Effects of low level of calcium on the biological behavior of rat primary ameloblasts cultured in vitro / 西安交通大学学报(医学版)

【Objective】 To explore the effect of low levels of calcium on the biological characteristics of ameloblasts. 【Methods】 Rat primary ameloblasts were cultured in standard DMEM medium. After five days they were identified by RT-PCR and immunohistochemistry. Then 0, 0.6 and 1.2 mmoL/L Ca2+ and 100 mL/L fetal bovine serum were added into DMEM medium without calcium. After 48 hours, the cell morphology was observed by inverted microscope. The proliferation and apoptosis of cells were separately examined by MTT and AnnexinV-PI. Real-time quantitative PCR was used to detect the expression levels of amelogenin and KLK4 mRNA. 【Results】 After Five days in standard DMEM medium, the cells were shaped like the paving pattern. RT-PCR showed that both amelogenin and KLK4 were expressed in the cells. Immunohistochemical staining showed that most cells had positive staining for amelogenin. After 48 hours of calcium intervention, some cells in 1.2 mmoL/L Ca2+ group had higher nuclear density and poor light transmittance, and more high columnar cells could be observed in 1.2 mmoL/L Ca2+ group than those in 0 and 0.6 mmoL/L Ca2+ groups. With the decrease in calcium concentration in the medium, MTT showed that the proliferation activity of ameloblasts reduced (P<0.01). Annexin V-PI showed that the percentage of apoptotic cells decreased, and there was a significant difference between 1.2 mmoL/L and 0 mmoL/L Ca2+ groups (P<0.05). Real time-PCR showed that the expressions of amelogenin and KLK4 mRNA reduced (P<0.01). 【Conclusion】 Low-level calcium may inhibit the differentiation of ameloblasts, thereby affecting the formation of enamel mineralization.

The effect of inhibiting p38 MAPK on the expression of genes related to enamel development in mice / 口腔疾病防治

Objective@#To study the effect of p38 mitogen activated protein kinase (p38 MAPK) on the expression of genes related to enamel development in the enamel epithelium and to provide a basis for the study of the molecular mechanism of enamel development.@*Methods@#The p38 MAPK-specific inhibitor SB203580 dissolved in DMSO was added to the culture medium of mouse mandibular molar tooth germs in vitro as experiment group, and mouse mandibular molar tooth germs treated with the same amount of DMSO were used as control group. Western blot was used to detect the protein expression level of phosphorylated p38 (p-p38) in the enamel epithelium. Real-time PCR was used to detect the mRNA expression levels of runt-related transcription factor 2 (Runx2), osteoblast-specific transcription factor (Osx), ameloblast markers odontogenic ameloblast associated protein (ODAM), amelotin (AMTN), matrix metalloproteinase 20 (MMP20) and kallikrein 4 (KLK4) in the enamel epithelium. @*Results @# Western blot results showed that under the action of the inhibitor SB203580, the phosphorylation level of p38 MAPK in mouse enamel epithelium decreased, and the difference was statistically significant (P < 0.05). Real-time PCR results showed that the expression levels of the transcription factors Runx2 and Osx and the ameloblast markers ODAM, AMTN, MMP20, and KLK4 in the SB203580 group were lower than those in the control group, and the difference was statistically significant (P < 0.05).@*Conclusion@#The p38 MAPK signaling pathway can mediate enamel development by regulating the expression of the transcription factors Runx2 and Osx and the ameloblast markers ODAM, AMTN, MMP20 and KLK4 in the mouse enamel epithelium.

The C-terminus of the amelogenin peptide promotes the proliferation of alc ameloblasts through accelerating cell cycle / 中国组织工程研究

BACKGROUND: C-terminus of the amelogenin peptide (AMG-CP) is a small molecular endogenous peptide that is highly shown that AMG-CP can regulate the proliferation and differentiation of cementoblasts, bone marrow mesenchymal stem cells and periodontal ligament fibroblasts, but the biological function of AMG-CP on ameloblasts has not been elucidated. O conserved among species. It is involved in important physiological processes during tooth development. Some studies have BJECTIVE: To investigate the effects of different concentrations of AMG-CP on the proliferation of ALC ameloblasts and its underlying mechanisms. METHODS: AMG-CP was successfully synthesized and determinated by liquid chromatography and mass spectrometry. The effects of AMG-CP at 0, 0.5, 1, 2 mg/L on the proliferation of ALC ameloblasts were observed by xCELLigence RTCA cell analysis system in real time. The effect of AMG-CP at 0, 1, 2 mg/L on cell cycle of ALC was detected by flow cytometry. Real-time PCR was used to detect the expression of cyclin D1, CDK4, MCM2, MCM5 mRNA in ALC cells treated with AMG-CP at 0, 1, 2 mg/L. Western blot was carried out to evaluate the effect of AGM-CP at 0, 1 mg/L on MAPK-ERK1/2 pathway by detecting the expression of phosphorylated ERK1/2 and total ERK1/2 in ALC cells. Pathway blockade assay was performed by using ERK1/2 blocker U0126 to pretreat ALC cells. Then cell proliferation ability as well as phosphorylated ERK1/2 expression was analyzed by xCELLigence RTCA cell analysis system and western blot. RESULTS AND CONCLUSION: Compared with the control group, AMG-CP promoted the proliferation of ALC cells, and decreased the population doubling time in a dose-depending manner. Flow cytometry detected the acceleration of cell cycle after treatment with AMG-CP. The results of Real-time PCR showed that AMG-CP upregulated cell cycle-related genes (cyclin D1, CDK4, MCM2, MCM5) expression. Western blot results showed that AMG-CP could upregulate the expression of phosphorylated ERK1/2 and activate MAPK-ERK1/2 signaling pathway in ALC cells. After U0126 was used to inhibit the MAPK-ERK1/2 pathway, the ability of AMG-CP promoting ALC proliferation was inhibited. These results suggest that AMG-CP has a potential to activate MAPK-ERK1/2 pathway, accelerate the process of cell cycle, and then promote the proliferation of ALC cells, all of which indicate that AMG-CP has the potential to promote the proliferation of ameloblasts.

The effects of varies osmotic pressure induced by adding fluoride into the medium on the biological characteristic of ameloblast / 西安交通大学学报(医学版)

Objective: To explore the effects of varies osmotic pressure induced by adding fluoride into the medium on the biological characteristic of ameloblast.Methods: Ameloblasts were cultured in vitro on the fifth day were seeded and purified. DMEM medium containing different concentrations of NaF (0, 42, 84 mg/L), NaCl (58.5 and 117 mg/L), glycerol (0.05 and 0.09 mg/L) and sorbitol (1 and 1.5 mg/L) was added into the cells, which were then cultured for 48 hours. The survival and proliferation of ameloblasts in different groups were detected by MTT. The apoptosis rate was measured by flow cytometry and the expressions of CK14 and KLK4 by Real-time PCR. Results: MTT showed that with the increase of osmolality, cell proliferation decreased; however, the most obvious one was in NaF group. Streaming apoptosis detection showed that with the increase of osmolality, the apoptosis rate of cells increased, which was most obvious in NaF group. Real-time PCR showed that the mRNA expressions of CK14 and KLK4 decreased with the increase of osmolality, and that they were lower in NaF 42 mg/L group than in the other control groups. Results: The change in osmotic pressure of the medium affects the proliferation, apoptosis and mRNA expression of ameloblasts. The biological change of ameloblasts cultured in medium containing different concentration of NaF is mainly due to the toxic effects of fluoride ions.

Expressions of cell polarity related proteins CDC42and PAR3 during tooth germ development in mice / 吉林大学学报(医学版)

Objective:To study the expressions of cell polarity related proteins CDC42 and PAR3 during tooth germ development in the mice,and to discuss their possible roles during tooth development in the mice.Methods:The whole heads were obtained from the mouse embryo on the days 13.5,14.5,16.5 and 18.5 (E13.5,E14.5,E16.5 and E18.5) and the mice on the postnatal days 1 (PN1) and 5 (PN5).The tissues were fixed in paraformaldehyde,decalcified,dehydrated,embedded in paraffin,and sectioned.The histology of tooth germ was observed by HE staining.The expressions of CDC42 and PAR3 during tooth germ development were detected by immunohistochemistry staining.Results:The HE staining results showed that E13.5,E14.5,E16.5 and E18.5were the bud stage,the cap stage,the early and the late bell stage of tooth germ development,respectively;the tooth germ of PN1 mice showed the matured odontoblasts and ameloblasts;the tooth germ of PN5 mice showed the completed tooth crown development.The immunohistochemistry staining results showed that CDC42 expressed in the tooth germ of the mice at E13.5,E14.5 and E16.5;the CDC42 expression at E 18.5 was reduced compared with E13.5,E14.5 and E16.5;CDC42 mainly expressed in the odontoblasts and ameloblasts of the tooth germ of the mice at PN1 and PN5;PAR3 weakly expressed in the tooth germ of the mice at E13.5 and E14.5,and it was increased at E16.5 and E18.5.At PN1 and PN5,the expressions of PAR3 were decreased compared with E18.5.Conclusion:CDC42 and PAR3 partieipat in the mouse tooth development;during the early stage of tooth germ development,they may be involved in the proliferation and migration of mouse dental germ;during the late stage,CD42 and PAR3 may be involved in the differentiation of the odontoblasts and the ameloblasts,especially in the establishment and maintenance of cell polarity.

Expressions of cell polarity related proteins CDC42 and PAR3 during tooth germ development in mice / 吉林大学学报(医学版)

Objective: To study the expressions of cell polarity related proteins CDC42 and PAR3 during tooth germ development in the mice, and to discuss their possible roles during tooth development in the mice. Methods: The whole heads were obtained from the mouse embryo on the days 13. 5, 14. 5, 16. 5 and 18. 5 (E13. 5, E14. 5, E16. 5 and E18. 5) and the mice on the postnatal days 1 (PN1) and 5 (PN5). The tissues were fixed in paraformaldehyde, decalcified, dehydrated, embedded in paraffin, and sectioned. The histology of tooth germ was observed by HE staining. The expressions of CDC42 and PAR3 during tooth germ development were detected by immunohistochemistry staining. Results: The HE staining results showed that E13. 5, E14. 5, E16. 5 and E18. 5 were the bud stage, the cap stage, the early and the late bell stage of tooth germ development, respectively; the tooth germ of PN1 mice showed the matured odontoblasts and ameloblasts; the tooth germ of PN5 mice showed the completed tooth crown development. The immunohistochemistry staining results showed that CDC42 expressed in the tooth germ of the mice at E13. 5, E14. 5 and E16. 5; the CDC42 expression at E 18. 5 was reduced compared with E13. 5, E14. 5 and E16. 5; CDC42 mainly expressed in the odontoblasts and ameloblasts of the tooth germ of the mice at PN1 and PN5; PAR3 weakly expressed in the tooth germ of the mice at E13. 5 and E14. 5, and it was increased at E16. 5 and E18. 5. At PN1 and PN5, the expressions of PAR3 were decreased compared with E18. 5. Conclusion: CDC42 and PAR3 participat in the mouse tooth development; during the early stage of tooth germ development, they may be involved in the proliferation and migration of mouse dental germ; during the late stage, CD42 and PAR3 may be involved in the differentiation of the odontoblasts and the ameloblasts, especially in the establishment and maintenance of cell polarity.

Expression and function of microRNAs in enamel development / 华西口腔医学杂志

microRNAs (miRNAs) are endogenous short, noncoding RNAs that can negatively regulate gene expression post-transcriptionally. miRNAs are involved in multiple developmental events in various tissues and organs, including dental enamel development. Any disruption during enamel development may result in inherited enamel malformations. This article reviews the expression and function of miRNAs in enamel development.

The effects of fluoride on proliferation and apoptosis of primary cultured rat ameloblast / 中华地方病学杂志

Objective To investigate the effects of fluoride at different concentrations on proliferation and apoptosis of primary rat ameloblast in vitro.Methods Ameloblasts were isolated from tooth germ of 4 days SD rat maxillomandibular molar and cultured in vitro.Cells were treated with NaF at 0.0 (control group),0.4,0.8,1.6,3.2 and 6.4 mmol/L for 24,48 and 72 h,respectively.Inverted microscope was used to observe cell morphology;immunochemistry method was used to identify ameloblasts;3-(4,5-dimethylthiazole-2)-2,5-diphenyl tetrazolium bromide (MTT) assay was applied to measure cell viability at each time point.The cells were treated with 1.6 mmol/L NaF for 24 and 48 h,or after 50 mol/L caspase pan-inhibitor Z-VAD-FMK pretreatment 1 h,1.6 mmol/L NaF treatment for 48 h.Cell apoptosis was then tested by flow cytometry.In addition,activation of caspase-3 and poly (ADP-ribose) polymerase (PARP) were assessed by Western blotting to explore potential involvement of caspase activation in NaF-induced apoptosis.All data analysis was performed using SigmaStat V 3.5 software.Results ①Primary rat ameloblasts were in polygonal shape at low density and appeared like paving stone at high density with obvious nucleus,showing typical morphological characteristics of cells with epithelial origin.②The results of immunochemistry assay indicated that the cultured cells were positive in cytokeratin 14 (CK14) and ameloblastin (AMBN) staining,in accordance with the immunocytochemical characteristics of ameloblasts.③The effects of NaF on ameloblast proliferation were in a dose-and time-dependent manner.For low dose NaF (0.4 and 0.8 mmol/L) groups,cells treated for 24 h had significantly higher cell proliferation rates than that of the control group (0.0 mmol/L,P ＜0.05),the proliferation indexes for 0.0,0.4 and 0.8 mmol/L groups were 1.00 ± 0.00,1.38 ± 0.11 and 1.29 ± 0.13,respectively;the same doses of NaF had no obvious influence on cell proliferation at 48 h (1.00 ± 0.00,1.16 ± 0.14 and 0.94 ± 0.07,P ＞ 0.05);cell proliferation indexes at 72 h were significantly lower than that of the control group (0.87 ± 0.03 and 0.80 ± 0.04,P ＜ 0.05).Medium dose of NaF (1.6 mmol/L) did not cause obvious alterations in cell proliferation at 24 h (0.90 ± 0.08,P ＞ 0.05);while cell proliferation indexes at 48 and 72 h were obviously reduced than that of the control group (0.38 ± 0.03 and 0.26 ± 0.04,P ＜ 0.01).For high NaF concentration (3.2 and 6.4 mmo]/L) groups,cell proliferation indexes were significantly decreased at all time points compared with control cells,the rates for 3.2 mmol/L groups were 0.57 ± 0.14,0.08 ± 0.03 and 0.00 ± 0.00,respectively,and the rates for 6.4 mmol/L groups were 0.11 ± 0.04,0.00 ± 0.00 and 0.00 ± 0.00,respectively (P ＜ 0.01).④Flow cytometry was used to detect apoptosis.The results showed that treatment with 1.6 mmol/L NaF resulted in significantly increased apoptosis in ameloblasts at both 24 h [(5.80 ± 2.03)％] and 48 h [(17.45 ± 4.97)％] compared to the control group [(2.59 ± 0.95)％,P ＜ 0.05].In cells pre-treated with pan-caspase inhibitor Z-VAD-FMK,NaF-induced apoptosis was significantly lower than that of cells treated with only 1.6 mmol/L NaF [(9.43 ± 3.79)％ vs (18.26 ± 3.39)％,P ＜ 0.05].⑤Cleavage of caspase-3 and PARP was detected in ameloblasts treated with 1.6 mmol/L NaF for 48 h.Conclusion Overdose fluoride could inhibit proliferation and induce apoptosis via activation of caspase cascade in primary cultured rat ameloblasts.

Establisihment of a fluorosis model induced by coal burning in ameloblasts of rat offspring / 中华地方病学杂志

Objective To establish a fluorosis model induced by coal burning and in ameloblasts of rat offsprings.Methods High fluoride air model was established based on the burning coal habit of the epidemic areas.Fluoride feed was made of corn dried by coal burning.Thirty-six SD rats were divided into 3 groups by random number table method according to body weight in a male and female ratio of 2 ∶ 1∶ in the high fluoride air room and rats were feed food with fluorine at 40 mg/kg (high fluoride group),25 mg/kg (low fluoride group);in the normal air room and rats were feed food with fluorine at 0 mg/kg (the control group),12 rats in each group.Mating litter in a ratio of 2 ∶ 1 at the end of 8 weeks.The offsprings were killed on postnatal day 3 and 7 to make mandible sections.Specimens were prepared for light microscope examination to observe the morphological changes of ameloblasts in the tooth germ.Results At the end of 0,2,4,6 and 8 weeks,serum fluoride of the high fluoride group were (0.031 ± 0.003),(0.060 ± 0.006),(0.085 ± 0.006),(0.110 ± 0.007) and (0.134 ± 0.008) mg/L;serum fluoride of the low fluoride group were (0.031 ± 0.003),(0.046 ± 0.005),(0.077 ± 0.006),(0.091 ± 0.007) and (0.104 ± 0.007) mg/L;serum fluoride of the control group were (0.030± 0.003),(0.037 ± 0.002),(0.044 ± 0.002),(0.046 ± 0.003) and (0.049 ± 0.003) mg/L.At the end of 2,4,6 and 8 weeks,serum fluoride of high fluoride group and low fluoride group were significantly higher than that of control group (all P ＜ 0.05).At 7 d,offspring rats in high fluoride group,adamantoblasts were in distortions and vacuole changes,but offspring rats in low fluoride group and the control group had no abnormality.Conclusion By providing rat with high fluoride air and food,we could establish a fluorosis model induced by coal burning in ameloblasts of rat offsprings.

The effect of fluoride on the viability of rat ameloblast HAT-7 cells and Ca2 + concentration in the cells / 实用口腔医学杂志

Objective:To evaluate the effect of fluoride on the viability of rat ameloblast HAT-7 cells and calcium concentration in the cells.Methods:HAT-7 cells were exposed to NaF at 0,0.4,0.8,1.6,3.2 and 6.4 mmol/L for 24,48 and 72 h respectively. CCK-8 assay was performed to examine the cells proliferation;the apoptosis rate was determined by flow cytometry;Ca2 +concentration in the cells was detected by laser scanning confocal microscopy.Results:The cell proliferation was increased by NaF at 0.4 mmol/L and 0.8 mmol/L,whereas inhibited at 1.6 mmol/L and above.The effects were in a time-dependent manner.NaF increased apoptosis of the cells and increased Ca2 + concentration in the cells in a concentration-dependent manner.Conclusion:Fluoride at low doses promotes proliferation,at high doses inhibits proliferation of HAT-7 cells.NaF of 1.6 mmol/L or more induces apoptosis of HAT-7 cells and in-duce Ca2 + overloading in the cells.

Ameloblast morphology and ultrastructure changes of dental fluorosis in rats / 中华地方病学杂志

Objective To observe the ameloblast morphology and ultrastructure changes of dental fluorosis in rats.Methods Forty SD rats were divided into experimental group and control group (20 rats in each group) by body weight using random number table method.Rats in experimental group were given drinking water with 50 mg/L fluoride to establish dental fluorosis model,and rats in control group were given drinking water without fluoride.All rats were killed at the 56th day,then ameloblast morphology and ultrastructure were observed by means of HE staining and transmission electron microscopy (TEM) at secretory,transition,maturation and post-maturation stages.Results Rats in experimental group showed typical symptoms of dental fluorosis,and lower incisors presented brown and white stripes on enamel surface,chalk color patches and other typical dental fluorosis changes,rats in control group no abnormality.HE staining results showed that differences were not observed between the two groups at the secretory and transformation stages,and few distortions and interstitial space widened were found in the experimental group at maturation and post-maturation stages.TEM displayed ultrastructure changes of ameloblasts,and no differences were observed between the two groups at secretory and transformation stages.Apoptosis was observed in some ameloblasts at the maturation stage in the experimental group,such as rough endoplasmic reticulum expansion,swelling,some ribosome particles exfoliating from endoplasmic reticulum,mitochondrial swelling and disappearance of crista,some nuclear membrane broken down,chromatin condensation and karyolysis.At post-maturation stage,intercellular space of the experimental group was wider than that of the control group.Conclusions Morphological and ultrastructure damage of ameloblasts may have occurred at the maturation stage in the process of dental fluorosis.Ameloblasts may be more sensitive to fluoride in dental enamel formation at maturation stage.

Smad4 Mediated TGF-beta/BMP Signaling in Tooth Formation Using Smad4 Conditional Knockout Mouse

Carcinoma ameloblástico / Ameloblastic Carcinoma

Se presenta el caso clínico de una paciente con antecedentes de gastritis, que acudió al consultorio médico de su área de salud por presentar aumento de volumen en paladar, con crecimiento progresivo, acompañado de molestias para masticar y deglutir, la cual fue remitida al Servicio de Cirugía Maxilofacial del Hospital Clinicoquirúrgico Docente "Dr. Joaquín Castillo Duany" de Santiago de Cuba. Los resultados de los exámenes complementarios, incluida la biopsia incisional en paladar, confirmaron que se trataba de un carcinoma ameloblástico. Se indicó seguimiento clínico en el Hospital Oncológico Docente "Conrado Benítez" de la mencionada ciudad.

The case report of a patient with history of gastritis that visited the doctor's office of her health area due to an increase of volume in palate, with progressive growth, accompanied by problems to chew and to swallow is presented. She was referred to the Maxillofacial Surgery Service of "Dr. Joaquín Castillo Duany" Teaching Clinical Surgical Hospital in Santiago de Cuba. The results of the additional tests, including the incisional biopsy in palate, confirmed that it was an ameloblastic carcinoma. Clinical follow-up in "Conrado Benítez" Teaching Oncological Hospital in Santiago de Cuba was suggested.

Expression of Frizzled Protein in Mouse Tooth Germ at Bell Stage / 中国医科大学学报

Objective To observe the expression of Frizzled protein in mouse tooth germ at bell stage and explore the possible function of Wnt/Frizzled signal molecules. Methods Mouse embryos at 17 and 19 days of gestation (E17 and E19) as well as mice postnatal day 2 (PN2) were employed in our study. The frozen sections of the first lower molar were made and indirect immunofluorescence technique was carried out on the sections. The location of Frizzled protein was observed under the fluorescence microscope. Results Frizzled protein was expressed weakly in the inner- and outer- dental epithelium at E17. At E19,the positive staining was found to distribute at the summit of both pre-ameloblasts and pre-odontoblasts. At P2,Frizzled protein was expressed strongly at the summit of ameloblasts and odontoblasts. Conclusion Wnt/Frizzled signal molecules may be involved in the differentiation of ameloblasts and odontoblasts.

A study of APin-protein interactions using protein microarray / 대한치과보존학회지

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.

Expression and function of OD314, Apin protein during ameloblast differentiation and amelogenesis / 대한치과보존학회지

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.

Expression of OD314 during ameloblast differentiation and maturation / 대한치과보존학회지

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.

Effect of short exposure to high levels of fluoride on cellular structural and the expression of BMP-4 in ameloblasts of mouse molar / 实用口腔医学杂志

Objective:To study the effect of short exposure to high levels of fluoride on structure and the BMP-4 expression of ameloblasts of mouse molar and explore the possible formation mechanisms of dental fluorosis. Methods:32 4-day-old ICR mice were randomly divided into two groups. The experimental animals and the control animals were shared equal numbers in each group. The experimental animals were received a single intraperitoneal dose of 20 (n=8) and 10 (n=8) mg NaF/kg(body weight) respectively,equal doses of NaCl were given to the controls (n=8 for each group). The injected volume was kept constant (10 ?l/g). After 24 h all mice were sacrificed. HE and immunohistochemical stainings were used to observe the structural changes and the expression of BMP-4 in ameloblasts of mouse molar at different stages. SPSS 13.0 software package was used to analyze the data. Results:The early and late secretory/transitional ameloblasts were sensitive to a short-lasting but high dose of F-. The immunohistochemistry results demonstrated that in experimental animals,the expression of BMP-4 was weaker than that in control animals with the significant differences(P

The effect of fluoride on the secretory ameloblast apoptosis / 西安交通大学学报(医学版)

Objective To elucidate the relationship between f luoride and cell apoptosis of ameloblasts and to further study the mechanism of dental fluorosis. Methods By establishing an experimental m odel of fluoride intoxication in pregnant C57BL/6N, we observed apoptosis in the fetal secretory ameloblasts in mice by means of transmission electron microscop e and TUNEL. Results High fluoride exposure in pregnant mic e could cause subameloblastic cysts in first lower molar of fetal mice. Programm ed cell death (PCD) occurred in the disturbed ameloblasts above cysts. Conclusion Intoxicat ion mechanism of fluoride on ameloblasts may occur by means of PCD.