核转录因子激活蛋白-1在牙胚发育中作用的实验研究
|
摘要
转录因子是一类可以通过与下游靶DNA结合，从而调控下游基因转录的分子，它们接受来自生长因子等方面的信号，再将这些指令具体地传递给直接执行功能的蛋白分子。核转录因子激活蛋白-1(AP-1)就是其中一类。近年来国内外学者发现AP-1在牙齿发育过程中有表达，而且牙齿特异性蛋白如牙本质涎磷蛋白(DSPP)、釉原蛋白(Am)、牙本质基质蛋白(DMP1)等基因的启动子上存在AP-1的结合位点。
AP-1最常见、最典型的形式是由c-fos和c-jun组成的二聚体形式，因此本研究从c-fos和c-jun切入，采用免疫组化、蛋白印迹、细胞培养等方法，研究AP-1在牙胚发育过程中的作用，观察人牙乳头细胞内c-fos和c-jun在生长因子BMP2、TGF-β1信号转导中的作用，以及生长因子对二者表达水平的调控方式，旨在从细胞内信号转导水平探讨成牙本质细胞分化和牙本质形成的分子机理，为进一步深入研究AP-1在牙齿发育过程中的分子调控机理提供一定的实验依据。研究内容及结果如下：
1．c-fos和c-jun在鼠牙胚发育过程中的表达
第四军医大学硕士学位论文
根据大鼠、小鼠牙胚发育时序，采用免疫组化方法观察大鼠、小鼠牙
胚发育不同阶段c一fos和c一jun的表达特性。结果显示:从表达模式来看，
大鼠和小鼠没有明显差异，而c一fos和c一jun表达略有不同。c一fos在牙
胚蓄状期呈阴性表达，帽状期表达广泛，成釉器各层细胞均有表达，其中
以内釉上皮表达最强。c一jun在鼠牙胚蕾状期、帽状期呈阴性表达。钟状
期，前成牙本质细胞、前成釉细胞及功能型成牙本质细胞和成釉细胞中
c一fos和c一jun均为阳性表达，且随着矿化组织的形成表达逐渐减弱。二
者在中间层亦呈强阳性表达，牙乳头细胞和星网层细胞弱阳性表达。总之，
c一fos和c一jun的表达随牙胚发育成熟有减弱的趋势，提示二者可能参与
了成牙本质细胞、成釉细胞的增殖分化以及上皮一间充质间信号转导过程。
2.c一fos和c一jun在人牙胚发育过程中的表达
利用免疫组化技术观察c一fos和c一jun在人牙胚发育不同时期的表
达。结果显示，二者的表达与鼠牙胚中的表达相似，但明显滞后于鼠.钟
状期以前c一fos、c一jun均为阴性表达。钟状期c一fos和c一jun在前成牙
本质细胞和前成釉细胞中呈阴性表达:但随着细胞分化和基质分泌，c一fos
和c一jun表达逐渐增强，且阳性部位多集中在成釉器的远中侧。牙本质形
成期，二者在成牙本质细胞中强阳性表达。结果提示c一fos和c一jim参与
了调控成牙本质细胞、成釉细胞的核内基因表达过程，以及牙本质形成过
程中细胞外基质蛋白的合成分泌。
3.BMPZ、TGF一pl作用下人牙乳头细胞。一fos和e一jun转位变化
本实验通过免疫组化和图像分析法半定蛋观察特定浓度的BMPZ、TGF-
pl在不同时间点对人牙乳头细胞c一fos、c一jun表达的影响。取生长良
好的第五代人牙乳头细胞，分为实验组与对照组。对照组人牙乳头细胞胞
浆c一fos、c一jun表达阴性，胞核不表达或弱阳性表达。实验组分别以
Zoong/mlBMpZ、sng/mlTGF一日1作用于人牙乳头细胞。刺激30min，胞核、
胞浆均有阳性着色;lh胞核着色明显加深;2h胞核着色达到最强，随后
逐渐渐弱;6h胞核仍呈阳性着色，但较2h弱;至12h胞核着色明显变浅，
24h基本已无阳性着色。结果说明BMPZ、TGF一旦1呈时间依赖性诱导人牙
第四军医大学硕士学位论文
乳头细胞c一fos和c一jun表达，整个过程约持续24小时。
4.BMPZ、TGF一pl作用下人牙乳头细胞内c一fos和c一jun蛋白表达水平
的变化
本组实验采用We Stern一blot方法，对BMPZ(200ng/ml)、TGF一日1(sng/ml)
分别作用于人牙乳头细胞zh、Zh、6h、12h、24h、48h后e一fos、e一jun
蛋白水平的表达变化进行研究。结果显示:细胞内c一fos、c一jim蛋白表
达水平在BMPZ、TGF一pl作用1h内有显著增加，Zh达峰值，在2h一12h
内蛋白表达水平逐渐渐弱，24h表达基本消失。提示:BMPZ、TGF一日1通
过增加c一fos和c一jun的蛋白表达数t来发挥作用。
由于蛋白质的表达是其功能的指征，因而牙齿发育过程中c一fos和
c一jun蛋白特异性的时空表达模式说明，转录因子AP一在成牙本质细胞
和成釉细胞分化、基质分泌及矿化阶段发挥着转录调节作用。BMPZ、TG卜
pl可诱导人牙乳头细胞c一fos、c一jun表达。本研究从体内、体外两方
面证实了AP一1在牙齿发育中的作用，为在转录水平深入探讨AP一1与牙齿
发育密切相关的DSPP、Am等相互作用的分子机理莫定一定的理论基础，
为最终实现牙再生提供重要的实验依据。
Transcription factors can control the transcription of their target genes when they bind to the DNA-binding sites on their promotors. They can transfer the messages of growth factors and so on to those effective molecules. Nuclear transcriptor activator protein-1(AP-1) is a transcription . factor of this kind. Some researchers, both at home and abroad, have found recently that AP-1 is expressed during tooth development and that there are AP-1 binding sites on the promotor of DSPP, Am and DMP1 genes which are expressed in tooth development.
Since the most typical complex of AP-1 is formed by c-fos and c-jun, we aimed at exploring the effect of c-fos and c-jun on teeth germ development and mineraliztion by cell culture in vitro, Western-blot, immunohistochemistry, etc. In order
to gain an insight into the molecular mechanisms of odontoblast differentiation and dentinogenesis and to give more groundwork for further research, the author sought to characterize the roles of c-fos and c-jun in BMP2 and TGF-B1 signaling, investigate the modulation of c-fos and c-jun expression by BMP2 and TGF-B1 in cultured human dental papilla cells in this study. The main content and results of the study are presented as follows.
1. The expression of c-fos and c-jun during rat and mice tooth development
The expression of c-fos and c-jun was observed on different stages during rat and mice tooth development with immunohisto-chemical technique. Results: The temporal and spatial pattern of c-fos and c-jun expression during rat tooth development was similar to that of mice. There was a little difference between the expression of c-fos and c-jun, however, c-fos was found negative during the bud stage, but stained positive in dental epithelium during the cap stage, especially in the inner enamel epithelia. There was no c-jun protein during the bud and cup stages. During the bell stage, c-fos and c-jun was mainly detected in the pre- and functional odontoblasts and ameloblasts. Furthermore c-fos and c-jun had decreasing intensity with the mineralization, c-fos and c-jun were also expressed weakly in dental papilla and stellate reticulum. The stratum intermedium was strongly positive for c-fos and c-jun. In a word, the expression of c-fos and c-jun showed a decreasing intensity with the maturation of tooth germs.
2. The expression of c-fos and c-jun during human tooth development
The expression of c-fos and c-jun on different stages of human tooth development was observed with immunohischeraical method. The results of the experiment indicated the time diathesis of the above expression, which was similar to that of rat and mice tooth development. During human tooth development, the expression of c-fos and c-jun appeared later than that during rat and mice tooth development. There were no postive signals in the enamel organ before bell stage. During the bell stage, c-fos and c-jun was negative in the pre-odontoblasts and pre-ameloblasts. The intensity of c-fos and c-jun increased in odontoblasts and ameloblasts when matrix was initially secreted. Furthermore most of the c-fos and c-jun positive cells were on the posterior sides of enamel organ. These results suggested that c-fos and c-jun could play an important role during the differentiation of odontoblasts, ameloblasts, dentin matrix secretion and mineralization
3. The translocation of c-fos and c-jun in the human dental papilla cells treated by BMP2 and TGF-B1
The effects of BMP2 and TGF-B1 were observed half-rationally with immunohistochemical technique and image analysis. Human dental papilla cells were treated with BMP2 or TGF-B1 at different time points. The human dental papilla cells of passage 5 were selected and grouped into experimental and control groups for this experiment. The result of the study demonstrated that the cytoplasm of the cells in the control group was found with negative signals, but there were slightly positive signals in the nucleus of these cells. Positive signals were found in both the nucleus and cytoplasm when treated with 200ng/ml BMP2 or 5ng/ml TGF-B1
AP-1最常见、最典型的形式是由c-fos和c-jun组成的二聚体形式，因此本研究从c-fos和c-jun切入，采用免疫组化、蛋白印迹、细胞培养等方法，研究AP-1在牙胚发育过程中的作用，观察人牙乳头细胞内c-fos和c-jun在生长因子BMP2、TGF-β1信号转导中的作用，以及生长因子对二者表达水平的调控方式，旨在从细胞内信号转导水平探讨成牙本质细胞分化和牙本质形成的分子机理，为进一步深入研究AP-1在牙齿发育过程中的分子调控机理提供一定的实验依据。研究内容及结果如下：
1．c-fos和c-jun在鼠牙胚发育过程中的表达
第四军医大学硕士学位论文
根据大鼠、小鼠牙胚发育时序，采用免疫组化方法观察大鼠、小鼠牙
胚发育不同阶段c一fos和c一jun的表达特性。结果显示:从表达模式来看，
大鼠和小鼠没有明显差异，而c一fos和c一jun表达略有不同。c一fos在牙
胚蓄状期呈阴性表达，帽状期表达广泛，成釉器各层细胞均有表达，其中
以内釉上皮表达最强。c一jun在鼠牙胚蕾状期、帽状期呈阴性表达。钟状
期，前成牙本质细胞、前成釉细胞及功能型成牙本质细胞和成釉细胞中
c一fos和c一jun均为阳性表达，且随着矿化组织的形成表达逐渐减弱。二
者在中间层亦呈强阳性表达，牙乳头细胞和星网层细胞弱阳性表达。总之，
c一fos和c一jun的表达随牙胚发育成熟有减弱的趋势，提示二者可能参与
了成牙本质细胞、成釉细胞的增殖分化以及上皮一间充质间信号转导过程。
2.c一fos和c一jun在人牙胚发育过程中的表达
利用免疫组化技术观察c一fos和c一jun在人牙胚发育不同时期的表
达。结果显示，二者的表达与鼠牙胚中的表达相似，但明显滞后于鼠.钟
状期以前c一fos、c一jun均为阴性表达。钟状期c一fos和c一jun在前成牙
本质细胞和前成釉细胞中呈阴性表达:但随着细胞分化和基质分泌，c一fos
和c一jun表达逐渐增强，且阳性部位多集中在成釉器的远中侧。牙本质形
成期，二者在成牙本质细胞中强阳性表达。结果提示c一fos和c一jim参与
了调控成牙本质细胞、成釉细胞的核内基因表达过程，以及牙本质形成过
程中细胞外基质蛋白的合成分泌。
3.BMPZ、TGF一pl作用下人牙乳头细胞。一fos和e一jun转位变化
本实验通过免疫组化和图像分析法半定蛋观察特定浓度的BMPZ、TGF-
pl在不同时间点对人牙乳头细胞c一fos、c一jun表达的影响。取生长良
好的第五代人牙乳头细胞，分为实验组与对照组。对照组人牙乳头细胞胞
浆c一fos、c一jun表达阴性，胞核不表达或弱阳性表达。实验组分别以
Zoong/mlBMpZ、sng/mlTGF一日1作用于人牙乳头细胞。刺激30min，胞核、
胞浆均有阳性着色;lh胞核着色明显加深;2h胞核着色达到最强，随后
逐渐渐弱;6h胞核仍呈阳性着色，但较2h弱;至12h胞核着色明显变浅，
24h基本已无阳性着色。结果说明BMPZ、TGF一旦1呈时间依赖性诱导人牙
第四军医大学硕士学位论文
乳头细胞c一fos和c一jun表达，整个过程约持续24小时。
4.BMPZ、TGF一pl作用下人牙乳头细胞内c一fos和c一jun蛋白表达水平
的变化
本组实验采用We Stern一blot方法，对BMPZ(200ng/ml)、TGF一日1(sng/ml)
分别作用于人牙乳头细胞zh、Zh、6h、12h、24h、48h后e一fos、e一jun
蛋白水平的表达变化进行研究。结果显示:细胞内c一fos、c一jim蛋白表
达水平在BMPZ、TGF一pl作用1h内有显著增加，Zh达峰值，在2h一12h
内蛋白表达水平逐渐渐弱，24h表达基本消失。提示:BMPZ、TGF一日1通
过增加c一fos和c一jun的蛋白表达数t来发挥作用。
由于蛋白质的表达是其功能的指征，因而牙齿发育过程中c一fos和
c一jun蛋白特异性的时空表达模式说明，转录因子AP一在成牙本质细胞
和成釉细胞分化、基质分泌及矿化阶段发挥着转录调节作用。BMPZ、TG卜
pl可诱导人牙乳头细胞c一fos、c一jun表达。本研究从体内、体外两方
面证实了AP一1在牙齿发育中的作用，为在转录水平深入探讨AP一1与牙齿
发育密切相关的DSPP、Am等相互作用的分子机理莫定一定的理论基础，
为最终实现牙再生提供重要的实验依据。
Transcription factors can control the transcription of their target genes when they bind to the DNA-binding sites on their promotors. They can transfer the messages of growth factors and so on to those effective molecules. Nuclear transcriptor activator protein-1(AP-1) is a transcription . factor of this kind. Some researchers, both at home and abroad, have found recently that AP-1 is expressed during tooth development and that there are AP-1 binding sites on the promotor of DSPP, Am and DMP1 genes which are expressed in tooth development.
Since the most typical complex of AP-1 is formed by c-fos and c-jun, we aimed at exploring the effect of c-fos and c-jun on teeth germ development and mineraliztion by cell culture in vitro, Western-blot, immunohistochemistry, etc. In order
to gain an insight into the molecular mechanisms of odontoblast differentiation and dentinogenesis and to give more groundwork for further research, the author sought to characterize the roles of c-fos and c-jun in BMP2 and TGF-B1 signaling, investigate the modulation of c-fos and c-jun expression by BMP2 and TGF-B1 in cultured human dental papilla cells in this study. The main content and results of the study are presented as follows.
1. The expression of c-fos and c-jun during rat and mice tooth development
The expression of c-fos and c-jun was observed on different stages during rat and mice tooth development with immunohisto-chemical technique. Results: The temporal and spatial pattern of c-fos and c-jun expression during rat tooth development was similar to that of mice. There was a little difference between the expression of c-fos and c-jun, however, c-fos was found negative during the bud stage, but stained positive in dental epithelium during the cap stage, especially in the inner enamel epithelia. There was no c-jun protein during the bud and cup stages. During the bell stage, c-fos and c-jun was mainly detected in the pre- and functional odontoblasts and ameloblasts. Furthermore c-fos and c-jun had decreasing intensity with the mineralization, c-fos and c-jun were also expressed weakly in dental papilla and stellate reticulum. The stratum intermedium was strongly positive for c-fos and c-jun. In a word, the expression of c-fos and c-jun showed a decreasing intensity with the maturation of tooth germs.
2. The expression of c-fos and c-jun during human tooth development
The expression of c-fos and c-jun on different stages of human tooth development was observed with immunohischeraical method. The results of the experiment indicated the time diathesis of the above expression, which was similar to that of rat and mice tooth development. During human tooth development, the expression of c-fos and c-jun appeared later than that during rat and mice tooth development. There were no postive signals in the enamel organ before bell stage. During the bell stage, c-fos and c-jun was negative in the pre-odontoblasts and pre-ameloblasts. The intensity of c-fos and c-jun increased in odontoblasts and ameloblasts when matrix was initially secreted. Furthermore most of the c-fos and c-jun positive cells were on the posterior sides of enamel organ. These results suggested that c-fos and c-jun could play an important role during the differentiation of odontoblasts, ameloblasts, dentin matrix secretion and mineralization
3. The translocation of c-fos and c-jun in the human dental papilla cells treated by BMP2 and TGF-B1
The effects of BMP2 and TGF-B1 were observed half-rationally with immunohistochemical technique and image analysis. Human dental papilla cells were treated with BMP2 or TGF-B1 at different time points. The human dental papilla cells of passage 5 were selected and grouped into experimental and control groups for this experiment. The result of the study demonstrated that the cytoplasm of the cells in the control group was found with negative signals, but there were slightly positive signals in the nucleus of these cells. Positive signals were found in both the nucleus and cytoplasm when treated with 200ng/ml BMP2 or 5ng/ml TGF-B1
引文
1 金岩 口腔颌面组织胚胎学 陕西科学技术出版 2002，150-2
2 Ruch JV, Lesot H, Begue-Kirn C. Odontoblast differentiation. Int J Der Biol. 1995, 39(1): 51-68
3 Osman M, Ruch JV. Behavior of odontoblasts and basal lamina of trypsin or EDTA-isolated mouse dental papillae in short-term culture. J Dent Res. 1981, 60(6): 1015-27
4 Sasaki T, Garant PR. Structure and organization of odontoblasts. Anat Rec. 1996, 245(2): 235-49
5 Kardos TB, Hunter AR, Hanlin SM. et al. Odontoblast differentiation: a response to environmental calcium? Endod Dent Traumatol. 1998, 14(3): 105-11
6 Thesleff I. Genetic basis of tooth development and dental defects. Acta Odontol Scand. 2000, 58(5): 191-4.
7 D'Souza RN, Aberg T, Gaikwad J et al. Cbfal is required for epithelial-mesenchymal inter actions regulating tooth development in mice. Development 1999, 126(13): 2911
8 Heikinheimo Ks. Tage-specific expression of dcapentaplegic-Vg-Related genes 2, 4 and 6 (bone morphogenetic proteins 2, 4, and 6) during human tooth morphogenesis. J Dent Res 1994, 73(3): 590
9 Vainio S, Karavanova I, Jowett A, et al. Identification of BMP-4 as a signal mediating secondary induction between epithelial and mesenchymal tissues during early tooth development. Cell 1993 75(1): 45-58
10 Thesleff I, Aberg T. Molecular regulation of tooth development. Bone. 1999, 25(1): 123-5
11 Thesleff I, Vaahtokari A, Kettunen P, et al. Epithelial-mesenchymal
signaling during tooth development. Connect Tissue Res. 1995, 32: 9
12 高玉好，杨连甲，张绍章等。骨形成蛋白诱导牙本质形成的实验研究。实用口腔医学杂志，1989，59(4)：250-1
13 Tziafas D. Mechanisms controlling secondary initiation of dentinogenesis: a review. Int Endod J. 1994, 27(2): 61-74
14 Boyan BD, Sylvia VL, Dean DD et al. Cell biology of calcified tissues: experimental models of differentiation and mechanisms by which local and systemic factors exert their effects. Connect Tissue Res. 1996, 35(1-4): 63-70
15 Mitsiadis TA, Hirsinger E, Lendahl U et al. Delta-notch signaling in odontogenesis: correlation with cytodifferen tiation and evidence for feedback regulation. Dev Biol. 1998, 204(2): 420-31
16 Ito Y, Zhao J, Mogharei A et al. Antagonistic effects of Smad2 versus Smad7 are sensitive to their expression level during tooth development. J Biol Chem. 2001, 276(47): 44163-72
17 Holland PW, Hogan BL. Expression of homeo box genes during mouse development: a review. Genes Dev. 1988, 2(7): 773-82
18 Satokata I, Maas R. Msxl deficient mice exhibit cleft palate and abnormalities of craniofacial and tooth development. Nat Genet. 1994, 6(4): 348-56
19 Qiu M, Bulfone A, Ghattas I, Meneses JJ, Christensen L, Sharpe PT, Presley R, Pedersen RA, Rubenstein JL. Role of the Dlx homeobox genes in proximodistal Patterning of the branchial arches: mutations of Dlx-1, Dlx-2, and Dlx-1 and -2 alter morphogenesis of proximal skeletal and soft tissue structures derived from the first and second arches. Der Biol. 1997, 185(2): 165-84
20 Bronckers AL, Engelse MA, Cavender A, et al. Cell-specific
patterns of Cbfal mRNA and protein expression in postnatal murine dental tissues. Mech Der 2001, 101(1-2): 255-8
21 Angel P, Karin M. The role of Jun, Fos and the AP-1 complex in cell-proliferation and transformation. Biochim Biophys Acta. 1991, 1072(2-3): 129-57.
22 Kouzarides T, Ziff E. The role of the leucine zipper in the fos-jun interaction. Nature. 1988, 336(6200): 646-51
23 Derynck R, Zhang Y, Feng X.F. Smads: transcriptional activators of TGF-β responses. Cell, 1998, 95: 737-40.
24 Shen X, Hu PP, Liberati NT, et al. TGF-beta-induced phosphorylation of Smad3 regulates its interaction with coactivator p3OO/CREB-bindingprotein. Mol Biol Cell. 1998, 9(12): 3309-19
25 Derynck R. TGF-beta-receptor-mediated signaling. Trends Biochem Sci. 1994, 19(12): 548-53
26 Lee W, et al. Natuer 1987: 325-68
27 Adcock IM. Transcription factors as activators of gene transcription: AP-1 and NF-kappa B. Monaldi Arch Chest Dis. 1997, 52(2): 178-86
28 K. Chida. et al. Nuclear translocation of Fos is stimulated by interaction with Jun through the leucine zipper. CMLS. 1999, (55): 297-302
29 Katai H. et al. An AP-1-like motif in the first intron of human Pro alpha 1(Ⅰ) collagen gene is a critical determinant of its transcriptional activity. Mol Cell Biochem. 1992, 118(2): 119-29
30 Curran T. et al. The AP-1 connection. Cell 1988, (55): 395.
31 Distel RJ, Spiegelman BM. Protooncogene c-fos as a transcription factor. Adv Cancer Res 1990, 55: 37-55
32 Foletta, Victora C. Transcription factor AP-1, and the role of Fra-2. Immu Cell Biol. 1996, 74(2): 121-33
33 Sheng M. et al. The regulation and function of c-fos and other immediate early genes in the nervous system. Neuron. 1990, 4(4): 477-85
34 Hattori, K. et al. Structure and chromosomal localization of the functional intronless human Jun protooncogene. Proc Natl Acad Sci U S A. 1988, 85(23): 9148-52.
35 Wisdom R. AP-1: one switch for many signals. Exp Cell Res. 1999, 253(1): 180-5
36 Nerlov C, Ziff EB. Three levels of functional interaction determine the activity of CCAAT/enhancer binding protein-alpha the serum albumin promoter. Genes Dev. 1994, 8(3): 350-62
37 Wang, Z-Q, et al. A novel target cell for c-fos-induced oncogensis: development of chondrogenic tumours in embryonic stem cell chimeras. EMBO J. 1991, (10): 2437-2450
38 Candeliere, G.A et al. Increased expression of the c-fos proto-oncogene in bone from patients with fibrous dysplasia. New Eng J Med. 1995, (332): 1546-51
39 Hilberg, F et al. c-Jun is essential for normal mouse development and hepatogensis. Nature, 1993, 179-81
40 Shaulian, E et al. AP-1 as a regulator of cell life and death. Nature Cell Biol. 2002, 131-36
41 Marry MacDougall. Genomic organization, chromosomal mapping, and promoter analysis of the mouse dentin sialophosphoprotein (aspp) gene, which codes for both dentin sialoprotein and dentin phosphoprotein. J Biol Chem. 1998, 273(16): 9457-64
42 Karthikeyan Narayananet al. Transcriptional regulation of dentin matrix protein 1 (DMP1) by AP-1 (c-fos/c-jun) factors. Con
tissue Res 2002, (43): 1-7
43 Strong DD, Merriman HL, Landale EC, Baylink DJ, Mohan S. The effects of the insulin-like growth factors and transforming growth factor beta on the Jun proto-oncogene family in MC3T3-E1 cells. Calcif Tissue Int. 1994, 55(4): 311-5
44 Janulis M, Silberman S, et al. Role of mitogen-activated protein kinases and c-Jun/AP-1 trans-activating activity in the regulation of protease mRNAs and the malignant phenotype in NIH 3T3 fibroblasts. J Biol Chem. 1999, 274(2): 801-13
45 Janknecht R, Cahill MA, Nordheim A. Signal integration at the c-fos promoter. Carcinogenesis. 1995, 16: 443-50
46 Robertson LM, Kerppola TK, Vendrell M, Luk D, Smeyne RJ, Bocchiaro C, Morgan JI, Curran T. Regulation of c-fos expression in transgenic mice requires multiple interdependent transcription control elements. Neuron 1995, 14: 241-52
47 SE Rutberg, TL Adams, M Olive, N Alexander, C Vinson, SH Yuspa. CRE DNA binding proteins bind to the AP-1 target sequence and suppress AP-1 transcriptional activity in mouse keratinocytes. Oncogene. 1999, 18(8): 1569-79
48 Ginty DD, Bonni A, et al. Nerve growth factor activates a Rasdependent protein kinase that stimulates c-fos transcription via phosphorylation of CREB. 1994, 7: 713-25
49 Price MA, Rogers AE, Treisman R. Comparative analysis of the ternarty complex factors Elk-1, SAP-1a and AP-2(ERP/NET). EMBO J. 1995, 14: 2589-2601
50 Gille H, Kortenjann M, Thomae O, et al. ERK phosphorylation potentiates Elk-1-mediated ternary complex formation and transactivation. EMBO J. 1995, 14: 951-62
51 Gille H, Strahl T, Shaw PE. Activation of ternary omplex actor
Elk-1 by stress-activated protein kinases. Curr Biol. 1995, 5: 1191-1200
52 Price MA, Rogers AE, Treisman R. Comparative analysis of the ternary complex factors Elk-1, SAP-1a and SAP-2 (ERP/NET). EMBO J, 1995, 14(11): 2589-2601
53 Whitmarsh AJ, Shore P, et al. Integration of MAP kinase signal transduction pathways at the serum response element. Science, 1995, 269: 403-7
54 Hill CS, Treisman R Differential activation of c-fos promoter elements by serum, lysophosphatidic acid, G proteins and polypeptide growth factors. EMBO J 1995, 14: 5037-47
55 Angel P. The role and regulation of the Jun proteins in response to phorbol ester and UV light. In: Baeuerle PA (ed) Inducible gene expression, Birkh(?)user, Boston, 1995, 63-92
56 Cbiu R, Boyle WJ, Meek J, Smeal T, Hunter T, Karin M. The c-Fos protein interacts with c-Jun/AP-1 to stimulate trenscription of AP-1 responsive genes. Cell. 1988 Aug 12; 54(4): 541-52
57 Deng T, Karin M. c-Fos transcriptional activity stimulated by H-Ras-activated protein kinase distinct from JNK and ERK. Nature (London) 1994, 371: 171-5
58 Arias J, Alberts AS, Brindle P, Claret F-X, Smeal T, Karin M. Activation of cAMP and mitogen responsive genes relies on a common nuclear factor. Nature (London). 1994, 370: 226-9
59 Smeal T, Hibi M, Karin M. Altering the specificity of signal transduction cascades: positive regulation of c-Jun transcriptional activity by protein kinase A. EMBO J. 1994, 13: 6006-10
60 Karin M, Hunter T. Transcriptional control by protein phosphorylation: signal transmission from the cell surface to the nucleus. Curr Biol. 1995, 5: 747-57
61 Byers MR et al. Focal c-fos expression in developing rat molars: correlations with subsequent intradental and epithelial sensory innervation. Int J Der Biol. 1995, 39(1): 181-89
62 G.S. Wise et al. Effects of epidermal growth factor(EGF) and colony-stimulating factor-1 (CSF-1) on expression of c-fos in rat mandibular molars: implications for tooth eruption. Cell Tissue Res. 1996, (284): 1-7
63 C. Kitamura et al. Expression of c-jun and jun-B proto-oncogene in odontoblasts during development of bovine tooth germs. J Dent Res, 1997, 76(4): 822-30
64 C. Kitamura et al. Temporal and spatial expression of c-jun and jun-B pro-oncogenes in pulp cells involved with reparative dentinogenesis after cavity preparation of rat molars. J Dent Res. 1997, 78(2): 673-80
65 Sumio Kishikawa et al. Localization transcription factor AP-1 family proteins in ameloblast nuclei of the rat incisor. J Histochem Cytochem. 2002, (48): 1511-20
66 Feng JQ, Luan X, Wallace J et al. Genomic organization, chromosomal mapping, and promoter analysis of the mouse dentin sialophosphoprotein (Dspp) gene, which codes for both dentin sialoprotein and dentin phosphoprotein. J Biol Chem. 1998, 273(16): 9457-64
67 Satokata I. and Maas R. Msx-1 deficient mice exhibit cleft palate and abnormalities of craniofacial and tooth development. Nat. Genet. 1994, 6: 348-56
68 Keranen SV, Aberg T, Kettunen P, Thesleff I, Jernvall J. Association of developmental regulatory genes with the development of different molar tooth shapes in two species of rodents. Dev Genes Evol. 1998, 208(9): 477-86
69 Ferrari D, Lichtler AC, Pan ZZ. et al. Ectopic expression of Msx-2 in posterior limb bud mesoderm impairs limb morphogenesis while inducing BMP-4 expression, inhibiting cell proliferation, and promoting apoptosis. Dev Biol. 1998, 197(1): 12-24
70 Bei M, Kratochwil K, Maas RL. BMP4 rescues a non-cell-autonomous function of Msxl in tooth development. Development. 2000, 127(21): 4711-8
71 Kratochwil K, Dull M, Farinas I, Galceran J, Grosschedl R. Lefl expression is activated by BMP-4 and regulates inductive tissue interactions in tooth and hair development. Genes Dev. 1996, 10(11): 1382-94
72 Wang E, Lee MJ, Pandey S. Control of fibroblast senescence and activation of programmed cell death. J Cell Biochem. 1994, 54(4): 432-9
73 Riabowol K, Schiff J, Gilman MZ. Transcription factor AP-1 activity is required for initiation of DNA synthesis and is lost during cellular aging. Proc Natl Acad Sci U S A. 1992, 89(1): 157-61
74 Fantozzi I, Zhang S, Platoshyn O, Remillard CV, Cowling RT, Yuan JX. Hypoxia increases AP-1 binding activity by enhancing capacitative Ca2+ entry in human pulmonary artery endothelial cells. Am J Physiol Lung Cell Mol Physiol. 2003, 285(6): L1233-45
75 McCabe LR, Kockx M, Lian J, Stein J, Stein G. Selective expression of fos- and jun-related genes during osteoblast proliferation and differentiation. Exp Cell Res. 1995, 218(1): 255-62
76 陈璐璐等 氟化钠对乳鼠成骨细胞c-fos、c-jun基因表达及细胞增殖的影响。中华预防医学杂志2000，34(6)：327-9
77 Slavkin HC. Molecular determinatiions during dental morphogenesis
and cytodifferentiation: A review. J Craniofac Genet Dev Biol. 1991, 11: 338
78 Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor Laboratory Press, 1989
79 Joan Massagu(?), David Wotton. Transcriptional control by the TGF-β/Smad signaling system. EMBO J. 2000, 9: 1745-54
80 Lai CF, Cheng SL. Signal transductions induced by bone morphogenetic protein-2 and transforming growth factor-beta in normal human osteoblastic cells. J Bio1Chem 2002, 277: 15514-22
81 Yamamoto N, Akiyama S, Katagiri T, et al. Smadl and smad5 act downstream of intracellular signalings of BMP-2 that inhibits myogenic differentiation and induces osteoblast differentiation in C2C12 my oblasts. Biochem Biophys Res Commun. 1997, 238: 574-80
82 Yanagisawa K, Osada H, Masuda A, et al. Induction of apoptosis by Smad3 and down-regulation of Smad3 expression in response to TGF-beta in human normal lung epithelial cells. Oncogene 1998, 17(13): 1743-7
83 Poncelet AC, De Caestecker MP, Schnaper HW. The transforming growth factor-beta/SMAD signaling pathway is present and functional in human mesangial cells. Kidney Int. 1999, 56(4): 1354-65
84 Liberati NT, Datto MB, Frederick JP, et al. Smads bind directly to the Jun family of AP-1 transcription factors. Proc Natl Acad Sci U S A 1999, 96(9): 4844-9
2 Ruch JV, Lesot H, Begue-Kirn C. Odontoblast differentiation. Int J Der Biol. 1995, 39(1): 51-68
3 Osman M, Ruch JV. Behavior of odontoblasts and basal lamina of trypsin or EDTA-isolated mouse dental papillae in short-term culture. J Dent Res. 1981, 60(6): 1015-27
4 Sasaki T, Garant PR. Structure and organization of odontoblasts. Anat Rec. 1996, 245(2): 235-49
5 Kardos TB, Hunter AR, Hanlin SM. et al. Odontoblast differentiation: a response to environmental calcium? Endod Dent Traumatol. 1998, 14(3): 105-11
6 Thesleff I. Genetic basis of tooth development and dental defects. Acta Odontol Scand. 2000, 58(5): 191-4.
7 D'Souza RN, Aberg T, Gaikwad J et al. Cbfal is required for epithelial-mesenchymal inter actions regulating tooth development in mice. Development 1999, 126(13): 2911
8 Heikinheimo Ks. Tage-specific expression of dcapentaplegic-Vg-Related genes 2, 4 and 6 (bone morphogenetic proteins 2, 4, and 6) during human tooth morphogenesis. J Dent Res 1994, 73(3): 590
9 Vainio S, Karavanova I, Jowett A, et al. Identification of BMP-4 as a signal mediating secondary induction between epithelial and mesenchymal tissues during early tooth development. Cell 1993 75(1): 45-58
10 Thesleff I, Aberg T. Molecular regulation of tooth development. Bone. 1999, 25(1): 123-5
11 Thesleff I, Vaahtokari A, Kettunen P, et al. Epithelial-mesenchymal
signaling during tooth development. Connect Tissue Res. 1995, 32: 9
12 高玉好，杨连甲，张绍章等。骨形成蛋白诱导牙本质形成的实验研究。实用口腔医学杂志，1989，59(4)：250-1
13 Tziafas D. Mechanisms controlling secondary initiation of dentinogenesis: a review. Int Endod J. 1994, 27(2): 61-74
14 Boyan BD, Sylvia VL, Dean DD et al. Cell biology of calcified tissues: experimental models of differentiation and mechanisms by which local and systemic factors exert their effects. Connect Tissue Res. 1996, 35(1-4): 63-70
15 Mitsiadis TA, Hirsinger E, Lendahl U et al. Delta-notch signaling in odontogenesis: correlation with cytodifferen tiation and evidence for feedback regulation. Dev Biol. 1998, 204(2): 420-31
16 Ito Y, Zhao J, Mogharei A et al. Antagonistic effects of Smad2 versus Smad7 are sensitive to their expression level during tooth development. J Biol Chem. 2001, 276(47): 44163-72
17 Holland PW, Hogan BL. Expression of homeo box genes during mouse development: a review. Genes Dev. 1988, 2(7): 773-82
18 Satokata I, Maas R. Msxl deficient mice exhibit cleft palate and abnormalities of craniofacial and tooth development. Nat Genet. 1994, 6(4): 348-56
19 Qiu M, Bulfone A, Ghattas I, Meneses JJ, Christensen L, Sharpe PT, Presley R, Pedersen RA, Rubenstein JL. Role of the Dlx homeobox genes in proximodistal Patterning of the branchial arches: mutations of Dlx-1, Dlx-2, and Dlx-1 and -2 alter morphogenesis of proximal skeletal and soft tissue structures derived from the first and second arches. Der Biol. 1997, 185(2): 165-84
20 Bronckers AL, Engelse MA, Cavender A, et al. Cell-specific
patterns of Cbfal mRNA and protein expression in postnatal murine dental tissues. Mech Der 2001, 101(1-2): 255-8
21 Angel P, Karin M. The role of Jun, Fos and the AP-1 complex in cell-proliferation and transformation. Biochim Biophys Acta. 1991, 1072(2-3): 129-57.
22 Kouzarides T, Ziff E. The role of the leucine zipper in the fos-jun interaction. Nature. 1988, 336(6200): 646-51
23 Derynck R, Zhang Y, Feng X.F. Smads: transcriptional activators of TGF-β responses. Cell, 1998, 95: 737-40.
24 Shen X, Hu PP, Liberati NT, et al. TGF-beta-induced phosphorylation of Smad3 regulates its interaction with coactivator p3OO/CREB-bindingprotein. Mol Biol Cell. 1998, 9(12): 3309-19
25 Derynck R. TGF-beta-receptor-mediated signaling. Trends Biochem Sci. 1994, 19(12): 548-53
26 Lee W, et al. Natuer 1987: 325-68
27 Adcock IM. Transcription factors as activators of gene transcription: AP-1 and NF-kappa B. Monaldi Arch Chest Dis. 1997, 52(2): 178-86
28 K. Chida. et al. Nuclear translocation of Fos is stimulated by interaction with Jun through the leucine zipper. CMLS. 1999, (55): 297-302
29 Katai H. et al. An AP-1-like motif in the first intron of human Pro alpha 1(Ⅰ) collagen gene is a critical determinant of its transcriptional activity. Mol Cell Biochem. 1992, 118(2): 119-29
30 Curran T. et al. The AP-1 connection. Cell 1988, (55): 395.
31 Distel RJ, Spiegelman BM. Protooncogene c-fos as a transcription factor. Adv Cancer Res 1990, 55: 37-55
32 Foletta, Victora C. Transcription factor AP-1, and the role of Fra-2. Immu Cell Biol. 1996, 74(2): 121-33
33 Sheng M. et al. The regulation and function of c-fos and other immediate early genes in the nervous system. Neuron. 1990, 4(4): 477-85
34 Hattori, K. et al. Structure and chromosomal localization of the functional intronless human Jun protooncogene. Proc Natl Acad Sci U S A. 1988, 85(23): 9148-52.
35 Wisdom R. AP-1: one switch for many signals. Exp Cell Res. 1999, 253(1): 180-5
36 Nerlov C, Ziff EB. Three levels of functional interaction determine the activity of CCAAT/enhancer binding protein-alpha the serum albumin promoter. Genes Dev. 1994, 8(3): 350-62
37 Wang, Z-Q, et al. A novel target cell for c-fos-induced oncogensis: development of chondrogenic tumours in embryonic stem cell chimeras. EMBO J. 1991, (10): 2437-2450
38 Candeliere, G.A et al. Increased expression of the c-fos proto-oncogene in bone from patients with fibrous dysplasia. New Eng J Med. 1995, (332): 1546-51
39 Hilberg, F et al. c-Jun is essential for normal mouse development and hepatogensis. Nature, 1993, 179-81
40 Shaulian, E et al. AP-1 as a regulator of cell life and death. Nature Cell Biol. 2002, 131-36
41 Marry MacDougall. Genomic organization, chromosomal mapping, and promoter analysis of the mouse dentin sialophosphoprotein (aspp) gene, which codes for both dentin sialoprotein and dentin phosphoprotein. J Biol Chem. 1998, 273(16): 9457-64
42 Karthikeyan Narayananet al. Transcriptional regulation of dentin matrix protein 1 (DMP1) by AP-1 (c-fos/c-jun) factors. Con
tissue Res 2002, (43): 1-7
43 Strong DD, Merriman HL, Landale EC, Baylink DJ, Mohan S. The effects of the insulin-like growth factors and transforming growth factor beta on the Jun proto-oncogene family in MC3T3-E1 cells. Calcif Tissue Int. 1994, 55(4): 311-5
44 Janulis M, Silberman S, et al. Role of mitogen-activated protein kinases and c-Jun/AP-1 trans-activating activity in the regulation of protease mRNAs and the malignant phenotype in NIH 3T3 fibroblasts. J Biol Chem. 1999, 274(2): 801-13
45 Janknecht R, Cahill MA, Nordheim A. Signal integration at the c-fos promoter. Carcinogenesis. 1995, 16: 443-50
46 Robertson LM, Kerppola TK, Vendrell M, Luk D, Smeyne RJ, Bocchiaro C, Morgan JI, Curran T. Regulation of c-fos expression in transgenic mice requires multiple interdependent transcription control elements. Neuron 1995, 14: 241-52
47 SE Rutberg, TL Adams, M Olive, N Alexander, C Vinson, SH Yuspa. CRE DNA binding proteins bind to the AP-1 target sequence and suppress AP-1 transcriptional activity in mouse keratinocytes. Oncogene. 1999, 18(8): 1569-79
48 Ginty DD, Bonni A, et al. Nerve growth factor activates a Rasdependent protein kinase that stimulates c-fos transcription via phosphorylation of CREB. 1994, 7: 713-25
49 Price MA, Rogers AE, Treisman R. Comparative analysis of the ternarty complex factors Elk-1, SAP-1a and AP-2(ERP/NET). EMBO J. 1995, 14: 2589-2601
50 Gille H, Kortenjann M, Thomae O, et al. ERK phosphorylation potentiates Elk-1-mediated ternary complex formation and transactivation. EMBO J. 1995, 14: 951-62
51 Gille H, Strahl T, Shaw PE. Activation of ternary omplex actor
Elk-1 by stress-activated protein kinases. Curr Biol. 1995, 5: 1191-1200
52 Price MA, Rogers AE, Treisman R. Comparative analysis of the ternary complex factors Elk-1, SAP-1a and SAP-2 (ERP/NET). EMBO J, 1995, 14(11): 2589-2601
53 Whitmarsh AJ, Shore P, et al. Integration of MAP kinase signal transduction pathways at the serum response element. Science, 1995, 269: 403-7
54 Hill CS, Treisman R Differential activation of c-fos promoter elements by serum, lysophosphatidic acid, G proteins and polypeptide growth factors. EMBO J 1995, 14: 5037-47
55 Angel P. The role and regulation of the Jun proteins in response to phorbol ester and UV light. In: Baeuerle PA (ed) Inducible gene expression, Birkh(?)user, Boston, 1995, 63-92
56 Cbiu R, Boyle WJ, Meek J, Smeal T, Hunter T, Karin M. The c-Fos protein interacts with c-Jun/AP-1 to stimulate trenscription of AP-1 responsive genes. Cell. 1988 Aug 12; 54(4): 541-52
57 Deng T, Karin M. c-Fos transcriptional activity stimulated by H-Ras-activated protein kinase distinct from JNK and ERK. Nature (London) 1994, 371: 171-5
58 Arias J, Alberts AS, Brindle P, Claret F-X, Smeal T, Karin M. Activation of cAMP and mitogen responsive genes relies on a common nuclear factor. Nature (London). 1994, 370: 226-9
59 Smeal T, Hibi M, Karin M. Altering the specificity of signal transduction cascades: positive regulation of c-Jun transcriptional activity by protein kinase A. EMBO J. 1994, 13: 6006-10
60 Karin M, Hunter T. Transcriptional control by protein phosphorylation: signal transmission from the cell surface to the nucleus. Curr Biol. 1995, 5: 747-57
61 Byers MR et al. Focal c-fos expression in developing rat molars: correlations with subsequent intradental and epithelial sensory innervation. Int J Der Biol. 1995, 39(1): 181-89
62 G.S. Wise et al. Effects of epidermal growth factor(EGF) and colony-stimulating factor-1 (CSF-1) on expression of c-fos in rat mandibular molars: implications for tooth eruption. Cell Tissue Res. 1996, (284): 1-7
63 C. Kitamura et al. Expression of c-jun and jun-B proto-oncogene in odontoblasts during development of bovine tooth germs. J Dent Res, 1997, 76(4): 822-30
64 C. Kitamura et al. Temporal and spatial expression of c-jun and jun-B pro-oncogenes in pulp cells involved with reparative dentinogenesis after cavity preparation of rat molars. J Dent Res. 1997, 78(2): 673-80
65 Sumio Kishikawa et al. Localization transcription factor AP-1 family proteins in ameloblast nuclei of the rat incisor. J Histochem Cytochem. 2002, (48): 1511-20
66 Feng JQ, Luan X, Wallace J et al. Genomic organization, chromosomal mapping, and promoter analysis of the mouse dentin sialophosphoprotein (Dspp) gene, which codes for both dentin sialoprotein and dentin phosphoprotein. J Biol Chem. 1998, 273(16): 9457-64
67 Satokata I. and Maas R. Msx-1 deficient mice exhibit cleft palate and abnormalities of craniofacial and tooth development. Nat. Genet. 1994, 6: 348-56
68 Keranen SV, Aberg T, Kettunen P, Thesleff I, Jernvall J. Association of developmental regulatory genes with the development of different molar tooth shapes in two species of rodents. Dev Genes Evol. 1998, 208(9): 477-86
69 Ferrari D, Lichtler AC, Pan ZZ. et al. Ectopic expression of Msx-2 in posterior limb bud mesoderm impairs limb morphogenesis while inducing BMP-4 expression, inhibiting cell proliferation, and promoting apoptosis. Dev Biol. 1998, 197(1): 12-24
70 Bei M, Kratochwil K, Maas RL. BMP4 rescues a non-cell-autonomous function of Msxl in tooth development. Development. 2000, 127(21): 4711-8
71 Kratochwil K, Dull M, Farinas I, Galceran J, Grosschedl R. Lefl expression is activated by BMP-4 and regulates inductive tissue interactions in tooth and hair development. Genes Dev. 1996, 10(11): 1382-94
72 Wang E, Lee MJ, Pandey S. Control of fibroblast senescence and activation of programmed cell death. J Cell Biochem. 1994, 54(4): 432-9
73 Riabowol K, Schiff J, Gilman MZ. Transcription factor AP-1 activity is required for initiation of DNA synthesis and is lost during cellular aging. Proc Natl Acad Sci U S A. 1992, 89(1): 157-61
74 Fantozzi I, Zhang S, Platoshyn O, Remillard CV, Cowling RT, Yuan JX. Hypoxia increases AP-1 binding activity by enhancing capacitative Ca2+ entry in human pulmonary artery endothelial cells. Am J Physiol Lung Cell Mol Physiol. 2003, 285(6): L1233-45
75 McCabe LR, Kockx M, Lian J, Stein J, Stein G. Selective expression of fos- and jun-related genes during osteoblast proliferation and differentiation. Exp Cell Res. 1995, 218(1): 255-62
76 陈璐璐等 氟化钠对乳鼠成骨细胞c-fos、c-jun基因表达及细胞增殖的影响。中华预防医学杂志2000，34(6)：327-9
77 Slavkin HC. Molecular determinatiions during dental morphogenesis
and cytodifferentiation: A review. J Craniofac Genet Dev Biol. 1991, 11: 338
78 Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor Laboratory Press, 1989
79 Joan Massagu(?), David Wotton. Transcriptional control by the TGF-β/Smad signaling system. EMBO J. 2000, 9: 1745-54
80 Lai CF, Cheng SL. Signal transductions induced by bone morphogenetic protein-2 and transforming growth factor-beta in normal human osteoblastic cells. J Bio1Chem 2002, 277: 15514-22
81 Yamamoto N, Akiyama S, Katagiri T, et al. Smadl and smad5 act downstream of intracellular signalings of BMP-2 that inhibits myogenic differentiation and induces osteoblast differentiation in C2C12 my oblasts. Biochem Biophys Res Commun. 1997, 238: 574-80
82 Yanagisawa K, Osada H, Masuda A, et al. Induction of apoptosis by Smad3 and down-regulation of Smad3 expression in response to TGF-beta in human normal lung epithelial cells. Oncogene 1998, 17(13): 1743-7
83 Poncelet AC, De Caestecker MP, Schnaper HW. The transforming growth factor-beta/SMAD signaling pathway is present and functional in human mesangial cells. Kidney Int. 1999, 56(4): 1354-65
84 Liberati NT, Datto MB, Frederick JP, et al. Smads bind directly to the Jun family of AP-1 transcription factors. Proc Natl Acad Sci U S A 1999, 96(9): 4844-9