过量氟对大鼠切牙牙本质涎蛋白mRNA表达的影响

目的研究过量氟对大鼠切牙发育过程中牙本质涎蛋白(dentin sialoprotein,DSP)mRNA表达的影响。方法选择6只Wistar大鼠,随机分成两组:对照组常规饲料喂养,自由饮用蒸馏水;实验组常规饲料喂养,自由饮用氟离子浓度为100 mg/L的氟化水,建立大鼠氟斑牙模型。饲养8周处死动物,提取大鼠切牙组织总RNA,逆转录为cDNA,利用实时荧光定量聚合酶链反应技术观察过量氟对大鼠切牙中DSP mRNA表达的影响。结果实时荧光定量聚合酶链反应结果显示DSP mRNA在实验组表达水平明显高于对照组表达水平(t=2.132,P<0.01)。结论过量氟可能通过增强DSP mRNA的表达,影响牙本质的发育矿化。     

短期高浓度氟对小鼠磨牙胚中牙本质涎蛋白表达的影响

目的:研究短期高浓度氟对小鼠磨牙成牙本质细胞形态及牙本质涎蛋白(dentin sialoprotein, DSP)表达的影响,探讨氟对牙本质发育的作用机制.方法:选择4 d龄的ICR小鼠共32 只,随机分为2 组,每组16 只,各组中实验动物和对照动物各半.实验动物单次腹腔注射剂量分别为10 mg/kg体重和20 mg/kg体重的NaF,对照动物单次腹腔注射等剂量的NaCl,注射量均为10 μl/g, 24 h后处死动物.采用HE染色、免疫组化染色观察高浓度氟对小鼠磨牙不同分化阶段成牙本质细胞形态及DSP的表达,采用SPSS 13.0软件对数据进行分析.结果:实验组分泌期成牙本质细胞形态紊乱,正常的高柱状形态丧失,DSP的表达明显强于对照动物,差异有统计学意义(P<0.01),而成熟期成牙本质细胞未见明显变化.结论:短期高浓度氟能增强分泌期成牙本质细胞中DSP的表达,抑制成牙本质细胞的增殖分化及随后的基质合成与分泌,从而影响牙本质的发育.     

根尖乳头干细胞向牙髓牙本质复合体分化能力的实验研究

目的:应用组织工程方法,探讨根尖乳头干细胞(SCAP)分别在炎性环境(肿瘤坏死因子-α)及正常环境下向牙髓牙本质复合分化的可能性.方法:分离大鼠根尖乳头组织,采用酶消化法结合组织块法获得SCAP,鉴定细胞来源,进行MTT生长曲线的测定,分正常组、炎症组(10 ng/mL TNF-α)及空白对照组行体外成骨能力茜素红及免疫组化实验,并将3个组分别移植到大鼠背部皮下观察8/12周后新生组织的形成情况.结果:MTT结果显示10 ng/mLTNF-α组显著高于正常SCAP组,差异有统计学意义(P＜0.5);茜素红染色显示均有矿化结节生成,与炎性组(10 ng/mL TNF-α)相比,正常组SCAP染色显著,矿化结节形成的范围大且呈深橘色;免疫组化染色显示SCAP胞浆中DSP/BSP均有表达;正常、炎症组移植物可见牙髓牙本质样结构形成,且DSP/BSP均阳性表达,空白对照组未见矿化组织形成.结论:SCAP具有高增殖及成骨向分化能力,可进行牙髓牙本质复合体的再生,炎性因子TNF-α对SCAP的生长增殖有促进作用同时不同程度上对SCAP矿化及成牙成骨向分化有抑制作用.     

Effect of overdose fluoride on the expression of DSP mRNA in rat incisors

目的 研究过量氟对大鼠切牙发育过程中牙本质涎蛋白( dentin sialoprotein,DSP) mRNA表达的影响.方法 选择6只Wistar大鼠,随机分成两组:对照组常规饲料喂养,自由饮用蒸馏水;实验组常规饲料喂养,自由饮用氟离子浓度为100 mg/L的氟化水,建立大鼠氟斑牙模型.饲养8周处死动物,提取大鼠切牙组织总RNA,逆转录为cDNA,利用实时荧光定量聚合酶链反应技术观察过量氟对大鼠切牙中DSP mRNA表达的影响.结果 实时荧光定量聚合酶链反应结果显示DSP mRNA在实验组表达水平明显高于对照组表达水平(t=2.132,P＜0.01).结论 过量氟可能通过增强DSP mRNA的表达,影响牙本质的发育矿化.     

bFGF在氟中毒大鼠切牙牙髓中的表达和意义

目的研究过量氟对大鼠牙髓细胞表达bFGF的影响。方法选择20只Wistar大鼠,随机分为2组I组(对照组)和II组(实验组)。8周后处死动物,利用磨片、HE染色、免疫组化染色技术观察过量氟对大鼠切牙形态及bFGF表达的影响。结果实验组大鼠切牙釉质、牙本质生长线明显,牙本质可见钙质小球,牙髓细胞及髓腔内侧牙本质bFGF表达明显弱于对照组,差异有显著性(P<0.01)。结论过量氟可抑制牙髓细胞表达bFGF,影响牙体硬组织的结构。     

正畸力对幼鼠牙本质中DSP表达的影响

目的:探讨正畸力对牙齿发育过程的牙本质中DSP表达的影响.方法:选取5周龄雄性SD大鼠,建立正畸牙齿移动模型.分别于加力后1、3、7、14、21 d处死动物,取含双侧上颌第一磨牙的上颌标本,制作石蜡切片,进行HE染色和DSP免疫组化染色.结果:牙冠部牙本质小管、前期牙本质、成牙本质细胞和牙根部成牙本质细胞DSP染色阳性.其中,牙尖部位的牙本质小管和前期牙本质呈强阳性染色,14 d时达到高峰,21 d时阳性染色有所减弱;根部成牙本质细胞在实验初期呈弱阳性染色,随时间的延长阳性染色逐渐增强.正常对照组DSP的表达明显弱于实验组.结论:对处于发育晚期的年轻恒牙施加适当的正畸力会促使成牙本质细胞进入活跃状态,牙本质中DSP表达上调,从而在一定程度上加速牙本质的形成及矿化.     

重建牙本质-牙髓复合体样结构的实验研究

目的 采用组织工程学的方法重建牙本质-牙髓复合体样结构。方法以大鼠牙髓干细胞为种子细胞，将羟基磷灰石-磷酸三钙（HA-TCP）复合支架材料接种至裸鼠背部皮下，8周后取材进行组织学及免疫组织化学检测。结果移植物组织学切片可见支架材料孔隙内有3层组织结构，由外向内分别为矿化牙本质、前期牙本质及牙髓组织，其中牙本质小管明显，牙髓组织外层细胞密集，部分出现极化，呈现出成牙本质细胞样细胞特征。免疫组织化学染色证实人牙本质特异性蛋白、DMP1在前期牙本质区及成牙本质细胞样细胞为阳性表达。结论采用牙髓干细胞为种子细胞、HA-TCP为支架材料可成功构建牙本质-牙髓复合体样结构，为进一步进行牙齿组织工程再造奠定基础。     

DSP、DMP1、CBFA1、BMP2在大鼠牙髓干细胞中的表达

目的检测DSP、DMP1、CBFA1、BMP2在大鼠牙髓干细胞中的表达,为大鼠牙髓干细胞的生物学功能研究提供基础。方法采用免疫组织化学染色的方法,对DSP、DMP1、CBFA1、BMP2在大鼠牙髓干细胞中的表达进行检测,然后采用矿化液对大鼠牙髓干细胞进行诱导,并对诱导后细胞进行DSP、DMP1表达的检测。结果大鼠牙髓干细胞DMP1仅个别细胞阳性表达,DSP少量细胞阳性表达,CBFA1、BMP2为阳性染色。经矿化液诱导后,大鼠牙髓干细胞DSP染色出现部分细胞强阳性,DMP1出现阳性结果。结论CBFA1、BMP2阳性表达表明大鼠牙髓干细胞具有一定的未成熟性,而经过诱导后出现牙本质特异性DSP的表达,表明大鼠牙髓干细胞可以被诱导向成牙本质细胞方向分化。     

永生化成牙本质细胞表达牙本质基质蛋白的实验研究

目的　探讨永生化人成牙本质细胞样细胞系hTERT-hOd-l表达牙本质基质蛋白的情况。方法　矿化液培养hTERT-hOd-l细胞5周,检测骨钙素(OC)分泌量和碱性磷酸酶(ALP)活性。采用免疫组织化学、RT-PCR和原位杂交方法检测Ⅰ型胶原、骨涎蛋白(BSP)、牙本质基质蛋白1 (DMP1)以及成牙本质细胞标志物牙本质涎磷蛋白(DSPP) 和牙本质涎蛋白(DSP)在细胞中的表达。结果　在矿化液诱导下,hTERT-hOd-l细胞ALP活性和OC分泌量升高。 hTERT-hOd-l细胞在mRNA水平上表达BSP、DMP1和DSPP,在蛋白质水平上表达DSP和Ⅰ型胶原。结论　hTERT- hOd-l细胞在体外表达牙本质基质蛋白,具有矿化的潜能。     

Notch 1信号蛋白在早期牙髓损伤修复中表达的免疫组化研究

目的 :通过建立大鼠牙髓损伤动物模型 ,观察Notch1信号蛋白在牙髓损伤修复过程中的表达。方法 :在大鼠第一磨牙牙合面开髓 ,分别对损伤后 2h、12h、2 4h、3d、7d及正常大鼠牙髓标本进行HE染色、免疫组化染色 ,检测各组标本中Notch1信号的表达。结果 :Notch1信号蛋白在正常成年大鼠牙髓组织中无表达 ,在牙髓损伤后表达上调。牙髓损伤 12h ,Notch1在成牙本质细胞层、部分牙髓细胞间质中呈弱阳性表达。损伤后 1～ 3d ,Notch1在成牙本质细胞层、部分牙髓细胞间质及接近损伤部位的牙髓细胞中呈阳性着色。损伤后 7d ,Notch1的表达位于成牙本质细胞层和穿髓孔下方的细胞增殖层。结论 :Notch1信号蛋白在大鼠牙髓损伤后早期阶段表达逐渐上调 ,随后在成牙本质细胞区及富细胞区牙髓细胞中表达增强 ,并沉积于穿髓点附近的细胞增殖层。提示它可能在牙髓损伤的早期自身修复和牙本质矿化中起作用     

The dentin sialoprotein (DSP) expression in rat tooth germs following fluoride treatment: an immunohistochemical study

Fluoride is known to alter expression of dentin matrix proteins and affect their posttranslational modifications. OBJECTIVE: The objective of our study was to examine dentin sialoprotein (DSP) expression in the early and late bell stages of development of the first molar tooth germs in rats treated with fluoride. DESIGN AND METHODS: Pregnant dumps were divided into three groups. They were fed a standard diet and from the fifth day of pregnancy, each group received either tap water (with trace amounts of fluoride), tap water with a low concentration of fluoride, or tap water with a high concentration of fluoride. Changes in DSP expression and distribution were visualized by immunohistochemistry. RESULTS: Immunoreactivity for DSP was detected in the cervical regions of the early bell stage in tooth germs of the 1-day-old animals. The earliest reaction was visible in the control group and the group supplemented with the low fluoride concentration (F(L)) but not in the group supplemented with the high fluoride concentration (F(H)). In early bell stages across all experimental groups, the immunoreactivity to DSP was observed in the cusp tip regions and was localized to preameloblasts, young and mature odontoblasts, dental pulp cells, predentin, and dentin. Generally, more intense positive staining for DSP was detected in animals supplemented with the high fluoride concentration. In the late bell stage found in the 4-day-old control group and the group supplemented with the low fluoride concentration, immunoreactivity for DSP was less intense compared with younger animals. However, immunoreactivity was greater in the group treated with the high dose of fluoride. In this group, the positive immunostaining for DSP, especially in young ameloblasts, was prolonged and relatively strong. CONCLUSIONS: Fluoride supplementation causes changes in the developmental pattern of DSP expression and its distribution in rat tooth germs.     

TGF-β1在氟中毒大鼠切牙牙髓中的表达

目的:研究过量氟对大鼠牙髓细胞TGF-β1表达的影响。方法:20只Wister大鼠,随机分为对照组和实验组。对照组用等量蒸馏水灌胃,实验组以20mg.kg-1.d-1氟化钠水灌胃。8周后处死动物,利用磨片、HE染色、免疫组化染色技术观察过量氟对大鼠切牙形态及TGF-β1表达的影响。采用SPSS10.0软件对数据进行t检验。结果:实验组大鼠切牙釉质牙本质生长线明显,球间牙本质增多,牙髓细胞及髓腔内侧牙本质TGF-β1表达显著弱于对照组,差异有显著性(P<0.01)。结论:过量氟可抑制牙髓细胞表达TGF-β1,影响牙体硬组织的结构。     

碱性成纤维因子用于犬牙盖髓剂的实验研究

目的探讨碱性成纤维因子(bFGF)对犬牙髓损伤修复的影响。方法选取健康英国小猎兔犬牙齿64颗,分为bFGF盖髓组、Dycal盖髓组及ZOE盖髓组。进行直接盖髓术,观察术后14天和28天修复性牙本质形成及牙髓组织学变化。免疫组化检测骨形成蛋白表达情况。结果术后14天:实验组及阳性对照组有纤维性基质形成,无牙本质桥形成;术后28天:实验组及阳性对照组有骨样牙本质桥、管状牙本质形成,阴性对照组无牙本质桥形成。结论bFGF在体内能够有效诱导牙髓细胞分化,形成修复性牙本质。     

五倍子提取液对牙本质龋损时胶原蛋白分解的影响

目的：评价五倍子提取液在牙本质龋损中对胶原分解的影响。方法；乳酸胶体系统致牙本质龋，20ml/L氟化钠，五倍子提取液，380ml/L氟化双氨银，去离子水4组试剂分别处理，再次脱矿及I型胶原酶牙本质基质，氯胺－T法对各组胶原分解量进行分析。结果：五倍子组和380ml／L氟化双氨银组对I型胶原酶分解胶原均有显著抑制作用（P＜0．01），且五倍子组明显优于380ml/L氟化双氨银组的抑制胶原分解作用（P＜0．01），结论：五倍子能抑制胶原分解，对脱矿的进一步进行产生影响，从而使牙本质龋的进展受到抑制。     

葡萄籽提取物促进早期牙本质龋损再矿化作用的体外研究

目的评价葡萄籽提取物（GSE）对人工牙本质龋的再矿化作用和显微结构的影响。方法制备60个人牙本质标本,采用化学法形成人工牙本质龋损,将样本随机分为4组,分别用10%GSE溶液、1 mg/L NaF溶液、10%GSE+ 1 mg/L NaF溶液和去离子水（DDW）处理,进行体外pH循环。采用显微硬度计测定处理前、后各样本的显微硬度值,扫描电镜观察pH循环后各组牙本质显微结构的变化,并用X线能谱仪分析牙本质元素百分含量。采用SPSS17.0软件包对数据进行统计学分析。结果显微硬度结果显示,GSE组牙本质的显微硬度获得百分比（SMHR）为40.87±9.92,NaF组为44.60±12.48,GSE+NaF组为48.54±9.27,去离子水组为15.98±8.33。与去离子水组相比,3个实验组的SMHR值均显著提高（P<0.05）,但相互之间无显著差异。扫描电镜显示,GSE组、NaF组和GSE+NaF组牙本质小管大部分呈封闭状态,而去离子水组牙本质小管呈开放状态。元素分析结果显示,3个实验组的Ca、P百分含量均显著高于去离子水组（P<0.05）。结论GSE能显著促进人工牙本质龋的再矿化作用,抑制牙本质龋的进展。     

不同氟离子导入条件对牙本质小管的封闭作用

目的通过体外实验研究导人氟离子的不同条件对牙本质小管封闭的效果。方法制备牙本质模型45个,每组5个共9组,其中正常牙本质组仅用打磨处理,其余8组模型用6％柠檬酸酸蚀后,除1个脱矿组之外,7个再矿化组用NF-Ⅱ型护齿仪在不同的电流、时间利NaF浓度下进行氟离子导入处理。扫描电镜观察各组牙本质模型表面及纵断面的形貌,将观测结果进行统计分析;另外,对钙、磷元素含量进行能量色散X射线分析。结果脱矿组大部分牙本质小管开放,再矿化组牙本质小管内可见明显的矿化物沉积,各组的牙本质小管面积和相对面积有显著性差异（P〈0．05）;而能量色散X射线分析显示脱矿组与再矿化各组表面钙磷比无显著性差异（P〉0．05）。结论NF-Ⅱ型护齿仪的氟离子导入对于牙本质小管有一定封闭作用,在本实验条件下,牙本质再矿化的最佳电流为0．1mA或0．3mA,最佳时间为6min,最佳的NaF浓度为1％。     

Effect of mineral trioxide aggregate on dentin bridge formation and expression of dentin sialoprotein and heme oxygenase-1 in human dental pulp

This study was conducted to evaluate the pulpal response to direct capping with either mineral trioxide aggregate (MTA) or calcium hydroxide (CH) cement in humans, with a focus on dentin bridge formation and dentin sialoprotein (DSP) and heme oxygenase-1 (HO-1) expression. Direct pulp capping was performed in 20 cases of caries-free human third molars. The pulps were exposed and capped with either MTA or hard-setting CH. After 2 months, the teeth were extracted, and the specimens were prepared for histologic and immunohistochemical evaluations. Histologically, 100% of the MTA group and 60% of the CH group developed dentin bridges. The mean thickness of the dentin bridges observed in the MTA group was statistically greater than that of CH group. In addition, DSP and HO-1 were expressed in the odontoblast-like cells and pulp fibroblasts beneath the dentin bridge; furthermore, significantly greater immunostaining was observed in the MTA group than in the CH group. Collectively, these results indicate that MTA is superior to CH in terms of inducing the dentinogenic process in human pulp capping.     

硼氟联合作用对大鼠切牙釉蛋白表达的影响

目的通过观察过量氟、硼以及氟硼联合作用对大鼠切牙釉蛋白表达的影响,初步探讨硼在预防氟斑牙中的作用。方法选择32只Wistar大鼠,随机分为4组。Ⅰ组常规饮用蒸馏水;Ⅱ组饮用含氟化钠质量浓度为220 mg/L的氟化水;Ⅲ组饮用含硼质量浓度为382 mg/L的水;Ⅳ组饮用含氟、硼质量浓度分别为220 mg/L、382 mg/L的水。8周后处死动物,获取切牙标本,利用免疫组织化学法和HE染色观察大鼠切牙成釉细胞和釉蛋白的表达。结果Ⅱ组釉蛋白的表达明显弱于Ⅰ组（P<0.01）,Ⅲ组与Ⅰ组表达差异无统计学意义,Ⅳ组与Ⅰ组表达差异无统计学意义,Ⅳ组与Ⅱ组表达差异有统计学意义（P<0.01）。结论过量氟可抑制釉蛋白的表达,而经硼与氟的联合作用后,釉蛋白表达所受影响降低。硼可降低氟对牙齿的危害性。     

Response of human pulps following acid conditioning and application of a bonding agent in deep cavities.

OBJECTIVES: The aim of this in vivo study was to evaluate the human dental pulp response when a one-bottle adhesive system was applied on etched or unetched deep dentine. METHODS: Eighteen class V deep cavity preparations were divided in three groups: group 1-total etching + two coats of single bond (SB) + composite resin (Z-100); group 2-enamel etching + two coats of SB + Z-100; group 3-cavity floor lined with a calcium hydroxide liner (Dycal) + acid-etching of enamel and lateral walls + two coats of SB + Z-100. Two teeth were used as intact control group. After 30 days the teeth were extracted and processed through H and E, Masson's trichrome and Brown and Brenn staining techniques. RESULTS: Moderate inflammatory response, disorganization of pulp tissue, as well as, deposition of thin layer of reactionary dentin were observed in group 1 teeth in which the remaining dentin thickness (RDT) was less than 300 microm. These histological findings appear to be related to long resin tags formation and bonding agent diffusion through dentinal tubules. In group 2, slight inflammatory response was observed only in one tooth in which the RDT was 162 microm. In group 3, all the teeth showed normal histological characteristics which were similar to the intact control group. Presence of bacteria was not correlated with the intensity of pulpal response. The patients reported no symptoms during the experiment. Radiographic evaluation showed no periapical pathology for any of the teeth. SIGNIFICANCE AND CONCLUSIONS: Acid-etched deep dentin (RDT less than 300 microm) lined with SB causes more intense pulpal response than unetched deep dentin. Based on the results observed in the present study and the conditions in which it was carried out, we recommend the application of a biocompatible liner before etching deep dentin and applying SB.