Notch1蛋白在小鼠下颌切牙发育中的表达

目的探讨Notch1在小鼠下颌切牙牙胚发育过程中的组织学分布。方法制作ICR小鼠下颌切牙不同发育阶段的冰冻组织切片,对小鼠下颌切牙牙胚自牙胚发育起始期至钟状晚期不同发育阶段组织Notch1的分布情况进行免疫组织化学染色。结果 Notch1在小鼠下颌切牙发育蕾状期牙胚的口腔侧上皮中表达,而在和间充质相邻的牙胚上皮中没有表达。从帽状期至钟状期,Notch1在牙胚的中间层表达,而在内釉上皮中没有表达。钟状期的唇侧颈环部位星网状层和部分牙上皮细胞也表达Notch1。此外,Notch1还在牙胚发育不同时期的间充质、牙乳头和早期牙髓中表达。结论 Notch1可能在小鼠下颌切牙发育过程中的牙上皮,特别是内釉上皮的细胞分化,以及牙囊和牙乳头细胞分化及分化完成后的牙髓干细胞的稳定性方面有重要作用。     

透明质酸在小鼠下颌第一磨牙牙胚不同发育时期的表达

目的:检测透明质酸在小鼠下颌第一磨牙牙胚不同发育时期的表达,探讨其在小鼠牙胚发育过程中的作用。方法:取不同胎龄的ICR胎鼠,制备小鼠下颌第一磨牙不同发育时期切片,用免疫组织化学实验方法检测透明质酸在下颌第一磨牙牙胚组织中的表达情况。结果:透明质酸在小鼠下颌第一磨牙牙胚的不同发育阶段表达各异。在小鼠磨牙开始发生时,透明质酸即在增厚的牙板上皮中表达,随后在蕾状期牙蕾中央的上皮细胞间可见透明质酸的表达,而在深层的牙源性间充质表达则不明显。自帽状期至钟状晚期,透明质酸在牙胚星网状层细胞间以及牙源性间充质细胞的表达逐渐增强,而在基底膜、内外釉上皮细胞、成釉细胞以及成牙本质细胞所在区域不表达。结论:透明质酸在牙胚发育过程中呈现时间-空间特异性表达。特别是其在星网状层细胞以及牙乳头间充质细胞中的表达随着牙胚的发育逐渐增强提示其可能与牙胚的形态发生密切相关。     

氯离子通道CLC-3在小鼠牙胚发育中的表达

目的:探讨CLC-3在BALB/c小鼠下颌第一磨牙牙胚不同发育时期的时空表达。方法:选取E13.5、E15.5、E18.5 d的BALB/c胎鼠和P1、P5 d的BALB/c小鼠,脱颈处死后取其头部,梯度乙醇脱水,石蜡包埋后连续切片,最后对石蜡切片行CLC-3免疫组化染色。结果:当胎鼠牙胚处于蕾状期时,CLC-3在牙胚的牙板上皮中可见表达;当牙胚发育为帽状期时,在牙乳头和牙囊细胞中表达;到钟状期时,CLC-3在牙乳头、牙囊、星网状层细胞和中间层细胞中广泛表达。出生后的小鼠CLC-3在牙乳头、牙囊、成釉细胞、成牙本质细胞、星网状层细胞、中间层细胞中均广泛表达。结论:CLC-3在小鼠牙胚发育的不同时期呈现不同的时空表达。     

Pik3cb在小鼠下颌第一磨牙牙胚发育不同阶段的表达研究

目的:检测Pik3cb在小鼠下颌第一磨牙牙胚发育不同阶段的表达分布及变化规律,进一步揭示Pik3cb对牙形态发生的影响。方法:制备原位杂交探针,及小鼠牙胚发育标志时间点的牙胚冰冻切片,通过原位杂交方法进一步显示Pik3cb在小鼠下颌第一磨牙牙胚发育不同时期的表达情况。结果:蕾状期Pik3cb明显表达于牙板上皮,帽状期开始大量表达于釉上皮与牙乳头,进入钟状期表达部位趋于集中,尤以内釉上皮及后续新形成硬组织周围的高柱状成釉细胞处最为明显。结论:Pik3cb在牙源性上皮及成釉细胞表达,并可能通过PI3K/PTEN/AKT/mTOR信号通路与成釉细胞瘤行为相关。     

Fibulin-7在小鼠下颌第一磨牙发育过程中的时空表达

目的：观察Fibulin-7蛋白在小鼠下颌第一磨牙牙胚发育过程中的时空表达特点。方法：取胚胎15.5 d、出生后第1天、第7天的C57BL/6J小鼠下颌骨标本,应用免疫组织化学方法检测Fibulin-7在下颌第一磨牙牙胚不同发育时期的表达情况。结果：Fibulin-7蛋白在小鼠下颌第一磨牙牙胚自钟状期末期至牙胚发育成熟过程中表达量逐渐增加。结论：Fibulin-7蛋白在牙胚发育的不同阶段均有表达,推测Fibulin-7参与了牙间充质细胞分化为成牙本质细胞、牙髓-牙本质复合体的形成过程。     

Barx1与Wnt-1在大鼠磨牙胚发育的时空表达及意义

目的 探讨Barx1、Wnt-1在SD大鼠磨牙胚发育过程中的时空表达及其在牙齿发育过程中的作用.方法 制备SD大鼠磨牙发育各期标本,免疫组织化学检测Barx1、Wnt-1、β-catenin的表达情况.结果 Barx1在帽状期开始表达于牙乳头、牙囊,在内釉上皮、星网状层中有少量表达;在钟状形态发生期广泛表达于内釉上皮、星网状层、中间层、牙乳头,牙囊及外釉层也有少量表达;钟状分化期在前成釉细胞、中间层、前牙本质细胞、牙乳头有表达;钟状分泌期主要在成釉细胞、星网层中表达,牙乳头、中间层、成牙本质细胞表达减弱.Wnt-1和β-catenin在蕾状期强表达于牙板、牙蕾上皮;在帽状期主要表达于牙板上皮、内釉上皮、星网状层,少量表达于外釉上皮、牙乳头、牙囊;钟状期后在各部分表达都逐渐减弱,Wnt-1与β-catenin的表达量正相关(r＝0.4332,P<0.001).结论 Barx1、Wnt-1、β-catenin在SD大鼠磨牙胚发育的不同阶段和不同的牙胚组织细胞间均存在差异,提示其在磨牙胚的增殖、分化过程中起着各自不同的作用.     

Shh在鼠磨牙牙胚发育晚期的基因表达

目的：观察信号分子Sonic hedgehog(Shh)在小鼠下颌第一磨牙牙胚发育晚期的基因表达，探讨其在成釉细胞、成牙本质细胞分化中的作用。方法：制备昆明小鼠磨牙牙胚发育晚期(E16．5—P1．5)标本，用原位杂交法分析Shh mRNA在牙胚中的表达和分布。结果：Shh mRNA在牙胚发育晚期的前成釉细胞和中间层细胞呈阳性表达，在成牙本质细胞层表达较弱。结论：在牙胚发育晚期，Shh可能通过自分泌途径和旁分泌途径，参与了成釉细胞和成牙本质细胞分化。     

同源盒基因Msx-1、Msx-2和Dlx-2在小鼠下颌第一磨牙发育阶段的表达

目的 观察同源盒基因Msx—1、Msx-2和Dlx-2 mRNA在小鼠下颌第一磨牙发育阶段的表达。方法 取胎龄E11～E18和新生P1～P3小鼠的头或下颌，制备5μm连续冠状切片。体外转录合成地高辛标记的Msx—1、Msx-2和Dlx-2 RNA探针。原位杂交后观察Msx—1、Msx—2和Dlx—2在小鼠下颌第一磨牙中的表达。结果 在牙胚发育过程中，Msx—1转录信号始终只在间质细胞中观察到，在上皮细胞中呈阴性。Msx-2和Dlx-2在牙源性上皮和间质中均表达，在磨牙发育起始期二者表达相似，但在随后的牙胚发育过程中，它们出现不同的表达特征。结论 牙胚发育过程中，Msx—1、Msx-2和Dlx-2有不同的表达特征，可能共同参与调控上皮和间质问的相互作用。     

Notch1及其配体Jagged1在小鼠颌下腺胚胎发育中的表达

目的探讨Notch1及其配体Jagged1在小鼠颌下腺胚胎发育过程中的组织学分布。方法制作ICR小鼠颌下腺不同发育阶段的冰冻组织切片,对小鼠颌下腺自始基至出生后2d不同发育阶段组织的Notch1和Jagged1的分布情况进行免疫组织化学染色。结果 Notch1在小鼠颌下腺发育的蕾状期和假腺管期上皮很少表达。从微管期早期开始,在顶端胚芽上皮中强表达,而在导管上皮中未见表达。在微管期后期,顶端胚芽上皮部分细胞Notch1仍强表达,部分细胞表达减弱。自终末分化期开始,导管上皮开始表达Notch1,而分化完成的腺泡中只有少量细胞仍表达Notch1。在Notch1表达细胞群的周围细胞有Jagged1表达。结论 Notch1可能在小鼠颌下腺发育过程中的细胞分化以及分化完成后维持干细胞的稳定性方面有重要作用。     

Ddit3在小鼠下颌第一磨牙牙胚不同时期的表达研究

目的:检测Ddit3在小鼠下颌第一磨牙牙胚不同时期的表达分布及变化规律,初步揭示Ddit3在小鼠牙胚发育中的作用。方法:取不同胚胎时间点的ICR妊娠小鼠,制备牙胚发育标本,通过免疫荧光和免疫组化的方法显示Ddit3在小鼠下颌第一磨牙牙胚发育不同时间点的表达分布。结果:Ddit3在牙胚发育的早期主要表达于胞浆中,从钟状期开始,Ddit3不仅在胞浆中有阳性表达,同时也微弱地表达于细胞核中。分布如下:在小鼠下颌第一磨牙发育的板状期,Ddit3几乎没有表达。在蕾状期,Ddit3主要表达于釉上皮的细胞质中,在牙外胚间充质细胞中几乎无表达。在帽状期,Ddit3的表达模式基本和蕾状期相同。在钟状期早期,Ddit3在成釉细胞、前成牙本质细胞和牙乳头胞质中呈阳性表达,在部分细胞核中也检测到了Ddit3的表达。钟状期晚期,Ddit3在新形成的硬组织周围的高柱状成釉细胞、成牙本质细胞和牙乳头细胞的胞浆中有阳性表达,在大多数成釉细胞、成牙本质细胞和牙乳头细胞的细胞核中也有表达。结论:Ddit3可能会调节成釉细胞和成牙本质细胞的分化及其硬组织的生物矿化。     

Expression of neural cell-adhesion molecule mRNA during mouse molar tooth development

This study employed in situ hybridisation using a probe recognising all isoforms of the molecule. Expression of the molecule in tooth germs started at embryonic day 13, when they were at the bud stage. Both inner cells of the epithelial bud and peripheral cells of the dental mesenchyme were positive. At the cap stage, positive cells were found in the inner part of the enamel organ but only in a limited area near the outer enamel epithelium. In the mesenchyme at the cap stage, expression was weak in the dental papilla and strong in the follicle. From the bell stage onward, epithelial cells in the enamel organ were negative except for the cells of the stratum intermedium, which were transiently positive at early and late bell stages. In the dental papilla, expression had mostly ceased during and after the bell stage, although transient expression was found in cuspal areas at the early bell stage. The dental follicle strongly expressed neural cell-adhesion molecule (NCAM) to the end of the experimental period, at post-natal day 4. In contrast to the first molar at its earliest stage of appearance, in which both the thickened epithelium and surrounding mesenchyme were negative for the expression of the molecule, the second molar appeared as a combination of extending epithelial thickenings and mesenchymal cells strongly positive for its expression. This study newly identifies the dental papilla and the stratum intermedium as NCAM-expressing sites.     

Expression of aquaporin isoforms during human and mouse tooth development

Previously, we described the development of hyaluronan (HA) deposition in human tooth germ tissues that are consistent with water transport in different stages of tooth development. The aquaporins (AQP) constitute a family of membrane water channels that are expressed in many organs. However, there are no data available about the expression pattern of aquaporin water channels in dental structures. In the present study we have characterised the expression of six different aquaporin isoforms (AQP1-5, AQP-9) in developing human and mouse tooth germs by immunohistochemistry using isoform specific antibodies. In the "bell stage" AQP1 was expressed in endothelial cells of small vessels whereas no other structures of the tooth primordial were labeled. AQP2, AQP3 and AQP9 immunoreactivity was not observed in tooth germs, whereas strong AQP4 and AQP5 expression was observed in dental lamina, inner enamel epithelium, stratum intermedium, stellate reticulum and the outer enamel epithelium. Oral epithelium also exhibited AQP4 and AQP5 immunolabeling. During development of the matrices of the dental hard tissues AQP4 and AQP5 immunostaining was observed in the odontoblasts and their processes, as well as in the secretory ameloblast and their apical processes. Immunolabeling controls were negative. In conclusion, AQP4 and AQP5 are expressed in tooth germ tissues in early development in cells that previously have been shown to express HA and/or CD44, indicating that AQP water channels may play a role for ECM hydration during tooth development.     

The distribution pattern of the hyaluronan receptor CD44 during human tooth development.

The aim was to investigate the expression pattern of the major cell-surface hyaluronan receptor CD44, as there are no existing data on its presence or absence in human dental structures at different developmental stages. Immunohistochemical localization of CD44 was studied using a monoclonal antibody, H3, that specifically recognizes an epitope in the common backbone of all CD44 isoforms. The dental lamina displayed a strong CD44 signal; the external enamel epithelium was negative. In the coronal region of the tooth germ the presecretory ameloblasts showed an intense reaction whereas the less differentiated inner enamel epithelial cells showed no signal at the cervical loop where they meet the external enamel epithelium. In the stellate reticulum a moderate reaction was detected. The secretory ameloblasts and the stratum intermedium showed a strong cell-surface CD44 signal. A strong signal was also observed on the odontoblasts and their processes. In the pulp, close to the odontoblastic layer, weak labelling was seen in the walls of capillary vessels. The distribution of CD44 in the human tooth germ corresponds to that of hyaluronan in most locations, suggesting that during tooth development this transmembrane protein plays an important part in hyaluronan-mediated events.     

跨膜蛋白Syndecan-1在不同发育时期小鼠磨牙组织中的表达

目的以小鼠磨牙为发育模型,研究其不同发育时期跨膜蛋白Syndecan- 1的表达特点,进而分析Syndecan-1在牙齿发育中的作用。方法取不同胎龄的胎鼠,制作其下颌第一磨牙的切片,进行免疫荧光染色,并在荧光显微镜下观察Syndecan- 1的表达情况。结果Syndecan- 1的表达随牙胚发育的时期不同而变化：蕾状期时在牙源性上皮和间充质中有弱的阳性分布,帽状期时成釉器上皮阳性表达减弱,而牙乳头及周围的间充质的阳性反应明显增强,钟状期时成釉器上皮又呈阳性表达,并以在中间细胞层的反应更为强烈,而牙乳头间充质的染色则很弱,同时前成釉细胞以及下方的成牙本质细胞的顶端也呈Syndecan- 1的阳性表达。结论Syndecan- 1参与了成釉器和牙乳头的发育和分化过程的调节,并与牙胚细胞的增殖以及成釉细胞和成牙本质细胞的分化相关。     

The separation and culture of cell populations from bovine molar tooth germs

Separated components of the maxillary molar tooth germs were cultured. The typical epithelial characteristics of enamel organ cells were retained. The cultured cells of the reduced enamel epithelium, outer enamel epithelium and stratum intermedium became polygonal with numerous microvilli, in contrast with the lack of microvilli and elongated profile of stellate reticulum and dental papilla cells. A plexiform organization of cytoskeletal microfila-ments was noted in cells which became polygonal, whereas in elongated cells, microfilaments were arranged in parallel bundles beneath the cell surface. In the cultured epithelial cells, but not dental papilla cells, cytoskeletal tonofilaments as well as cell surface specializations at contact sites between cells were common.     

Shh、Ptch1及Ptch2基因在小鼠磨牙发育过程中的表达

目的通过原位杂交的方法研究Sonic Hedgehog（Shh）、Patched1（Ptch1）及Patched2（Ptch2）在小鼠下颌磨牙发育过程中的表达，以探讨该信号通路在牙胚发育中的作用。方法制备小鼠下颌第一磨牙发育各期标本，原位杂交检测Shh、Ptch1及Ptch2的表达部位及强度。结果Shh在帽状期表达于釉结，钟状早期表达于内釉上皮，钟状晚期表达于成釉细胞；Ptch1在帽状期表达于牙囊、外釉上皮及星网状层，钟状早期表达于牙囊、外釉上皮及牙乳头，钟状晚期表达于成牙本质细胞及牙乳头；Ptch2在帽状期表达于釉结，钟状早期表达于内釉上皮，钟状晚期无表达。结论Shh信号系统在牙胚发育各期有各自的表达特点，提示可能在牙胚形态的调控，成釉细胞、成牙本质细胞分化的诱导中起重要作用。     

组蛋白去甲基化酶JMJD3在小鼠牙发育中的表达

目的 研究组蛋白去甲基化酶JMJD3在牙发育过程中的表达.方法 采用免疫组织化学方法观察小鼠胚胎E14~18天及出生后P1~P3磨牙牙胚中组蛋白去甲基化酶JMJD3的表达.结果 JMJD3表达于细胞核内,包括上皮性细胞和间充质细胞.在小鼠磨牙牙胚发育的蕾状期(E14)和帽状期(E16),JMJD3在牙蕾上皮,成釉器内釉细胞、外釉细胞、星网状细胞中弱表达.在小鼠磨牙牙胚发育的钟状期(E18),JMJD3在成釉器的外釉细胞层,内釉细胞层,中间层、星网状层及邻近间充质细胞内呈强阳性表达.在小鼠磨牙牙胚发育钟状晚期(P3), JMJD3在成釉细胞、成牙本质细胞及邻近间充质细胞呈强阳性表达.结论 JMJD3在小鼠牙发育过程中呈时空特异性表达,提示JMJD3参与小鼠磨牙的发育.     

肝细胞生长因子受体在大鼠牙胚发育中的表达及其意义

目的：研究肝细胞生长因子受体(c—Met)在大鼠牙胚发育过程中的时空表达模式。方法：用免疫组织化学方法，检测c—Met在大鼠牙胚发育不同阶段表达的位置。结果：c—Met在牙胚发育的不同阶段表达于牙胚中特定的部位：帽状期(胚胎15d)，内釉上皮阳性表达，中间层及部分星网状层细胞弱阳性表达；钟状期(胚胎19d)，内、外釉上皮阳性表达，但内釉上皮较外釉上皮表达弱，中间层及星网状层细胞弱阳性表达；矿化期(生后7d)，成牙本质细胞阳性表达。结论：c—Met在大鼠牙胚发育过程中的表达具有时空特异性，在牙齿形态发生的过程中具有重要的作用，还可能与牙齿的矿化有关。