牙本质基质蛋白1在人牙胚发育过程中的表达和意义

目的：观察牙本质基质蛋白1(dental matrix protein，DMP1)在人牙胚发育过程中的表达和分布变化，探讨DMP1在人牙齿发育过程中的作用。方法：制备人牙胚发育各期组织标本，采用免疫组织化学方法观察DMP1在人牙胚不同阶段的表达情况。结果：DMP1主要表达于钟状晚期分泌型成牙本质细胞，在前成牙本质细胞、前成釉细胞和牙乳头细胞中表达弱阳性，在成釉细胞中有一过性表达，即在釉基质开始形成但没有形成硬组织时的分泌型成釉细胞中有表达，但在随后的分泌型成釉细胞中表达渐弱至阴性。结论：观察到DMP1在人牙胚发育不同时期的表达和表达变化，提示其可能参与成牙本质细胞和成釉细胞的分化及牙本质形成过程。     

牙本质基质蛋白1在组织中的表达及其对牙本质形成调控作用的研究进展

牙本质基质蛋白1(DMPI)是非胶原蛋白中的重要一员,属于SIBLINGs家族,在硬组织形成和发育中具有重要作用.该文就近几年对DMP 1基因、蛋白结构、分布以及在牙本质形成调控方面的研究作一综述.     

人牙本质涎蛋白在人牙胚发育过程中的表达和意义

目的：探讨人牙本质涎蛋白(human dentin sialoprotein，hDSP)在人牙胚发育过程中的表达和意义。方法：用免疫组化方法检测人牙本质涎蛋白在人牙胚不同发育阶段中的表达。结果：hDSP在蕾状期、帽状期釉上皮以及钟状早期内釉上皮有弱阳性表达，钟状中期正在分泌基质的成牙本质细胞、牙本质小管有强阳性表达，钟状晚期，牙本质小管仍有强阳性表达，而成牙本质细胞转为弱阳性表达。前成釉细胞、成釉细胞有一过性表达。前期牙本质始终无阳性表达。牙胚周围骨组织、软骨组织和口腔软组织无阳性表达。结论：提示hDSP可能参与了牙本质的形成。另外前期牙本质阴性表达，表明hDSP蛋白由成牙本质细胞分泌后，可能通过成牙本质细胞突起穿过前期牙本质分泌至矿化前沿，参与牙本质的形成。     

Shh在大鼠牙胚发育过程中的表达

目的 :通过检测Shh(SonicHedgehog)在大鼠牙胚发育不同时期的表达 ,探讨其在牙齿发育过程中的作用。方法 :采用免疫组化方法 ,观察大鼠牙齿发育各阶段标本中Shh的表达。结果 :大鼠牙齿发育早期 ,Shh表达于口腔上皮和牙上皮 ;帽状期表达于成釉器 ;钟状期 ,成釉细胞和成牙本质细胞表达阳性。结论 :Shh在牙齿发育过程中的表达有时空特异性 ,提示它参与牙釉质和牙本质的形成     

甲状旁腺激素对人牙乳头细胞牙本质基质蛋白1表达的影响

目的：观察甲状旁腺激素(parathyroid hormone，PTH)对体外培养的人牙乳头细胞(human dental papilla cells，HDPC)牙本质基质蛋白1(dental matrix protein，DMP1)合成分泌的影响。方法：原代方法培养出人牙乳头细胞，用不同浓度的甲状旁腺激素刺激培养的第5代人牙乳头细胞，免疫组化观察结果，并采用图像分析的方法，进行半定量分析。结果：PTH可刺激人牙乳头细胞DMP1的表达，诱导的DMP1主要表达于细胞胞浆内，呈一定的浓度依赖性，0．3μg／mL为刺激最佳浓度。结论：PTH能够刺激人牙乳头细胞产生DMP1，对牙乳头细胞向成牙本质细胞分化有一定促进作用，可能是成牙本质细胞分化的重要介质之一。     

人牙本质基质蛋白1 cDNA序列的克隆

目的：克隆人牙本质基质蛋白１（ＤＭＰ１）成熟肽编码区基因片段。方法：用异硫氰酸胍一步法从引产胎儿牙乳头组织中抽提总ＲＮＡ,用Ｏｌｉｇｏ（ｄｔ）作引物逆转录合成牙乳头ｃＤＮＡ,然后利用ＰＣＲ方法,从ｃＤＮＡ中扩增出人ＤＭＰ１成熟区的基因片段（约１．８ｋｂｐ）,将所得基因片段插入ｐＢｌｕｅｓｃｒｉｐｔ质粒载体,转化到大肠杆菌ＸＬ１－Ｂｌｕｅ后挑选阳性克隆,提取重组质粒ＤＮＡ,通过酶切分析和核苷酸序列分析鉴定阳性克隆。结果：酶切图谱和部分序列分析结果与国外文献报道一致。结论：克隆到人ＤＭＰ１成熟肽编码区基因片段。     

牙本质基质蛋白-1与骨的形成及发育

摘要牙本质基质蛋白-1作为牙特异性细胞外基质蛋白,其在骨形成及发育的作用与功能得到了广泛的关注与研究。研究表明,牙本质基质蛋白-1在骨组织中高度表达,牙本质基质蛋白-1在骨形成及发育中起重要作用,本文拟就牙本质基质蛋白-1在骨发育中的作用与机理研究进展作一综述。     

大鼠牙本质基质蛋白1对人牙髓细胞增殖和ALP活性的影响

目的 :探讨大鼠牙本质基质蛋白 1(DMP1)对体外培养的人牙髓细胞增殖和ALP活性的影响。方法 :利用MTT法研究牙髓细胞的细胞增殖情况 ,碱性磷酸酶检测试剂盒检测牙髓细胞内ALP活性。结果 :DMP1在细胞培养 3d和 5d时 ,5 μg/mL组可促进人牙髓细胞增殖 (P <0 .0 5 )。细胞培养 3d时 ,仅5 μg/mL组可促进人牙髓细胞ALP活性 (P <0 .0 5 ) ;培养 5d时 1μg/mL和 5 μg/mL均可促进人牙髓细胞ALP活性 (P <0 .0 5 ) ;培养 7d时促进作用进一步加强 (P <0 .0 5 )。结论 :DMP1在一定浓度下可促进牙髓细胞的增殖 ;DMP1对牙髓细胞ALP活性的促进作用呈浓度依赖性 ,高浓度DMP1可以促进人牙髓细胞ALP活性 ,随着时间的延长 ,促进ALP活性的作用也增强。     

牙本质基质蛋白1在人不同龋坏牙齿中的免疫定位

目的 :观察牙本质基质蛋白 1(dentalmatrixprotein ,DMP1)在不同龋坏人牙齿中的表达 ,探讨DMP1在牙髓损伤修复中的作用。方法 :采用免疫组化SABC方法观察DMP1在不同龋坏人恒牙中的表达 ,并用图像分析系统进行半定量分析。结果 :DMP1在正常人恒牙组表达阴性 ,浅龋组部分成牙本质细胞和前期牙本质中弱阳性表达 ;中龋组成牙本质细胞数目增多 ,在胞浆和胞突以中阳性表达 ;深龋组成牙本质细胞层排列紊乱 ,出现空泡变性 ,成牙本质细胞样细胞形态细长、扁平 ,两种细胞胞浆和胞突中DMP1表达阳性 ,在修复性牙本质层强阳性表达。与对照组相比 ,成牙本质胞浆中DMP1含量在中龋组和深龋组明显增加 ,浅龋组无显著意义。结论 :DMP1参与成牙本质细胞样细胞分化分泌 ,并在修复性牙本质和反应性牙本质形成过程中起重要作用     

牙本质基质蛋白-1的研究进展

牙本质基质蛋白-1是牙发育过程中一种非常重要的非胶原性基质蛋白,对牙本质的形成和矿化起着重要的作用,但其具体的作用机制有待于进一步的研究.下面就牙本质基质蛋白-1的表达定位、基因结构、生物学功能以及表达调控因素作一综述.     

Inositol hexasulphate, a casein kinase inhibitor, alters the distribution of dentin matrix protein 1 in cultured embryonic mouse tooth germs

Immunohistochemical studies using a polyclonal antibody, raised against the recombinant form of dentin matrix protein 1 (DMP1), show that DMP1 was detected mainly in odontoblasts in cultured mouse embryonic tooth germs. However, in restricted areas, DMP1 staining was also observed in secretory ameloblasts, in the stratum intermedium and stellate reticulum, but only when the odontoblasts located in front of them were unstained. When the embryonic tooth germs were cultured in the presence of inositol hexasulfate, a casein kinase I and II inhibitor, staining of odontoblasts was weak or nil, whereas, in contrast, ameloblasts and enamel organ were strongly immunolabelled, suggesting an enhanced translocation of DMP1 after secretion to the secretory ameloblasts and/or stratum intermedium and stellate reticulum. Moreover, DMP1--was shown to be a good substrate for gelatinase A (MMP-2), but not to gelatinase B (MMP- 9). We hypothesized that DMP1--or the sub-fractions cleaved by the MMP--could behave as diffusible signaling molecule (s) rather than as a true dentin extracellular matrix component.     

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。     

Glycosylation of DMP1 maintains cranial sutures in mice

Craniosynostosis, a severe craniofacial developmental disease, can only be treated with surgery currently. Recent studies have shown that proteoglycans are involved in the suture development. For the bone matrix protein, dentin matrix protein 1 (DMP1), glycosylation on the N-terminal of it could generate a functional proteoglycan form of DMP1 during osteogenesis. We identified that the proteoglycan form of DMP1 (DMP1-PG) is highly expressed in mineralisation front of suture. But, the potential role of DMP1-PG in suture fusion remain unclear. To investigate the role of DMP1-PG in cranial suture fusion and craniofacial bone development. By using a DMP1 glycosylation site mutation mouse model, DMP1-S89G mice, we compared the suture development in it with control mice. We compared the suture phenotypes, bone formation rate, expression levels of bone formation markers in vivo between DMP1-S89G mice and wild-type mice. Meanwhile, cell culture and organ culture were performed to detect the differences in cell differentiation and suture fusion in vitro. Finally, chondroitin sulphate (CHS), as functional component of DMP1-PG, was employed to test whether it could delay the premature suture fusion and the abnormal differentiation of bone mesenchymal stem cells (BMSCs) of DMP1-PG mice. DMP1-S89G mice had premature closure of suture and shorter skull size. Lack of DMP1-PG accelerated bone formation in cranial suture. DMP1-PG maintained the essential stemness of BMSCs in suture through blocking the premature differentiation of BMSCs to osteoblasts. Finally, chondroitin sulphate, a major component of DMP1-PG, successfully delayed the premature suture fusion by organ culture of skull in vitro. DMP1-PG could inhibit premature fusion of cranial suture and maintain the suture through regulating the osteogenic differentiation of BMSCs.     

Porcine dentin matrix protein 1: gene structure, cDNA sequence, and expression in teeth

Dentin matrix protein 1 (DMP1) is an acidic non-collagenous protein that is necessary for the proper biomineralization of bone, cartilage, cementum, dentin, and enamel. Dentin matrix protein 1 is highly phosphorylated and potentially glycosylated, but there is no experimental data identifying which specific amino acids are modified. For the purpose of facilitating the characterization of DMP1 from pig, which has the advantage of large developing teeth for obtaining protein in quantity and extensive structural information concerning other tooth matrix proteins, we characterized the porcine DMP1 cDNA and gene structure, raised anti-peptide immunoglobulins that are specific for porcine DMP1, and detected DMP1 protein in porcine tooth extracts and histological sections. Porcine DMP1 has 510 amino acids, including a 16-amino acid signal peptide. The deduced molecular weight of the secreted, unmodified protein is 53.5 kDa. The protein has 93 serines and 12 threonines in the appropriate context for phosphorylation, and four asparagines in a context suitable for glycosylation. Dentin matrix protein 1 protein bands with apparent molecular weights between 30 and 45 kDa were observed in partially purified dentin extracts. In developing teeth, immunohistochemistry localized DMP1 in odontoblasts and the dentinal tubules of mineralized dentin and in ameloblasts, but not in the enamel matrix.     

尼古丁对大鼠牙槽骨内ALP和DMP1 C末端表达的影响

目的:观察尼古丁对大鼠牙槽骨内碱性磷酸酶(ALP)和牙本质基质蛋白1(DMP1)C末端表达的影响. 方法:将20只5周龄健康雄性Wistar大鼠随机等分为实验组和对照组,每组再随机等分为30天和60天两个亚组,实验组每天腹腔注射尼古丁0.73 mg·kg-1. 分别使用钙钴法和免疫组化法检测ALP和DMP1 C末端在大鼠牙槽骨内的表达. 结果: 对照组:ALP和DMP1 C末端都为深棕色线形表达于牙槽骨的钙化前缘, 形成明显的矿化条带;ALP和DMP1 C末端分别为棕色沉淀和棕黄色颗粒状密布于骨细胞周围.实验组:ALP和DMP1 C末端表达部位同对照组,表达强度减弱;相对于30天组,60天组变化更明显. 结论:尼古丁可能通过抑制大鼠牙槽骨内ALP和DMP1 C末端的表达,抑制大鼠牙槽骨的矿化.     

Immunohistochemical study of small integrin-binding ligand, N-linked glycoproteins in reactionary dentin of rat molars at different ages

Small integrin-binding ligand, N-linked glycoproteins (SIBLING) are believed to play key roles in the process of biomineralization. Reactionary dentin (RD), formed by odontoblasts in response to external stimuli, differs morphologically from primary dentin (PD). To test our hypothesis that the microscopic changes reflect variations in molecular mechanisms involved in formation of the two forms of dentin, and to characterize RD further, we compared the distributions of four SIBLING proteins [bone sialoprotein (BSP), osteopontin (OPN), dentin matrix protein 1 (DMP-1) and dentin sialophosphoprotein (DSPP)] in naturally occurring RD with those in PD. Molars of rats aged 12, 18, 24 and 36 wk were analyzed using immunohistochemistry with antibodies against BSP, OPN, DMP-1, and dentin sialoprotein (a fragment of DSPP). Differences in the distribution of the four SIBLING proteins were evident. Bone sialoprotein, not seen in PD, was consistently observed in RD. Osteopontin, almost absent from PD, was clearly observed in RD. The expression levels of DMP-1 and DSP in RD were lower than in PD. Elevated expression of BSP and OPN, along with a marked decrease of dentin sialoprotein and DMP-1 in RD, suggests a difference in the mechanism of formation of the two forms of dentin.     

Distribution of SIBLING proteins in the organic and inorganic phases of rat dentin and bone

The SIBLING protein family is a group of non-collagenous proteins (NCPs) that includes dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP1), bone sialoprotein (BSP), and osteopontin (OPN). In the present study, we compared these four proteins in different phases of rat dentin and bone. First, we extracted NCPs in the unmineralized matrices and cellular compartments using guanidium-HCl (G1). Second, we extracted NCPs closely associated with hydroxyapatite using an EDTA solution (E). Last, we extracted the remaining NCPs again with guanidium-HCl (G2). Each fraction of Q-Sepharose ion-exchange chromatography was analyzed using sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE), Stains-All stain, and with western immunoblotting. In dentin, the NH₂-terminal fragment of DSPP and its proteoglycan form were primarily present in the G1 extract, whereas the COOH-terminal fragment of DSPP was present exclusively in the E extract. The processed NH₂-terminal fragment of DMP1 was present in G1 and E extracts, whereas the COOH-terminal fragment of DMP1 existed mainly in the E extract. Bone sialoprotein was present in all three extracts of dentin and bone, whereas OPN was present only in the G1 and E extracts of bone. The difference in the distribution of the SIBLING proteins between organic and inorganic phases supports the belief that these molecular species play different roles in dentinogenesis and osteogenesis.