Abstract Introduction This study aimed to evaluate the efficacy of epigallocatechin gallate (EGCG), an antibacterial cross-linking agent, on the proliferation and differentiation of human dental pulp ...cells (hDPCs) cultured in hydrogel collagen scaffolds. Methods The odontogenic differentiation induced by EGCG was evaluated by alkaline phosphatase (ALP) activity and odontogenic-related gene expression using real-time polymerase chain reaction. The antibacterial effect of EGCG was investigated by a disc diffusion assay in comparison with glutaraldehyde. Proliferation was analyzed by cell number counting under both optical and confocal laser scanning microscopes. To assess the mechanical properties of collagen treated with EGCG, the setting time, surface roughness, and compressive strength were measured. Results EGCG itself did not up-regulate the odontogenic-related markers ( P > .05) although ALP activity was slightly increased. The proliferation and differentiation of hDPCs cultured in collagen increased significantly in the presence of EGCG ( P < .05). The antibacterial activity of EGCG was similar to that of glutaraldehyde. The setting time of collagen was significantly shortened when it was treated with EGCG ( P < .05). The surface roughness and compressive strength of the cross-linked collagen were higher than those of collagen without EGCG ( P < .05). Conclusions Our results showed that EGCG, the antibacterial cross-linking agent, promoted the proliferation and differentiation of hDPCs cultured in collagen scaffolds. Furthermore, the enhanced mechanical properties of collagen scaffolds induced by EGCG may play important roles in cell behavior. Consequently, the application of EGCG to collagen scaffolds might be beneficial for regenerative endodontic therapy.
Abstract Introduction This study tested the null hypothesis that there is no difference between the use of 1.3% NaOCl/17% ethylenediaminetetraacetic acid (EDTA) and 5.25% NaOCl/17% EDTA irrigation ...regimens on the collagen degradation and flexural strength reduction in mineralized dentin. Methods Dentin powder and mineralized dentin sections were immersed in 1.3% or 5.25% NaOCl for 10–240 minutes and then rinsed with 17% EDTA as the final irrigant for 2 minutes. Untreated mineralized dentin powder/sections served as controls in the respective experiments. Dentin powders were examined by using Fourier transform infrared (FT-IR) spectroscopy to analyze their relative subsurface intact collagen content with the apatite/collagen ratio. Hydrated dentin sections were subjected to 3-point flexure under water for determining their flexural strengths. Results Collagen degradation was significantly increased and the flexural strength of mineralized dentin was significantly reduced after the use of 5.25% NaOCl as the initial irrigant for more than 1 hour ( P < .05). Conversely, changes were insignificant when 1.3% NaOCl was used as the initial irrigant for up to 4 hours (Kruskal-Wallis analysis of variance, n = 10, P < .05). Conclusions The null hypothesis was rejected. The deleterious effects attributed to the use of NaOCl on dentin are concentration-dependent and time-dependent and are not associated with the demineralization caused by the use of EDTA as the final active irrigant.
Abstract Introduction Four generations of methacrylate resin–based sealers have been available commercially. Three of these were introduced during the last 5 years when the concept of simultaneous ...bonding of root canal sealers to root filling materials and dentin was popularized. Methods This article presents an overview of methacrylate resin–based sealers, with the objectives of clarifying the behavior of these materials and delineating their limitations in clinical application. Results The first generation sealer was introduced in the mid-1970s. The initial enthusiasm associated with its use eventually diminished as a result of its suboptimal physical, biologic, and clinical properties. With advances in self-etching adhesive technology acquired from adhesive dentistry, methacrylate resin–based sealers were reintroduced in the beginning of the 21st century to support the introduction of bondable root canal filling materials. Three different generations of these sealers have since been available commercially. Although some in vitro studies on the sealing ability, self-etching potential, biocompatibility, and removability of the sealers showed better potential over conventional nonbonding sealers, accomplishing the ideal goal of a monoblock in the root canal space with these materials is still regarded as a major challenge. Conclusions On the basis of the in vitro and in vivo data available to date, there appears to be no clear benefit with the use of methacrylate resin–based sealers in conjunction with adhesive root filling materials at this point in their development.
Abstract Introduction The aim of this study was to investigate the effects of genipin, a natural collagen cross-linking agent, on odontogenic differentiation of human dental pulp cells (hDPCs) ...because the mechanical properties of collagen allow it to serve as a scaffold for engineering of pulp-dentin complex. Furthermore, the role of extracellular signal–regulated kinase (ERK) was investigated as a mediator of the differentiation. Methods The odontogenic differentiation was analyzed by alkaline phosphatase activity, real time-polymerase chain reaction, Western blotting, and alizarin red S staining. The morphologic features of hDPCs cultured in genipin-treated collagen were evaluated by scanning electron microscopy. For the assessment of mechanical properties of collagen treated with genipin, the surface roughness and compressive strength were measured. Results Alkaline phosphatase activity, the expression of odontogenic markers, and mineralized nodule formation increased in the genipin-treated group. Genipin also activated ERK, and treatment with ERK inhibitor blocked the expression of the markers. The cells cultured in genipin-treated collagen spread across the substrate and attached in close proximity to one another. The proliferation and differentiation of hDPCs cultured in genipin-treated collagen were facilitated. The mechanical properties of collagen, such as surface roughness and compressive strength, were increased by treatment with genipin. Conclusions Our results show that genipin promotes odontogenic differentiation of hDPCs via the ERK signaling pathway. Furthermore, the enhanced mechanical properties of the collagen scaffold induced by genipin may play important roles in cell fate. Consequently, the application of genipin might be a new strategy for dentin-pulp complex regeneration.
Abstract Background Fibroblast growth factor-2 (FGF-2) participates in both hematopoiesis and osteogenesis; however, the effects of FGF-2 on chemokines during odontoblastic differentiation have not ...been reported. This study investigated whether human dental pulp cells (HDPCs) treated with FGF-2 could express chemokines during differentiation into odontoblastic cells and sought to identify its underlying mechanism of action. Methods To analyze differentiation, we measured alkaline phosphatase (ALP) activity, calcified nodule formation by alizarin red staining, and marker RNA (mRNA) expression by reverse-transcriptase polymerase chain reaction (RT-PCR). Expression of chemokines, such as interleukin-6 (IL-6), IL-8, monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein-1α (MIP-1α), and MIP-3α, were evaluated by RT-PCR. Results ALP activity, the mineralization, and mRNA expression for odontoblastic markers were enhanced by FGF-2 in HDPCs. FGF-2 also up-regulated the expression of IL-6, IL-8, MCP-1, MIP-1α, and MIP-3α mRNAs, which were attenuated by inhibitors of p38, ERK1/2 and p38 MAP kinases, protein kinase C, phosphoinositide-3 kinase, and NF-κB. Conclusion Taken together, these data suggest that FGF-2 plays a role not only as a differentiation inducing factor in the injury repair processes of pulpal tissue but also as a positive regulator of chemokine expression, which may help in tissue engineering and pulp regeneration using HDPCs. However, the fate of odontoblastic or osteoblastic differentiation, effective local delivery for FGF-2, interaction of chemotatic and odontogenic factors, and other limitations will need to be overcome before a major modality for the treatment of pulp disease.
Abstract Introduction Peroxisome proliferator-activated receptor gamma (PPARγ) has well-known anti-inflammatory action in human dental pulp cells (HDPCs). The purpose of this study was to investigate ...whether the anti-inflammatory action of PPARγ involves in cellular cytoprotection and supports odontoblast differentiation under oxidative stress in HDPCs. Methods To simulate long-term oxidative stress, pulp cells were treated with 150 μmol hydrogen peroxide (H2 O2 ) for 12 days. The replication deficiency adenovirus (adenovirus PPARγ) was introduced for PPARγ overexpression in pulp cells. The cellular cytotoxicity and reactive oxygen species formation by H2 O2 were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and 2′,7′-dichlorodihydrofluorescein diacetate with fluorescence-activated cell sorting assay. To determine the roles of PPARγ, several molecules of odontogenic/osteogenic and signal pathway were analyzed by reverse-transcription polymerase chain reaction and Western hybridization. Dentin mineralization was determined by alizarin red stain and alkaline phosphatase activity assay. Results Pulp cells treated with long-term H2 O2 showed high reactive oxygen species formation, low cell viability, down-expression of antioxidant molecules (Cu/Zn and Mn superoxide dismutase), and odontogenic/osteogenic markers (eg, dentin sialophosphoprotein, dentin matrix protein-1, osteopontin, bone sialoprotein, Runx-2, and bone morphogenetic protein 2 and 7). In addition, pulp cells with oxidative stress underwent the activation of ERK1/2, activator protein-1, and nuclear factor-κB translocation to the nucleus. However, the PPARγ-overexpressed cells gave opposite results although under oxidative stress. Furthermore, PPARγ and its agonist rosiglitazone exhibited an induction of dentin mineralization under oxidative stress. Conclusions PPARγ in pulp cells increases cell viability, odontoblastic differentiation, and dentin mineralization under oxidative stress. These results offer new insights into the potential antioxidative activity of PPARγ and its agonist for therapeutic agents for pulp vitality in HDPCs.
Abstract Introduction Information on the nerve fibers innervating the dental pulp is crucial for understanding dental pain and hypersensitivity. This study investigated the morphologic differences of ...parvalbumin (PV)-positive (+) myelinated fibers in 3 different regions of the human dental pulp. Methods Light and electron microscopic immunohistochemistry for parvalbumin, a marker for myelinated fibers, and quantitative analysis were performed in the apical root, core of coronal pulp, and peripheral pulp of human premolar teeth. Results About 40% of the myelinated fibers in the apical root pulp became unmyelinated in the core of the coronal pulp, and virtually all the remaining fibers became unmyelinated at the peripheral pulp. The size of myelinated axons decreased from root to peripheral pulp. PV+ axons showed extensive axonal varicosities in the peripheral pulp. Conclusions These findings suggest that the myelinated fibers innervating the human dental pulp undergo extensive morphologic change in the extrapulpal region and in the coronal and peripheral pulp, and that PV-mediated regulation of calcium concentration and its downstream events may occur primarily in axonal varicosities in the peripheral pulp.
Abstract Introduction Although bacterial infection and heat stress are common causes of injury in human dental pulp cells (HDPCs), little is known about the potential defense mechanisms mediating ...their effects. This study examined the role of SIRT1 in mediating heat stress and lipopolysaccharide (LPS)-induced immune and defense gene expression in HDPCs. Methods HDPCs were exposed to heat stress (42°C) for 30 minutes after stimulation with LPS (1 μg/mL) for 48 hours. The expression of defense genes was evaluated by reverse-transcriptase polymerase chain reaction, Western blotting, and enzyme-linked immunosorbent assay. Results LPS and heat stress synergistically increased the expression of SIRT1 and immune and defense genes such as interleukin (IL)-8, hemeoxygenase-1 (HO-1), and human β-defensin 2 (hBD-2). Resveratrol enhanced LPS- and heat stress–induced expression of HO-1 and hBD-2 but reduced IL-8 messenger RNA levels. The stimulation of HO-1 and hBD-2 messenger RNA expression by LPS and heat stress was inhibited by sirtinol; SIRT1 small interfering RNA; and inhibitors of p38, ERK, JNK, and nuclear factor κB. Conclusions These results show for the first time that SIRT1 mediates the induction of immune and defense gene expression in HDPCs by LPS and heat stress. SIRT1 may play a pivotal role in host immune defense system in HDPCS.
Abstract Introduction Although simvastatin has multiple demonstrable effects, its function in dentinogenesis remains unclear. In this study, we tested the hypothesis that the addition of simvastatin ...to human dental pulp cells (HDPCs) stimulates odontogenesis both by promoting odontoblastic differentiation and by favoring the release of angiogenic factors. In addition, the role of heme oxygenase-1 (HO-1) in these effects was investigated. Methods The expression of markers for odontoblastic differentiation and angiogenesis was analyzed by means of alkaline phosphatase (ALP) activity, alizarin red staining, and Western blotting. Results Simvastatin enhanced the differentiation of HDPCs by up-regulating mineralization nodules and odontogenic markers as well as angiogenic markers. These phenomena were then correlated with the induction of HO-1 protein levels. The inducing effect of simvastatin on odontoblastic differentiation and angiogenesis was nullified by an HO-1 inhibitor and a carbon monoxide (CO) scavenger. Conclusions These results suggested that simvastatin exerts its odontoblastic differentiation and angiogenesis-inducing effects in HDPCs through a mechanism that involves the action of HO-1 and its product CO.
Abstract This study was conducted to investigate the effects of mechanical stress, particularly cyclic strain, on proinflammatory cytokines as well as antioxidant properties and their interactions ...with cellular defense systems in human dental pulp (HDP) cells. Exposure of HDP cells to mechanical strain induced inflammatory cytokines such as interleukin-1β, tumor necrosis factor–α, and interleukin-6, as well as antioxidant genes such as heme oxygenase-1, superoxide dismutases, reduced nicotinamide adenine dinucleotide phosphate quinone oxidoreductase-1, and glutathione peroxidases. In addition, treatment with N-acetylcysteine, indomethacin, and heme oxygenase–1 inhibitors blocked reactive oxygen species production, antioxidant response element (ARE) gene expression, and Nrf2 accumulation that occurred in response to mechanical stress. These data demonstrate that mechanical strain activates inflammatory cytokines and oxidative stress, which then act in concert to induce the Nrf2-/ARE-mediated antioxidant enzymes. Therefore, we suggest that the activation of a compensatory adaptation or defense antioxidant system might represent a novel mechanism for protecting HDP cells against mechanical stress.
