaction in the rat mandibular incisor (normal and experimentally inflamed). Rodent mandibular incisors were utilised as a model throughout this thesis with structural and functional investigations on demineralised teeth, non-demineralised freshly extracted pulp tissues and tissue explants. The work described in this thesis used immunohistochemical, ELISA and quantitative reverse transcription polymerase chain reaction (q-RT-PCR) techniques. Cellular heterogeneity was observed both in the odontoblasts population and in the interstitial cells forming the bulk of the pulp. Cellular processes were also observed in addition to the observation of cellular processes extending from interstitial cells in the cell-rich zone to odontoblasts. The presence and localisation of immunoreactivity to the above mentioned targets (COX1, COX2, NOS1 and prostanoid receptors) are novel and confirmed with q-RT-PCR. Isolated pulp tissues exposed to LPS were found to release an increased amount of PGE2, which was found to be inhibited by other factors like the presence of NO and ATP. Functional cross-talk between PG and NO was investigated by the ELISA technique, with experimentally-induced inflammation increasing PG release, whereas NO and ATP caused inhibition of PGE2 release. The effects of carefully selected pro-inflammatory agents (LPS, NO and ATP) on PG pathway were examined at the mRNA level by q-RT-PCR. Exposure to LPS was found to cause upregulation of all target genes, whereas variable reactions were observed in response to incubation with NO, ATP and PGE2.
Inhibition of odontogenic differentiation of human dental pulp cells by dental resin monomers
Biomaterials research, 2015
Dental resin monomers that are leached from the resin matrix due to incomplete polymerization can affect the viability and various functions of oral tissues and cells. In this study, the effects of triethylene glycol dimethacrylate (TEGDMA) and 2-hydroxyethyl methacrylate (HEMA) on odontogenic differentiation of human dental pulp cells (HDPCs) were examined. To mimic clinical situations, dental pulp cells were treated with resin monomers for 24 h prior to the analysis of alkaline phosphatase (ALP) activity and mRNA expression of genes related to pulp cell differentiation. To elucidate the underlying signaling pathways, regulation of mitogen-activated protein (MAP) kinases by resin monomers was also investigated. The ALP activity of HDPCs was reduced by TEGDMA and HEMA at noncytotoxic concentrations. The mRNA expression of dentin sialophosphoprotein (DSPP), osteocalcin (OCN), and osteopontin (OPN) was also downregulated by resin monomers. However, DSPP expression was not affected by ...
Oxidative stress and cytotoxicity generated by dental composites in human pulp cells
Clinical Oral Investigations, 2012
Dental composites are a source of residual monomers that are released into the oral environment. Since monomers act on cultured cells through reactive oxygen species (ROS), we hypothesized that composites generate ROS associated with cytotoxicity. Human pulpderived cells were exposed to extracts of methacrylatebased materials including triethylene glycol dimethacrylate and 2-hydroxyethyl methacrylate-free composites (Tetric Ceram, Tetric EvoCeram, els, els flow, Solitaire 2) and a silorane-based composite (Hermes III). The materials were polymerized in the presence and absence of a polyester film and then extracted in culture medium. The generation of ROS was measured by flow cytometry, and cytotoxicity was determined as well. Methacrylate-based composites reduced cell survival but varied in efficiency. Undiluted extracts of Solitaire 2 specimens prepared in the absence of a polyester film reduced cell survival to 26% compared with untreated cultures. Cytotoxicity was reduced when specimens were covered with a polyester film during preparation. Cytotoxicity of the composites was ranked as follows: Solitaire 2 >> els flow > Tetric Ceram = Tetric EvoCeram = els > Hermes III. The generation of ROS followed the same pattern as detected with cytotoxic effects. A positive correlation was found between ROS production and cell survival caused by extracts made from materials not covered with a polyester film. These findings suggest that components released from composites affect cellular signaling networks through ROS formation. Regenerative and reparative capacities of the dentine-pulp complex may be impaired by biologically active resin monomers released from composite restorations.
Characterization of human dental pulp cells grown in chemically defined serum-free medium
Biomedical reports, 2018
Dental pulp cells (DPCs) are promising candidates for use as transplantable cells in regenerative medicine. However, expansion of these cells typically requires culture media containing fetal bovine serum, which may cause infection and immunological reaction following transplantation. In addition, the proliferation and differentiation of DPCs markedly depend upon serum batches. Therefore, the present study examined whether DPCs could be expanded under serum-free conditions. DPCs obtained from four donors were identified to proliferate actively in the serum-free medium, STK2, when compared with those cells in control medium (Dulbecco's modified Eagle's medium containing 10% serum). The high proliferative potential with STK2 was maintained through multiple successive culture passages. DNA microarray analyses demonstrated that the gene expression profile of DPCs grown in STK2 was similar to that of cells grown in the control medium; however, a number of genes related to cell pr...