Genetic information such as DNA sequences has been limited to fully explain mechanisms of gene regulation and disease process. Epigenetic mechanisms, which include DNA methylation, histone modification and non-coding RNAs, can regulate gene expression and affect progression of disease. Although studies focused on epigenetics are being actively investigated in the field of medicine and biology, epigenetics in dental research is at the early stages. However, studies on epigenetics in dentistry deserve attention because epigenetic mechanisms play important roles in gene expression during tooth development and may affect oral diseases. In addition, understanding of epigenetic alteration is important for developing new therapeutic methods. This review article aims to outline the general features of epigenetic mechanisms and describe its future implications in the field of dentistry.

While it is reasonably well known that certain dental procedures increase the temperature of the tooth's surface, of greater interest is their potential damaging effect on the pulp and tooth-supporting tissues. Previous studies have investigated the responses of the pulp, periodontal ligament, and alveolar bone to thermal irritation and the temperature at which thermal damage is initiated. There are also many in vitro studies that have measured the temperature increase of the pulp and tooth-supporting tissues during restorative and endodontic procedures. This review article provides an overview of studies measuring temperature increases in tooth structures during several restorative and endodontic procedures, and proposes clinical guidelines for reducing potential thermal hazards to the pulp and supporting tissues.

The limited durability of resin-dentin bonds severely compromises the longevity of composite resin restorations. Resin-dentin bond degradation might occur via degradation of water-rich and resin sparse collagen matrices by host-derived matrix metalloproteinases (MMPs). This review article provides overview of current knowledge of the role of MMPs in dentin matrix degradation and four experimental strategies for extending the longevity of resin-dentin bonds. They include: (1) the use of broad-spectrum inhibitors of MMPs, (2) the use of cross-linking agents for silencing the activities of MMPs, (3) ethanol wet-bonding with hydrophobic resin, (4) biomimetic remineralization of water-filled collagen matrix. A combination of these strategies will be able to overcome the limitations in resin-dentin adhesion.

Numerous cases about additional growth of roots or pulp tissue regeneration by using various intracanal medicaments in immature permanent teeth with periapical or pulpal disease have been reported. The underlying mechanism has not been clearly delineated, but it has been widely accepted that undifferentiated mesenchymal cells and stem cells are involved. Moreover, the growth and deposition of osteoid or cementoid tissues have been observed in regenerated pulp and roots. This new and non-invasive treatment has brightened the future of endodontics, and enlarged the vision of regenerative root canal treatment with multi-potent stem cells and various tissue engineering techniques.

During a composite resin restoration, an anticipating contraction gap is usually tried to seal with low-viscosity resin after successive polishing, etching, rinsing and drying steps, which as a whole is called rebonding procedure. However, the gap might already have been filled with water or debris before applying the sealing resin. We hypothesized that microleakage would decrease if the rebonding agent was applied before the polishing step, i.e., immediately after curing composite resin. On the buccal and lingual surfaces of 35 extracted human molar teeth, class V cavities were prepared withthe occlusal margin in enamel and the gingival margin in dentin. They were restored with a hybrid composite resin Z250 (3M ESPE, USA) using an adhesive AdperTM Single Bond 2 (3M ESPE). As rebonding agents, BisCover LV (Bisco, USA), ScotchBond Multi-Purpose adhesive (3M ESPE) and an experimental adhesive were applied on the restoration margins before polishing step or after successive polishing and etching steps. The infiltration depth of 2% methylene blue into the margin was measured using an optical stereomicroscope. The correlation between viscosity of rebonding agents and mciroleakage was also evaluated. There were no statistically significant differences in the microleakage within the rebonding procedures, within the rebonding agents, and within the margins. However, when the restorations were not rebonded, the microleakage at gingival margin was significantly higher than those groups rebonded with 3 agents (p < 0.05). The difference was not observed at the occlusal margin. No significant correlation was found between viscosity of rebonding agents and microleakage, except very weak correlation in case of rebonding after polishing and etching at gingival margin (r = -0.326, p = 0.041).

This study aimed to assess whether the gender of the dental practitioner affects operative techniques in class 2 and class 5 resin composite restorations. In 2008, a nationwide survey was given to Korean dentists. Total 12,193 e-mails were distributed, 2,632 were opened by recipients, and 840 responses were collected. Of the respondents, 78.9% were male and 21.1% were female. The gender distribution in the age groups between respondents and the total population did not differ (p > 0.05). A chi-square test was used to compare technical differences between female and male dentists. A multiple logistic regression analysis was performed to assess the association between gender and operative techniques in resin composite restoration. For class 2 resin composite restoration, female dentists were 1.87 times more likely than male dentists to do multiple incremental fillings (four layers or more) and 2.72 times more likely than males to spend 30 minutes or more for the treatment (p < 0.05). For class 5 resin composite restoration, female dentists were 2.69 times more likely than their male counterparts to use a cavity base or liner, 1.83 times more likely to do multiple incremental fillings (four layers or more) and 1.63 times more likely to spend 20 minutes or more for the procedure (p < 0.05). The gender factor was influential to individual operative techniques in restorative treatment.

The purpose of this study was to assess the current materials, methods and difficulties according to the year of licence and educational background of Korean dentists in Class II direct composite resin restorations.

Total 17 questions were included in the questionnaire. Questions were broadly divided into two parts; first, operator's information, and second, the materials and methods used in Class II posterior composite restoration. The questionnaire was sent to dentists enrolled in Korean Dental Association via e-mail. Total 12,193 e-mails were distributed to dentists, 2,612 e-mails were opened, and 840 mails (32.2%) were received from respondents. The data was statically analyzed by chi-square test using SPSS(v. 12.0.1, SPSS Inc, Chicago, IL, USA).

Male dentists among respondents was 79%. 60.3% of the respondents acquired their licences recently (1998-2007), and 77% practiced in private offices. 83.4% have acquired their knowledge through school lectures, conferences and seminars.

For the Class II restorations, gold inlays were preferred by 65.7% of respondents, while direct composite resin restorations were used by 12.1% amalgam users were only 4.4% of respondents.

For the restorative technique, 74.4% of respondents didn't use rubber dam as needed. For the matrix, mylar strip (53.4%), metal matrix (33.8%) and Palodent system (6.5%) were used. 99.6% of respondents restored the Class II cavity by incremental layering.

Obtaining of the tight interproximal contact was considered as the most difficult procedure (57.2%) followed by field isolation (21%).

Among various bonding systems, 22.6% of respondents preferred SE Bond and 20.2% used Single Bond. Z-250 was used most frequently among a variety of composite resins.

The purpose of this study was to perform quantitative comparisons of water permeable zones in both the adhesive and the hybrid layer before and after thermocycling in order to assess the integrity of the bonding interface. Twenty eight flat dentin surfaces were bonded with a light-cured composite resin using one of four commercial adhesives [OptiBond FL (OP), AdheSE (AD), Clearfil SE Bond (CL), and Xeno III (XE)]. These were sectioned into halves and subsequently cut to yield 2-mm thick specimens; one specimen for control and the other subjected to thermocycling for 10,000 cycles. After specimens were immersed in ammoniacal silver nitrate for 24 h and exposed to a photo developing solution for 8 h, the bonded interface was analyzed by scanning electron microscopy (SEM) and wavelength dispersive spectrometry (WDS) at five locations per specimen. Immediately after bonding, the adhesive layer of OP showed the lowest silver uptake, followed by CL, AD, and XE in ascending order (p < 0.0001); the hybrid layer of CL had the lowest silver content among the groups (p = 0.0039). After thermocycling, none of the adhesives manifested a significant increase of silver in either the adhesive or the hybrid layer. SEM demonstrated the characteristic silver penetrated patterns within the interface. It was observed that integrity of bonding was well maintained in OP and CL throughout the thermocycling process. Adhesive-tooth interfaces are vulnerable to hydrolytic degradation and its permeability varies in different adhesive systems, which may be clinically related to the restoration longevity.

Deterioration of long-term dentin adhesion durability is thought to occur by hydrolytic degradation within hydrophilic domains of the adhesive and hybrid layers. This study investigated the hypothesis that priming the collagen network with an organic solvent displace water without collapse and thereby obtain good bond strength with an adhesive made of hydrophobic monomers and organic solvents. Three experimental adhesives were prepared by dissolving two hydrophobic monomers, bisphenol-A-glycidylmethacrylate (Bis-GMA) and triethylenegly-col dimethacrylate (TEGDMA), into acetone, ethanol or methanol. After an etching and rinsing procedure, the adhesives were applied onto either wet dentin surfaces (wet bonding) or dentin surfaces primed with the same solvent (solvent-primed bonding). Microtensile bond strength (MTBS) was measured at 48 hrs, 1 month and after 10,000 times of thermocycles. The bonded interfaces were evaluated using a scanning electron microscope (SEM). Regardless of bonding protocols, well-developed hybrid layers were observed at the bonded interface in most specimens. The highest mean MTBS was observed in the adhesive containing ethanol at 48 hrs. With solvent-primed bonding, increased MTBS tendencies were seen with thermocycling in the adhesives containing ethanol or methanol. However, in the case of wet bonding, no increase in MTBS was observed with aging.

The objective of this study was to compare dentin shear bond strength (DSBS) of dentin bonding agents (DBAs) cured with a plasma arc (PAC) light curing unit (LCU) and those cured with a light emitting diode (LED) LCU. Optical properties were also analyzed for Elipar freelight 2 (3M ESPE); LED LCU, Apollo 95E (DMT Systems); PAC LCU and VIP Junior (Bisco); Halogen LCU. The DBAs used for DSBS test were Scotchbond Multipurpose (3M ESPE), Singlebond 2 (3M ESPE) and Clearfil SE Bond (Kuraray). After DSBS testing, fractured specimens were analyzed for failure modes with SEM.

The total irradiance and irradiance between 450 nm and 490 nm of the LCUs were different. LED LCU showed narrow spectral distribution around its peak at 462 nm whereas PAC and Halogen LCU showed a broad spectrum. There were no significant differences in mean shear bond strength among different LCUs (P > 0.05) but were significant differences among different DBAs (P < 0.001)

This study was aimed to develop an instrument for real-time measurement of fluid conductance and to investigate the hydrodynamics of dentinal fluid. The instrument consisted of three parts; (1) a glass capillary and a photo sensor for detection of fluid movement, (2) a servo-motor, a lead screw and a ball nut for tracking of fluid movement, (3) a rotary encoder and software for data processing.

To observe the blocking effect of dentinal fluid movement, oxalate gel and self-etch adhesive agent were used. BisBlock (Bisco) and Clearfil SE Bond (Kuraray) were applied to the occlusal dentin surface of extracted human teeth. Using this new device, the fluid movement was measured and compared between before and after each agent was applied.

The instrument was able to measure dentinal fluid movement with a high resolution (0.196 nL) and the flow occurred with a rate of 0.84 to 15.2 nL/s before treatment. After BisBlock or Clearfil SE Bond was used, the fluid movement was decreased by 39.8 to 89.6%.

The purpose of this study was: (1) to compare nanoleakage patterns of a conventional 3-step etch and rinse adhesive system and two experimental hydrophobic adhesive systems and (2) to investigate the change of the nanoleakage patterns after load cycling. Two kinds of hydrophobic experimental adhesives, ethanol containing adhesive (EA) and methanol containing adhesive (MA), were prepared. Thirty extracted human molars were embedded in resin blocks and occlusal thirds of the crowns were removed. The polished dentin surfaces were etched with a 35% phosphoric acid etching gel and rinsed with water. Scotchbond Multi-Purpose (MP), EA and MA were used for bonding procedure. Z-250 composite resin was built-up on the adhesive-treated surfaces. Five teeth of each dentin adhesive group were subjected to mechanical load cycling. The teeth were sectioned into 2 mm thick slabs and then stained with 50% ammoniacal silver nitrate. Ten specimens for each group were examined under scanning electron microscope in backscattering electron mode. All photographs were analyzed using image analysis software. Three regions of each specimen were used for evaluation of the silver uptake within the hybrid layer. The area of silver deposition was calculated and expressed in gray value. Data were statistically analyzed by two-way ANOVA and post-hoc testing of multiple comparisons was done with the Scheffe's test. Silver particles were observed in all the groups. However, silver particles were more sparsely distributed in the EA group and the MA group than in the MP group (p < .0001). There were no changes in nanoleakage patterns after load cycling.

Protein microarray or protein chips is potentially powerful tools for analysis of protein-protein interactions. APin cDNA was previously identified and cloned from a rat odontoblast cDNA library. The purpose of this study was to investigate the APin-protein interactions during ameloblast differentiation. Protein microarray was carried with recombinant APin protein and MEF2, Aurora kinase A, BMPR-IB and EF-hand calcium binding protein were selected among 74 interacting proteins. Immortalized ameloblast cells (ALCs) were transfected with pCMV-APin construct and U6-APin siRNA construct. After transfection, the expression of the mRNAs for four proteins selected by protein micoarrays were assessed by RT-PCR.

The results were as follows:

1. APin expression was increased and decreased markedly after its over-expression and inactivation, respectively.

2. Over-expression of the APin in the ALCs markedly down-regulated the expression of MEF2 and Aurora kinase A, whereas their expression remained unchanged by its inactivation.

3. Expression of BMPR-IB and EF-hand calcium binding protein were markedly increased by the overexpression of the APin in the ALCs, whereas expression of BMPR-IB remained unchanged and expression of EF-hand calcium binding protein was markedly decreased by its inactivation.

These results suggest that APin plays an important role in ameloblast differentiation and mineralization by regulating the expression of MEF2, Aurora kinase A, BMPR-IB and EF-hand calcium binding protein.

This study was aimed to elucidate the biological function of OD314 (Apin protein), which is related toameloblast differentiation and amelogenesis. Apin protein, calcifying epithelial odontogenic (pindborg) tumors (CEOTs)-associated amyloid, were isolated from CEOTs, and has similar nucleotide sequences to OD314. We examined expression of the OD314 mRNA using in-situ hybridization during tooth development in mice. Expression of OD314 and several enamel matrix proteins were examined in the cultured ameloblast cell line up to 28 days by reverse transcription-polymerase chain reaction (RT-PCR) amplification. After inactivation and over-expression of the OD314 gene in ameloblast cell lines using U6 vector-driven RNA interference and CMV-OD314 construct, RT-PCR were performed to evaluate the effect of the OD314 during amelogenesis.

The results were as follows:

1. In in-situ hybridization, OD314 mRNAs were more strongly expressed in ameloblast than odontoblast.

2. When ameloblast cells were cultured in the differentiation and mineralization medium for 28 days, the tuftelin mRNA expression was maintained from the beginning to day 14, and then gradually decreased to day 28. The expressions of amelogenin and enamelin were gradually decreased according to the ameloblast differentiation.

3. Inactivation of OD314 by U6-OD314 siRNA construct down-regulated the expression of OD314, MMP-20, and tuftelin, whereas over-expression of OD314 by CMV-OD314 construct up-regulated the expression of OD314 and MMP-20 without change in tuftelin.

These results suggest that OD314 is considered as an ameloblast-enriched gene and may play the important roles in ameloblast differentiation and mineralization.

The purpose of this study is to evaluate prospectively the effect of different bonding systems and retention grooves on the clinical performance of resin restorations in non-carious cervical lesions (NCCLs). Thirty-nine healthy adults who had at least 2 NCCLs in their premolar areas were included in this study. One hundred and fifty teeth were equally assigned to six groups: (A) Scotchbond Multi-Purpose (SBMP, 3M ESPE, St. Paul, MN, USA, 4th generation bonding system) without retention grooves; (B) SBMP with retention grooves; (C) BC Plus (Vericom Co., Anyang, Gyeonggido, Korea, 5th generation bonding system) without retention grooves; (D) BC Plus with retention grooves; (E) Adper Prompt (3M ESPE, Seefeld, Germany, 6th generation bonding system) without retention grooves; (F) Adper Prompt with retention grooves. All cavities were filled with a hybrid composite resin, Denfil (Vericom Co., Anyang, Gyeonggido, Korea) by one operator. Restorations were evaluated at baseline and at 6-month recall, according to the modified USPHS (United States Public Health Service) criteria. Additionally, clinical photographs were taken and epoxy resin replicas were made for SEM evaluation. At 6-month recall, there were some differences in the number of alpha ratings among the experimental groups. But, despite the differences in the number of alpha ratings, there was no significant difference among the 3 adhesive systems (p > 0.05). There was also no significant difference between the groups with or without mechanical retention (p > 0.05). Follow-ups for longer periods than 6 months are needed to verify the clinical performance of different bonding systems and retention grooves.

This study evaluated the effectiveness and safety of an experimental bleaching strip (Medison dental whitening strip, Samsung medical Co., Anyang, Korea) containing 2.9% hydrogen peroxide. Twenty-three volunteers used the bleaching strips for one and a half hour daily for 2 weeks. As control group, the same strips in which hydrogen peroxide was not included were used by 24 volunteers with the same protocol. The shade change (ΔE*, color difference) of twelve anterior teeth was measured using Shade Vision (X-Rite Inc., S.W. Grandville, MI, USA), Chroma Meter (Minolta Co., Ltd. Osaka, Japan) and Vitapan classical shade guide (Vita Zahnfabrik, Germany). The shade change of overall teeth in the experimental group was significantly greater than that in the control group (p < 0.05) and was easily perceivable. The change resulted from the increase of lightness (CIE L* value) and the decrease of redness (CIE a* value) and yellowness (CIE b* value). The shade change of individual tooth was greatest in canine, and smallest in central incisor. The safety of the bleaching strip was also confirmed.

This study investigated the hypothesis that the dentin bond strength of self-etching adhesive (SEA) might be improved by applying additional layer of bonding resin that might alleviate the pH difference between the SEA and the restorative composite resin. Two SEAs were used in this study; Experimental SEA (Exp, pH: 1.96) and Adper Prompt (AP, 3M ESPE, USA, pH: 1.0). In the control groups, they were applied with two sequential coats. In the experimental groups, after applying the first coat of assigned SEAs, the D/E bonding resin of All-Bond 2 (Bisco Inc., USA, pH: 6.9) was applied as the intermediate adhesive. Z-250 (3M ESPE, USA) composite resin was built-up in order to prepare hourglass-shaped specimens. The microtensile bond strength (MTBS) was measured and the effect of the intermediate layer on the bond strength was analyzed for each SEA using t-test. The fracture mode of each specimen was inspected using stereomicroscope and Field Emission Scanning Electron Microscope (FE-SEM). When D/E bonding resin was applied as the second coat, MTBS was significantly higher than that of the control groups. The incidence of the failure between the adhesive and the composite or between the adhesive and dentin decreased and that of the failure within the adhesive layer increased. According to the results, applying the bonding resin of neutral pH can increase the bond strength of SEAs by alleviating the difference in acidity between the SEA and restorative composite resin.

Ameloblasts are responsible for the formation and maintenance of enamel which is an epithelially derived protective covering for teeth. Ameloblast differentiation is controlled by sequential epithelial-mesenchymal interactions. However, little is known about the differentiation and maturation mechanisms. OD314 was firstly identified from odontoblasts by subtraction between odontoblast/pulp cells and osteoblast/dental papilla cells, even though OD314 protein was also expressed in ameloblast during tooth formation.

In this study, to better understand the biological function of OD314 during amelogenesis, we examined expression of the OD314 mRNA and protein in various stages of ameloblast differentiation using in-situ hybridization and immunohistochemistry.

The results were as follows :

1. The ameloblast showed 4 main morphological and functional stages referred to as the presecretory, secretory, smooth-ended, and ruffle-ended.

2. OD314 mRNA was expressed in secretory ameloblast and increased according to the maturation of the cells.

3. OD314 protein was not expressed in presecretory ameloblast but expressed in secretory ameloblast and maturative ameloblast. OD314 protein was distributed in entire cytoplasm of secretory ameloblast. However, OD314 was localized at the proxiamal and distal portion of the cytoplasm of smooth-ended and ruffle-ended ameloblast.

These results suggest that OD314 may play important roles in the ameloblast differentiation and maturation.

The purpose of this study was to monitor the secretion of matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinase (TIMP) by human periodontal ligament (PDL) fibroblasts stimulated with Prevotella nigrescens lipopolysaccharide (LPS), and to examine the effect of calcium hydroxide treatment on P. nigrescens LPS.

LPS was extracted and purified from anaerobically cultured P. nigrescens. PDL fibroblasts were stimulated by the LPS (0, 0.1, 1, 10 ug/ml) or LPS (10 ug/ml) pretreated with 12.5 mg/ml of Ca(OH)₂ for 3 days, for various periods of time (12, 24, 48 h). Immunoprecipitation were performed for protein level analysis of MMP-1, MMP-2 and TIMP-1. Total RNA was isolated and real-time quantitative polymerase chain reaction (PCR) was performed for quantification of MMP-1 mRNA.

According to this study, the results were as follows:

1. The production of MMP-1 by stimulation with P. nigrescens LPS increased in time-dependent manner, and showed maximum value at 48 h in both protein and mRNA level. But there was no dose-dependent increase.

2. MMP-2 production time-dependently increased when stimulated with 1 and 10 ug/ml LPS, but there was no dose-dependent increase.

3. TIMP-1 production increased to 24 h, but decreased at 48 h. It increased when stimulated with 0.1 and 1 ug/ml LPS, but suppressed at 10 ug/ml.

4. P. nigrescens LPS pretreated with Ca(OH)₂ markedly downregulated MMP-1 gene expression.

A variety files made of stainless steel (S-S) or nickel-titanium (Ni-Ti) are used during endodontic treatment. The purpose of this study was to evaluate the corrosion susceptibility of S-S and Ni-Ti endodontic files. Three brands of files were used for this study: K-flex® S-S files (Maillefer, USA), Profile® Ni-Ti files (Maillefer, USA), K-3® Ni-Ti files (SybronEndo, USA). 120 files of each brands (21mm, ISO size #20) were divided into 12 groups according to 1) sterilization methods using Autoclave or Ethylene Oxide (E-O) gas, 2) Irrigation solutions using 5.25 % NaOCl or Saline, 3) the number of sterilization (1, 5, 10 times). After above procedures, each of the files was inspected by three examiners with a light microscope and camera at X25. Each file was judged and ranked according to the following criteria: 0; no corrosion, 1; mild corrosion, 2; moderate corrosion, and 3; severe corrosion. The files of high score were examined under the Scanning Electron Microscope.

Data were statistically analyzed with the Kruskal-Wallis test (p < 0.05). Most of the ten time-autoclaved files had showed mild to moderate corrosion. But, one or five time-autoclaved files did not show corrosive surface. NaOCl treatment and E-O gas sterilization did not influence on corrosion. There was a significant difference in corrosion susceptibility between sterilization methods and the number of autoclaving. However, there was no significant difference between brands and file materials.

The purpose of this study is to monitor the secretion of matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) produced by human osteosarcoma cell line (MG63) stimulated with Prevotella nigrescens lipopolysaccharides (LPS), and to compare the level of secretion before and after the treatment of calcium hydroxide on P. nigrescens LPS.

LPS was extracted and purified from anaerobically cultured P. nigrescens. MG63 cells were stimulated by the LPS (0, 1, 10 µg/ml) or LPS (10 µg/ml) pretreated with 12.5 mg/ml of Ca(OH)₂ for 3 days.

Total RNA was isolated from the cell, and real-time quantitative polymerase chain reaction (PCR) was performed for quantification of MMP-1 and TIMP-1.

The results were as follows.

1. MMP-1 mRNA expression at 48 hr was highly increased by stimulation with P. nigrescens LPS. The increase was dose-dependent.

2. When stimulated with 1 µg/ml of LPS, TIMP-1 mRNA expression was highly increased at 24 hr and 48 hr. However, TIMP-1 expression was suppressed at higher concentration (10 µg/ml).

3. When P. nigrescens LPS was pretreated with Ca(OH)₂, MMP-1 and TIMP-1 gene expression was downregulated.

The results of this study suggest that transcriptional regulation of MMP-1 and TIMP-1 by P. nigrescens LPS could be one of the important mechanisms in bone resorption of periapical inflammation. The result of calcium hydroxide on MMP-1 and TIMP-1 gene expression suppression shows that calcium hydroxide detoxified bacterial LPS and thus should be used the medication of choice for intracanal dressings in root canal infected with black-pigmented bacteria.

Odontoblasts are responsible for the formation and maintenance of dentin. They are known to synthesize unique gene products including dentin sialophosphoprotein (DSPP). Another unique genes of the cells remain unclear.

OD314 was isolated from the odontoblasts/pulp cells of rats and partially characterized as an odontoblast-enriched gene (Dey et al., 2001). This study aimed to elucidate the biological function of OD314, relating to odontoblast differentiation and dentinogenesis. After determining the open reading frame (ORF) of OD314 by transient transfection analysis using green fluorescent protein (GFP) expression vector, mRNA in-situ hybridization, immunohistochemistry, reverse transcription-polymerase chain reaction (RT-PCR) and western analysis were performed.

The results were as follows:

1. In in-situ hybridization, OD314 mRNAs were expressed in odontoblasts of developing coronal and root pulp.

2. OD314 was a novel protein encoding 154 amino acids, and the protein was mainly expressed in cytoplasm by transient transfection analysis.

3. Mineralized nodules were associated with multilayer cell nodules in the culture of human dental pulp cells and first detected from day 21 using alizarin-red S staining.

4. In RT-PCR analysis, OD314, osteocalcin (OC) and DSPP strongly expressed throughout 28 days of culture. Whereas, osteonectin (ON) mRNA expression stayed low up to day 14, and then gradually decreased from day 21.

5. Western blots showed an approximately 17 kDa band. OD314 protein was expressed from the start of culture and then increased greatly from day 21.

In conclusion, OD314 is considered as an odontoblast-enriched gene and may play important roles in odontoblast differentiation and dentin mineralization.