OBJECTIVETo observe the effects of nicotine on the proliferation of odontoblasts and explore the possible mechanism.

METHODSOdontoblasts MDPC-23 were cultured, inoculated and divided into two groups randomly. With no stimuli added for the control group, the experimental group was stimulated by 100 microg/mL nicotine. After 8 hours, 10 micromol/L BrdU was added to label cells at S stage in cell cycle. 24 hours later, odontoblasts were fixed and immunofluorescence staining was performed with specific mouse BrdU antibody. After counterstaining with propidium iodide, BrdU positive cells were arbitrarily scored microscopically by an independent estimation conducted three times, and the corresponding total cell number in the same vision were counted in both groups. BrdU positive cell rates were calculated and compared statistically. At the same time, odontoblasts MDPC-23 were cultured and stimulated by 100 microg/mL nicotine, the dynamic Ca2+ concentration inside the cytoplasm were detected immediately by a confocal laser scanning microscope.

RESULTSThe ratio of S stage cells in the experimental group was 36.3% significantly lower than that (48.2%) in the control group. After the addition of 100 microg/mL nicotine, the Ca2+ concentration inside the cytoplasm rose rapidly, sustained at a high level for a short time and then relapsed gradually.

CONCLUSIONNicotine had inhibitory effects on the proliferation of odontoblasts MDPC-23, which might be related to the increased Ca2+ concentration in the cytoplasm.

The polarity of ameloblasts and odontoblasts is crucial for their differentiation and function. Polarity-related molecules play an important role in this process. This review summarizes the process of polarity formation of ameloblasts and odontoblasts and their related regulators.
Ameloblasts ; Cell Differentiation ; Odontoblasts

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.
Ameloblasts* ; Amelogenesis ; Cytoplasm ; Dental Enamel ; Immunohistochemistry ; Odontoblasts ; RNA, Messenger ; Tooth

OBJECTIVEStudies have showed that L type calcium channel plays an important role in dentin calcification and affects tooth development and tooth reparation after injury. The objective of this article is to study the effects of nimodipine, blocking agent of L type calcium channel, on human dentinogenesis using human tooth slice organ culture in vitro.

METHODSYoung healthy human premolars were collected, and cut into 2 mm-thick transverse slices by low speed diamond saw. Agarose beads dipped in nimodipine solution and PBS weresy minetrically placed on tooth slices, and the slices were then embedded in a semisolid agarose-based medium and cultured with organ culture method for 1 week. Fluorescent band of tetracycline, Von-Kossa staining, immunohistochemical staining of the slices and transmission electron microscopy (TEM) of odontoblasts were observed to evaluate dentinogenesis changes of the slices.

RESULTSTooth slices were successfully cultured in vitro for 1 week and the odontoblasts could maintain their original morphology. After treatment with nimodipine, the fluorescent band of tetracycline was narrow and weak, and globular calcification in predentine was decreased compared with the control. TEM showed that secretory vesicles in odontoblast were somewhat increased, hut iminunohistochemical staining for collagen I showed no difference between the two groups.

CONCLUSIONNimodipine can influence the calcification of dentine, but has no obvious influence on the synthesis and secretion of dentine matrix. The results show that L type calcium channel is important in dentin calcification.

The effects of chitosan upon the experimentally induced differentiation of MDPC-23 cells, derived from mouse dental papilla cells, were investigated by RT-PCR, observations of cell morphology and Alizaline red-S staining. Chitosan was found to significantly increase and accelerate the expression of ALP mRNA but decrease the ColI transcript levels, as compared with the control, in a time-dependent manner during the differentiation of MDPC-23 cells. Chitosan also significantly downregulated ON mRNA expression and accelerated mineralization in differentiating MDPC-23 cells. These results suggest that chitosan facilitates odontoblast differentiation and mineralization and may have potential clinical applications as a dentin regeneration material.
Animals ; Chitosan ; Dental Papilla ; Dentin ; Mice ; Odontoblasts ; Regeneration ; RNA, Messenger

OBJECTIVEThe Sonic hedgehog signalling peptide has been demonstrated to play important roles in the growth and patterning of the tooth development. This study aims on whether the antagonist beta-transduction repeat containing protein of Sonic hedgehog signal transduction expressed in tooth germs ameloblast and odontoblast or not.

METHODSThe mouse embryo heads of different developmental stages of E10.5, E13.5, E14.5, E16.5, E18.5 and P0, P3, P6 after birth were acquired fixed with 4% paraformaldehyde for 48 hours, embeded with Paraffin and examined using LsAB (labelled streptavidin-biotin) method to observe the beta-TrCP expression pattern in tooth germs, ameloblast and odontoblast.

RESULTSIt was demonstrated that beta-TrCP expressed in oral epithelium, tooth bud, mesenchymal cell cytoplasm of ameloblast and odontoblast of different stage of tooth development.

CONCLUSIONbeta-TrCP expressed from early stage to later stage of murine tooth development. And these findings provide the evidence of antagonist regulatory pathways for shh in teeth development.

The ultimate goal of a regenerative pulp treatment strategy is to reconstitute normal tissue continuum at the pulp-dentin border, regulating tissue-specific processes of reparative dentinogenesis. However, little is known about the molecular mechanism of reparative dentinogenesis. The purpose of this study was to investigate the pulpal response after direct pulp capping and pulpotomy with mineral trioxide aggregate (MTA) by histological and immunohistochemical studies. There was continuous reparative dentin bridge formation at 2 weeks after treatment with MTA in both the pulp capping and the pulpotomy groups. The cells in the pulp capping group showed typical odontoblast characteristics, while the cells of reparative dentin in pulpotomy group were round in shape, lost their polarity, organized as a sheet of cells, and trapped in osteodentin-like mineralized tissue. In pulp capping group, upper layer of the reparative dentin showed cell lacunae indicating osteoblastic characteristics, whereas lower layer of the reparative dentin contained predentin and dentinal tubule-like structures as normal dentin. However, there was osteodentin formation in pulpotomy group. DSP protein was expressed at 4 weeks in odontoblasts of pulp capping group, while BSP was expressed at 4 weeks after pulpotomy. These results suggest that two different types of reparative dentin formation, dentin-like and bone-like dentin, may depend on the type and extent of the injury and the effect of the associated defense reaction on the structural and functional integrity at the dentin-pulp border.
Dental Pulp Capping* ; Dentin* ; Dentinogenesis ; Odontoblasts ; Osteoblasts ; Pulpotomy* ; Pemetrexed

Odontoblasts are responsible for the formation and maintenance of dentin which is a mineralized part in dentin-pulp complex of tooth. OD314 was obtained by subtractive hybridization between odontoblasts and osteoblast/dental papilla cells, and differentiatially expressed in the odontoblasts but not in osteoblasts and dental papilla cells. In this study, to better understand the biological function of new odontoblast-enriched gene, OD314, we examined expression of OD314 in cultured MDPC-23 cells and intracellular localization of OD314 protein. We also evaluate the effect of OD314 over-expression and inactivation on the cells by northern analysis. When MDPC-23 cells are cultured in the differentiation and mineralization medium for 28 days, OD314 mRNA expression was gradually increased from the beginning to day 21 and remained relatively high on day 28. Immunofluorescent staining of cultured MDPC-23 revealed localization of OD314 on the cytoplasm, especially near the nuclear membrane. However, a small amount of fluorescence was also observed in the nucleus. Inactivation of OD314 by RNA interference up-regulated the expression of DSPP, whereas over-expression of OD314 by CMV-OD314 plasmid down-regulated the expression of ON. These results suggest that OD314, a odontoblat-enriched gene, may play important roles in the odontoblast differentiation and dentin mineralization.
Cytoplasm ; Dental Papilla ; Dentin ; Fluorescence ; Nuclear Envelope ; Odontoblasts* ; Osteoblasts ; Plasmids ; RNA Interference ; RNA, Messenger ; Tooth

Tooth development involves bud, cap, bell and hard tissue formation stages, each of which is tightly controlled by regulatory molecules. The aim of this study was to identify genes that are differentially expressed during dental hard tissue differentiation. Sprague-Dawley rats at postnatal days 3, 6 and 9 were used in the analysis. Differential display RT-PCR (DD-PCR) was used to screen differentially expressed genes between the 2nd (root formation stage, during mineralization) and 3rd (cap stage, before mineralization) molar germs at postnatal day 9. The DNA detected in the 2nd molar germs showed homology to osteonectin only (GenBank accession no. NM_012656.1). The level of osteonectin mRNA expression was much higher in the 2nd molar germs than in the 3rd molar germs and was found to increase in a time-dependent manner from the early bell stage to the root formation stage in the 2nd molar germs. The pattern of osteonectin protein expression was consistent with these RT-PCR results. Osteonectin protein was found by immunofluorescent analysis to localize in odontoblasts and preodontoblasts rather than the dentin matrix itself. Further studies are needed to validate the involvement of osteonectin in mineralization and root formation.
Animals ; Dentin ; DNA ; Molar ; Odontoblasts ; Osteonectin ; Rats ; Rats, Sprague-Dawley ; RNA, Messenger ; Tooth

Nuclear factor I-C (NFI-C) plays a pivotal role in various cellular processes such as odontoblast and osteoblast differentiation. Nfic-deficient mice showed abnormal tooth and bone formation. The transplantation of Nfic-expressing mouse bone marrow stromal cells rescued the impaired bone formation in Nfic(-/-) mice. Studies suggest that NFI-C regulate osteogenesis and dentinogenesis in concert with several factors including transforming growth factor-β1, Krüppel-like factor 4, and β-catenin. This review will focus on the function of NFI-C during tooth and bone formation and on the relevant pathways that involve NFI-C.
Animals ; Bone Development* ; Dentinogenesis ; Mesenchymal Stromal Cells ; Mice ; NFI Transcription Factors* ; Odontoblasts ; Osteoblasts ; Osteogenesis ; Osteoporosis ; Tooth*