• Biomaterials Research
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• v.22 no.4
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• pp.346-351
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• 2018

Background: Carbonate apatite ($CO_3Ap$) and silica-calcium phosphate composite (SCPC) are bone substitutes with good prospect for dental application. SCPC creates a hydroxyapatite surface layer and stimulate bone cell function while, $CO_3Ap$ induce apatite crystal formation with good adaptation providing good seal between cement and the bone. Together, these materials will add favorable properties as a pulp capping material to stimulate mineral barrier and maintain pulp vitality. The aim of this study is to investigate modification of $CO_3Ap$ cement combined with SCPC, later term as $CO_3Ap-SCPC$ cement (CAS) in means of its chemical (Calcium release) and physical properties (setting time, DTS and pH value). Methods: The study consist of three groups; group 1 (100% calcium hydroxide, group 2 $CO_3Ap$ (60% DCPA: 40% vaterite, and group 3 CAS (60% DCPA: 20% vaterite: 20% SCPC. Distilled water was employed as a solution for group 1, and $0.2mol/L\;Na_3PO_4$ used for group 2 and group 3. Samples were evaluated with respect to important properties for pulp capping application such as pH, setting time, mechanical strength and calcium release evaluation. Results: The fastest setting time was in $CO_3Ap$ cement group without SCPC, while the addition of 20% SCPC slightly increase the pH value but did not improved the cement mechanical strength, however, the mechanical strength of both $CO_3Ap$ groups were significantly higher than calcium hydroxide. All three groups released calcium ions and had alkaline pH. Highest pH level, as well as calcium released level, was in the control group. Conclusion: The CAS cement had good mechanical and acceptable chemical properties for pulp capping application compared to calcium hydroxide as a gold standard. However, improvements and in vivo studies are to be carried out with the further development of this material.

• Biomaterials and Biomechanics in Bioengineering
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• v.3 no.2
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• pp.105-114
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• 2016

The purpose of this study is to evaluate the effects of cryopreservation on dental pulp-derived stem cells (DPSC) viability over a period of three years. Dental pulp-derived stem cells were isolated and cultured from thirty-one healthy teeth. DPSC isolates were assessed for doubling-time and baseline viability prior to cryopreservation and were assessed again at three time points; one week (T1), 18 months (T2), and 36 months (T3). DPSC can be grouped based on their observed doubling times; slow (sDT), intermediate (iDT), and rapid (rDT). Viability results demonstrated all three types of DPSC isolates (sDT, iDT and rDT) exhibit time-dependent reductions in viability following cryopreservation, with the greatest reduction observed among sDT-DPSCs and the smallest observed among the rDT-DPSC isolates. Cryopreserved DPSCs demonstrate time-dependent reductions in cellular viability. Although reductions in viability were smallest at the initial time point (T1) and greatest at the final time point (T3), these changes were markedly different among DPSC isolates with similar doubling times (DTs). Furthermore, the analysis of various DPSC biomarkers - including both intracellular and cell surface markers, revealed differential mRNA expression. More specifically, the relative high expression of Sox-2 was only found only among the rDT isolates, which was associated with the smallest reduction in viability over time. The expression of Oct4 and NANOG were also higher among rDT isolates, however, expression was comparatively lower among the sDT isolates that had the highest reduction in cellular viability over the course of this study. These data may suggest that some biomarkers, including Sox-2, Oct4 and NANOG may have some potential for use as biomarkers that may be associated with either higher or lower cellular viability over long-term storage applications although more research will be needed to confirm these findings.

• Restorative Dentistry and Endodontics
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• v.44 no.2
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• pp.20.1-20.8
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• 2019

Objectives: To achieve pulp-dentin complex regeneration with tissue engineering, treatment efficacies and safeties should be evaluated using in vivo orthotopic transplantation in a sufficient number of animals. Mice have been a species of choice in which to study stem cell biology in mammals. However, most pulp-dentin complex regeneration studies have used large animals because the mouse tooth is too small. The purpose of this study was to demonstrate the utility of the mouse tooth as a transplantation model for pulp-dentin complex regeneration research. Materials and Methods: Experiments were performed using 7-week-old male Institute of Cancer Research (ICR) mice; a total of 35 mice had their pulp exposed, and 5 mice each were sacrificed at 1, 2, 4, 7, 9, 12 and 14 days after pulp exposure. After decalcification in 5% ethylenediaminetetraacetic acid, the samples were embedded and cut with a microtome and then stained with hematoxylin and eosin. Slides were observed under a high-magnification light microscope. Results: Until 1 week postoperatively, the tissue below the pulp chamber orifice appeared normal. The remaining coronal portion of the pulp tissue was inflammatory and necrotic. After 1 week postoperatively, inflammation and necrosis were apparent in the root canals inferior to the orifices. The specimens obtained after experimental day 14 showed necrosis of all tissue in the root canals. Conclusions: This study could provide opportunities for researchers performing in vivo orthotopic transplantation experiments with mice.

• Journal of the korean academy of Pediatric Dentistry
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• v.35 no.1
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• pp.11-17
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• 2008

The purpose of this study was to investigate early histological changes and calcitonin gene-related peptide (CGRP) expression in the dental pulp of the rat after Er:YAG laser preparation. Occlusal cavities were prepared in the upper first molars using either Er:YAG laser and conventional bur. At 48 hours after cavity preparation, the teeth were processed for hematoxylin-eosin stain and CGRP immunohistochemistry. The results were as follows : 1. The cavity floor by Er:YAG laser preparation was more irregular shape compared with those by bur preparation and there are some cracks in the directions of dentinal tubules. 2. There were more inflammatory cell infiltration and disruption of odontoblast in the dental pulp by Er:YAG laser preparation in comparison with the dental pulp by bur preparation. 3. CGRP expression in the pulp tissue by both Er:YAG laser and bur preparations were increased and higher than in the normal pulp. The expression pattern of CGRP was more strong in the pulp by Er:YAG laser preparation. These results indicate that Er:YAG laser is useful in the operative dentistry such as caries removal and cavity preparation if properly applied.

• Restorative Dentistry and Endodontics
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• v.35 no.3
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• pp.152-163
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• 2010

This study investigated the changes in gene expression when mineral trioxide aggregate (MTA) was applied in vitro to human dental pulp cells (HDPCs). MTA in a teflon tube (diameter 10 mm, height 2 mm) was applied to HDPCs. Empty tube-applied HDPCs were used as negative control. For microarray analysis, total RNA was extracted at 6, 24, and 72 hrs after MTA application. The results were confirmed selectively by performing reverse transcriptase polymerase chain reaction for genes that showed changes of more than two-fold or less than half. Of the 24,546 genes, 109 genes were up-regulated greater than twofold (e.g., FOSB, THBS1, BHLHB2, EDN1, IL11, FN1, COL10A1, and TUFT1) and 69 genes were down-regulated below 50% (e.g., SMAD6 and DCN). These results suggest that MTA, rather than being a bio-inert material, may have potential to affect the proliferation and differentiation of pulp cells in various ways.

• Journal of the korean academy of Pediatric Dentistry
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• v.40 no.3
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• pp.185-193
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• 2013

Mineral trioxide aggregate (MTA) has recently been used as a pulpotomy medicament for primary molars. The aim of this study was to evaluate and compare the proliferation and differentiation potential of dental pulp stromal cells of permanent teeth and deciduous teeth cultured on MTA-coated surface. Human dental pulp stromal cells were obtained from human permanent premolars and deciduous teeth and cultured on MTA-coated culture plates. The cells were subjected to proliferation assay and cell cycle analysis. Their differentiation potential was evaluated by analysing changes in the mRNA expressions of runt-related transcriptional factor 2 (Runx2) and alkaline phosphatase (ALP). Morphological changes of cells in direct contact with MTA were observed using scanning electron microscopy (SEM). The proliferation rates, distribution of cell cycles and mRNA expression patterns of Runx2 and ALP were similar in both types of pulpal cells. SEM observations revealed that both types changed into more dendrite-like cells. On the surface of MTA, human dental pulp stromal cells from deciduous and permanent teeth were able to both proliferate and differentiate into cells that induce mineralization. MTA is suitable as a biocompatible pulpotomy medicament for primary teeth.

• Restorative Dentistry and Endodontics
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• v.41 no.2
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• pp.98-105
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• 2016

Objectives: The biocompatibility of two experimental scaffolds for potential use in revascularization or pulp regeneration was evaluated. Materials and Methods: One resilient lyophilized collagen scaffold (COLL), releasing metronidazole and clindamycin, was compared to an experimental injectable poly(lactic-co-glycolic) acid scaffold (PLGA), releasing clindamycin. Human dental pulp stem cells (hDPSCs) were seeded at densities of $1.0{\times}10^4$, $2.5{\times}10^4$, and $5.0{\times}10^4$. The cells were investigated by light microscopy (cell morphology), MTT assay (cell proliferation) and a cytokine (IL-8) ELISA test (biocompatibility). Results: Under microscope, the morphology of cells coincubated for 7 days with the scaffolds appeared healthy with COLL. Cells in contact with PLGA showed signs of degeneration and apoptosis. MTT assay showed that at $5.0{\times}10^4$ hDPSCs, COLL demonstrated significantly higher cell proliferation rates than cells in media only (control, p < 0.01) or cells co-incubated with PLGA (p < 0.01). In ELISA test, no significant differences were observed between cells with media only and COLL at 1, 3, and 6 days. Cells incubated with PLGA expressed significantly higher IL-8 than the control at all time points (p < 0.01) and compared to COLL after 1 and 3 days (p < 0.01). Conclusions: The COLL showed superior biocompatibility and thus may be suitable for endodontic regeneration purposes.

• Restorative Dentistry and Endodontics
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• v.34 no.5
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• pp.415-423
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• 2009

The purpose of this study is to investigate the response of human pulp cell on Portland cement mixed with $\beta$-glycerophosphate. To investigate the effect of $\beta$-glycerophosphate and/or dexamethasone on human pulp cell, ALP activity on various concentration of $\beta$-glycerophosphate and dexamethasone was measured and mineral nodule of human pulp cell was stained with Alizarin red S. MTS assay and ALP activity of human pulp cell on Portland cement mixed with various concentration of $\beta$-glycerophosphate (10 mM, 100mM, 1M) was measured and the specimens were examined under SEM. Addition of $\beta$-glycerophosphate or dexamethasone alone had no effect however, the addition of 5 mM $\beta$-glycerophosphate and 100 nM dexamethasone had the largest increasement in ALP activity. There was no toxicity in all samples and the data showed that Portland cement mixed with 10 mM $\beta$-glycerophosphate had more increase in ALP activity compared with control. In conclusion, Portland cement mixed with $\beta$-glycerophosphate has no toxicity and promotes differentiation and mineralization of pulp cell compared with additive-free Portland cement. This implicated that application of Portland cement mixed with $\beta$-glycerophosphate might form more reparative dentin and in turn it would bring direct pulp capping to success.

• The Korean Journal of Physiology and Pharmacology
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• v.18 no.1
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• pp.25-32
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• 2014

Nitric oxide (NO) is recognized as a mediator and regulator of inflammatory responses. NO is produced by nitric oxide synthase (NOS), and NOS is abundantly expressed in the human dental pulp cells (HDPCs). NO produced by NOS can be cytotoxic at higher concentrations to HDPCs. However, the mechanism by which this cytotoxic pathway is activated in cells exposed to NO is not known. The purpose of this study was to elucidate the NO-induced cytotoxic mechanism in HDPCs. Sodium nitroprusside (SNP), a NO donor, reduced the viability of HDPCs in a dose- and time-dependent manner. We investigated the in vitro effects of nitric oxide on apoptosis of cultured HDPCs. Cells showed typical apoptotic morphology after exposure to SNP. Besides, the number of Annexin V positive cells was increased among the SNP-treated HDPCs. SNP enhanced the production of reactive oxygen species (ROS), and N-acetylcysteine (NAC) ameliorated the decrement of cell viability induced by SNP. However, a soluble guanylate cyclase inhibitor (ODQ) did not inhibited the decrement of cell viability induced by SNP. SNP increased cytochrome c release from the mitochondria to the cytosol and the ratio of Bax/Bcl-2 expression levels. Moreover, SNP-treated HDPCs elevated activities of caspase-3 and caspase-9. While pretreatment with inhibitors of caspase (z-VAD-fmk, z-DEVD-fmk) reversed the NO-induced apoptosis of HDPCs. From these results, it can be suggested that NO induces apoptosis of HDPCs through the mitochondria-dependent pathway mediated by ROS and Bcl-2 family, but not by the cyclic GMP pathway.

• International Journal of Oral Biology
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• v.39 no.4
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• pp.177-185
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• 2014

Transfection is a gene delivery tool that is a popular means of manipulating cellular properties, such as induced pluripotent stem cell (iPSC) generation by reprogramming factors (Yamanaka factors). However, the efficiency of transfection needs to be improved. In the present study, three transfection protocols - non-liposomal transfection (NLT), magnetofection and electroporation - were compared by analysis of their transfection efficiencies and cell viabilities using human dental pulp cells (hDPC) and bovine fetal fibroblasts (bFF) as cell sources. Enhanced green fluorescent protein gene was used as the delivery indicator. For magnetofection, Polymag reagent was administrated. NLT, FuGENE-HD and X-treme GENE 9 DNA transfection reagents were used for NLT. For electroporation, the $Neon^{TM}$ and $NEPA21^{TM}$ electroporators were tested. $Neon^{TM}$ electroporation showed highest transfection efficiency when compared with NLT, magnetofection, and $NEPA21^{TM}$ electroporation, with transfection efficiency of about 33% in hDPC and 50% in bFF, based on viable cell population in each cell type. These results suggest that transfection by $Neon^{TM}$ electroporation can be used to deliver foreign genes efficiently in human and bovine somatic cells.