Egr-1与骨桥蛋白在大鼠血管平滑肌细胞中的相关性及其机制的研究
摘要
Egr-1与骨桥蛋白在大鼠血管平滑肌细胞中的相关性及其机制的研究
前言
动脉粥样硬化性疾病、血管重建术后再狭窄(restenosis,RS)、高血压等血管重塑相关性疾病已经愈来愈严重地威胁着人类的健康。目前认为,血管中膜平滑肌细胞(smooth muscle cell,SMC)向内膜下迁移与增殖是血管重塑的主要病理基础之一。
研究表明,某些转录因子(transcription factors,TF)可以调节多个血管平滑肌细胞(vascular smooth muscle cell,VSMC)增殖相关基因的表达,从不同层面调控VSMC的迁移与增殖。同时,人们也逐渐注意到细胞外基质(extracellular matrix,ECM)在血管重塑过程中的重要地位。研究发现,ECM在各种外界刺激的作用下,可以作为细胞外信号,经跨膜信号传递系统到达核内,激活一系列调控VSMC增殖的TF,从而推动VSMC的分裂、迁移及增殖。TF与ECM在血管重塑的过程中密不可分,因此将二者有机地联系在一起,并对其相互作用关系进行深入探讨,将有助于全面了解血管重塑的细胞与分子机制。
早期生长反应因子-1(eally growth response factor-1,Egr-1)作为一种锌指结构TF,参与多种基因的调控,与细胞增殖关系密切。大量研究表明,Egr-1在介导VSMC迁移、增殖过程中发挥着重要的作用,成为目前血管重塑机制研究中的焦点。骨桥蛋白(osteopontin,OPN)作为ECM中一种重要的功能性蛋白,被认为是血管损伤重塑过程中重要的始动因素,应用抗OPN抗体可以抑制VSMC的表型转化、迁移和增殖。尽管Egr-1和OPN在介导VSMC迁移、增殖过程中均起着重要的作用,但二者之间的关系尚不清楚。
本课题拟在以往研究的基础上,对培养的大鼠主动脉VSMCs分别稳定转染Egr-1、OPN基因,并检测转染前后OPN与Egr-1的表达变化,从而分析二者的相关性。并应用染色质免疫沉淀(chromatin immunoprecipitation,ChIP)方法检测Egr-1与OPN启动子的结合情况,同时通过向OPN稳定转染细胞株中加入细胞外信号调节蛋白激酶(extracellular signal-regulated kinase,ERK)的磷酸化抑制剂,检测抑制ERK磷酸化后,OPN上调Egr-1的水平变化,从而对Egr-1与OPN相关性的内在机制进行深入探讨,并为抑制VSMC的迁移、增殖,控制血管壁的不适当重塑提供理论及实验基础。
材料与方法
1、大鼠VSMC的培养
大鼠A10主动脉VSMC细胞株购自ATCC细胞库,用含10%胎牛血清(fetalbovine serum,FBS)的DMEM培养基,在37℃、5%CO_2、饱和湿度的培养箱中培养,用0.25%的胰酶消化、传代。
2、质粒的提纯、扩增及稳定转染
委托TaKaRa公司对pCMV-Egr-1/NEO、pCMV(-)/NEO质粒及pET-28-rOPN/NEO、pET-28(-)/NEO质粒进行提纯、扩增及鉴定,其中pET-28-rOPN/NEO和pET-28(-)/NEO质粒被连于CMV启动子上,以便在VSMC中表达。
待培养的VSMCs生长到80%汇合度时,以100μg/ml～1mg/ml的新霉素(neomycin418,G418)浓度进行最低G418浓度筛选。当培养的VSMCs在6孔板内达到80%汇合度时,应用FuGENE6分别向VSMCs内转染pCMV-Egr-1/NEO、pCMV(-)/NEO质粒及pCMV-ET-28-rOPN/NEO、pCMV-ET-28(-)/NEO质粒。细胞转染24小时(hour,h)后,加入400μg/ml G418的培养液进行培养。2周后,采用有限稀释法将细胞传至96孔板中进行单克隆化培养(于含200μg/ml G418的条件培养液中),待长满后进行扩大培养,并检测细胞中Egr-1、OPNmRNA的表达。
3、应用ERK磷酸化抑制剂阻断ERK信号传导通路
当OPN稳定转染的VSMCs生长至80%汇合度时,向细胞培养液中加入10μMERK磷酸化抑制剂PD98059,继续培养24h后,进行Western blot检测。
4、反转录多聚酶链反应(reverse transcriptase polymerase chainreaction,RT-PCR)检测Egr-1、OPNmRNA
用Trizol Reagent提取VSMCs的总RNA。用RNA PCR Kit Ver.3.0扩增目的片段。PCR产物电泳后,用溴化乙啶(ethidium bromide,EB)染色,BioImaging Systems系统成像,用NIH image软件分析产物条带灰度,以Egr-1、OPN产物条带灰度值与甘油醛-3-磷酸脱氢酶(glyceralde-3-phosphate dehydrogena,GAPDH)产物条带灰度值的比值分别作为Egr-1、OPNmRNA的相对表达量。
5、Western blot检测Egr-1、OPN、ERK、P-ERK蛋白
提取细胞总蛋白,上样量为80μg。转印到PVDF膜上后用5%脱脂奶粉封闭,用Egr-1兔抗大鼠多克隆抗体(1:300)、OPN兔抗大鼠多克隆抗体(1:500)、ERK兔抗大鼠多克隆抗体(1:200)、磷酸化细胞外调节蛋白激酶(phosphoextracellularsignal-regulated kinase,P-ERK)兔抗大鼠多克隆抗体(1:200)及兔抗大鼠α-tubulin多克隆抗体(1:500)4℃孵育过夜,分别用相应的二抗37℃孵育2h,ECL发光1分钟,胶片中的蛋白条带经BioImaging Systems采集后进行灰度值测定。目的蛋白灰度值与内对照α-tubulin比值为其相对蛋白定量。
6、ChIP方法检测Egr-1与OPN启动子的结合
分别取2×10~6个对数生长期的稳定转染Egr-1的VSMCs及正常VSMCs,用甲醛交联,0.125M甘氨酸终止交联。超声剪切染色质脱氧核糖核酸(deoxyribonucleicacid,DNA)成500～1000个碱基对(basepair,bp)片段。分别取50μl全细胞抽提物作为实验和阴性对照样本。用ChIP缓冲液10倍稀释后各取其中10μl合并作为输入对照(input),在实验样本中加入5g Egr-1兔抗大鼠多克隆抗体(1:300),将实验样品及阴性对照样本4℃孵育过夜。几次洗涤后,复合物从珠子上洗脱下来。用酚氯仿抽提纯化DNA,最后应用于PCR分析。
7、统计学分析
采用SPSS11.0统计软件进行数据分析。Egr-1与OPN的相关性用双变量相关分析法(Spearman相关分析),不同组间的参数比较用One-way ANOVA分析,两两比较采用LSD方法,P＜0.05有统计学意义。
实验结果
1、RT-PCR结果显示:Egr-1稳定转染组的Egr-1mRNA较正常对照组和空载体组表达明显增强(p＜0.01);OPN稳定转染组OPNmRNA与正常对照组和空载体组比较,表达也明显增强(p＜0.01)。
2、RT-PCR及Western blot方法检测Egr-1和OPN稳定转染VSMCs后OPN和Egr-1的表达水平。结果显示:与正常对照组和空载体组比较,Egr-1转染组的OPNmRNA及蛋白表达均随着Egr-1表达的增强而明显上调(p＜0.01);同样,OPN稳定转染组的Egr-1mRNA及蛋白表达较正常对照组和空载体组明显升高(p＜0.01)。
3、ChIP结果显示:无论正常对照组还是Egr-1稳定转染组的Egr-1抗体免疫沉淀DNA中均可以扩增出含OPN启动子结合位点的基因调控区片段。
4、Western blot结果显示:正常对照组与OPN转染组给予PD98059后,与未处理组比较,P-ERK/ERK表达均明显减少(p＜0.01),同时,Egr-1表达明显下调(p＜0.01)。
结论
1、成功建立了Egr-1、OPN的稳定转染细胞株。
2、Egr-1无论在转录水平还是在翻译水平均可影响OPN的表达,同样,OPN也可以影响Egr-1的表达,二者的表达呈正相关性。
3、VSMC中,OPN启动子区域存在着Egr-1的结合位点,Egr-1参与了对OPN基因的转录调控。
4、OPN可以通过ERK途径反馈上调Egr-1的表达。
5、VSMC中,Egr-1与OPN之间形成一个作用级联放大的正反馈环。
Objective
Vascular remodeling diseases,such as atherosclerosis,restenosis(RS)following reconstructive vascular operation,and hypertension has become an increasingly serious threat to human health.Currently,many studies have demonstrated that migration and proliferation of the vascular smooth muscle cells(VSMCs)in the vascular tunica media is one pathological basis of vascular remodeling.
Studies have found that some transcription factors can regulate the expressions of several VSMC proliferation-associated genes,and thereby regulate VSMC proliferation. At the same time,under the influence of various external stimuli,some extracellular matrix can be used as extracellular signal reaches the nucleus and then through an intracellular signal transduction system to induces the expression of a series of VSMC proliferation-associated transcription factors,and thereby promotes VSMC migration and proliferation.With careful attention to the reciprocal relationship between transcription factors and extracellular matrix,which conduce to finding out the mechanism underlying vascular remodeling.
Early growth response factor-1(Egr-1),a zinc finger transcription factor,regulates the expression of multiple proliferation-associated genes,and its expression level is closely correlated with cell proliferation.Studies found that Egr-1 could mediate the migration and proliferation of VSMCs.Osteopontin(OPN)is a functionally important protein in the extracellular matrix(ECM),and anti-OPN antibodies can inhibit the phenotypic modulation,proliferation and migration of VSMCs.Although OPN and Egr-1 both play a significant role in mediating the process of migration and proliferation of the VSMCs,relatively little is known about the relationship between them.
We used Egr-1 and OPN cDNA to transfect VSMCs and then detected the expression changes of Egr-1 and OPN and observed the relationship between Egr-1 and OPN before and after transfection.Meanwhile,we performed a chromatin immunoprecipitation(CHIP)assay to examine whether Egr-1 could bind to the OPN promoter,and In order to clarify the mechanism of OPN-mediated changes in Egr-1 expression,extracellular signal-regulated kinase(ERK)inhibitor was added to OPN-transfected cells.These results of the interaction between Egr-1 and OPN will likely provide an important theoretical and experimental basis needed to control the inappropriate remodeling of vessel walls.
Materials and Methods
1.Rat A10 aortic VSMCs were cultured
Rat A10 aortic VSMCs were purchased from ATCC.Cells were cultured in Dulbecco's Modified Eagle Medium(DMEM)with 10%fetal bovine serum(FBS)at 37℃in a humid atmosphere of 5%CO_2.
2.Vectors and stable transfection
pCMV-Egr-1/NEO and pCMV(-)/NEO were kindly provided by Dr.Lorraine E. Chalifour,Division of Experimental Medicine,McGill University,Montreal,Canada. pET-28-rOPN/NEO and pET-28(-)/NEO were kindly provided by Dr.Harvey A Goldberg,University of Western Ontario,London,Canada.The plasmids were purified, amplified,identified and the ET-28-rOPN and ET-28(-)cassette were placed under control of a CMV promoter to drive OPN expression by TaKaRa.
To establish stable cell lines,cells were cultured at 80%confluence in 6-well dishes and then transfected with either pCMV-Egr-1/NEO or pCMV(-)/NEO or pCMV-ET-28-rOPN/NEO or pCMV-ET-28(-)/NEO,which both express Geneticin 418(G418)resistance.All transfections were performed using FuGENE6 according to the manufacturer's instructions.One day later,cells were subcultured and grown in the presence of 400μg/ml G418.After 2 weeks,single G418-resistant colonies were obtained by serial dilution in 96-well dishes.Colonies were maintained in a medium containing 200μg/ml of G418 and analyzed individually for expression of Egr-1 or OPN.
3.ERK inhibitor to inhibit the ERK pathway
Cells expressing OPN were cultured at 80%confluence and then treated with 10μMPD98059,the ERK pathway inhibitor.24h after PD98059 treatment,we analyzed Egr-1 expression via Western blot.
4.Reverse transcriptase polymerase chain reaction(RT-PCR)for Egr-1、OPNmRNA
Total RNA was extracted from VSMCs using Trizol Reagent according to the manufacturer's instructions.RT-PCR analysis was performed using TaKaRa RNA PCR Kit(AMV)Ver.3.0 according to the manufacturer's instructions.Products were resolved by 1%agarose gel and bands visualized by ethidium bromide staining. Densitometric analysis of bands was performed using BioImaging Systems.
5.Western blot analysis for Egr-1、OPN、ERK、P-ERK protein
Protein lysate(80μg)from cells was resolved by 12%sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE)and transferred to PVDF membranes. The filters were blocked with TBST buffer containing 5%skim milk,incubated with primary antibodies(anti-Egr-1(1:300),anti-OPN(1:500),anti-ERK(1:200), anti-P-ERK(1:200),andα-tubulin(1:500)overnight at 4℃Samples were then incubated with HRP-IgG secondary antibody and enhanced chemiluminescence(ECL) was used to visualize the bands.Band quantification was performed using Quantity One.Experiments were performed in triplicate.
6.Chromatin immunoprecipitation(ChIP)analysis to observe the Egr-1 binds to the OPN gene promoter.
ChIP assays were performed using the ChIP assay kit according to the manufacturer's instructions.The procedure included DNA-protein cross-linking inchromatin,shearing DNA into smaller fragments,immunoprecipitation with anti-Egr-1 antibody,and PCR identification of associated DNA sequences.Input was used as the positive control.
7.Statistical analyses
All values were expressed as mean±SD.SPSS11.0 software was used for all statistical analysis.The relationship between Egr-1 and OPN was analyzed using bivariate correlation analytical method(Spearman's correlation analysis).Data were analyzed using one-way analysis of variance followed by a least significant difference test(LSD)for multiple comparisons.Differences were considered significant If p＜0.05.
Results
1.RT-PCR results showed that Egr-1 mRNA was more highly expressed in VSMCs which were stably transfected with Egr-1 than in control cells and vector-transfected cells(p＜0.01),similarly,OPN mRNA was more highly expressed in cells stably transfected with OPN cDNA than controls group and vector-transfected group(p＜0.01).
2.We analyzed Egr-1 and OPN expression via RT-PCR and Western blot.The results showed that OPN mRNA and protein expression levels in the Egr-1-transfected VSMCs were significantly increased(p＜0.01),compared to untransfected cells and vector-transfected cells.Similarly,Egr-1 mRNA and protein levels were increased in the OPN-transfected VSMCs(p＜0.01).
3.ChIP results showed that the OPN promoter sequence can be amplified in both control and Egr-1-transfected cells.
4.Western blot results showed that levels of phospho-ERK were decreased relative to ERK in the PD98059 treated group compared to controls(p＜0.01). Furthermore,we found that Egr-1 expression was reduced in PD98059 treated cells compared to control-treated cells(p＜0.01).
Conclusion
1.Egr-1 cDNA or OPN cDNA was successfully transfected into VSMC_S.
2.Egr-1 can affect the expression of OPN,Similarly,OPN can also affect the expression of Egr-1,and Egr-1 and OPN expression were positively correlated.
3.Egr-1 can bind to the OPN promoter,and is likely to directly regulate its transcription.
4.OPN acts through the ERK pathway to upregulate Egr-1.
5.Egr-1 and OPN factors are likely to operate in a positive feedback loop in VSMC.
前言
动脉粥样硬化性疾病、血管重建术后再狭窄(restenosis,RS)、高血压等血管重塑相关性疾病已经愈来愈严重地威胁着人类的健康。目前认为,血管中膜平滑肌细胞(smooth muscle cell,SMC)向内膜下迁移与增殖是血管重塑的主要病理基础之一。
研究表明,某些转录因子(transcription factors,TF)可以调节多个血管平滑肌细胞(vascular smooth muscle cell,VSMC)增殖相关基因的表达,从不同层面调控VSMC的迁移与增殖。同时,人们也逐渐注意到细胞外基质(extracellular matrix,ECM)在血管重塑过程中的重要地位。研究发现,ECM在各种外界刺激的作用下,可以作为细胞外信号,经跨膜信号传递系统到达核内,激活一系列调控VSMC增殖的TF,从而推动VSMC的分裂、迁移及增殖。TF与ECM在血管重塑的过程中密不可分,因此将二者有机地联系在一起,并对其相互作用关系进行深入探讨,将有助于全面了解血管重塑的细胞与分子机制。
早期生长反应因子-1(eally growth response factor-1,Egr-1)作为一种锌指结构TF,参与多种基因的调控,与细胞增殖关系密切。大量研究表明,Egr-1在介导VSMC迁移、增殖过程中发挥着重要的作用,成为目前血管重塑机制研究中的焦点。骨桥蛋白(osteopontin,OPN)作为ECM中一种重要的功能性蛋白,被认为是血管损伤重塑过程中重要的始动因素,应用抗OPN抗体可以抑制VSMC的表型转化、迁移和增殖。尽管Egr-1和OPN在介导VSMC迁移、增殖过程中均起着重要的作用,但二者之间的关系尚不清楚。
本课题拟在以往研究的基础上,对培养的大鼠主动脉VSMCs分别稳定转染Egr-1、OPN基因,并检测转染前后OPN与Egr-1的表达变化,从而分析二者的相关性。并应用染色质免疫沉淀(chromatin immunoprecipitation,ChIP)方法检测Egr-1与OPN启动子的结合情况,同时通过向OPN稳定转染细胞株中加入细胞外信号调节蛋白激酶(extracellular signal-regulated kinase,ERK)的磷酸化抑制剂,检测抑制ERK磷酸化后,OPN上调Egr-1的水平变化,从而对Egr-1与OPN相关性的内在机制进行深入探讨,并为抑制VSMC的迁移、增殖,控制血管壁的不适当重塑提供理论及实验基础。
材料与方法
1、大鼠VSMC的培养
大鼠A10主动脉VSMC细胞株购自ATCC细胞库,用含10%胎牛血清(fetalbovine serum,FBS)的DMEM培养基,在37℃、5%CO_2、饱和湿度的培养箱中培养,用0.25%的胰酶消化、传代。
2、质粒的提纯、扩增及稳定转染
委托TaKaRa公司对pCMV-Egr-1/NEO、pCMV(-)/NEO质粒及pET-28-rOPN/NEO、pET-28(-)/NEO质粒进行提纯、扩增及鉴定,其中pET-28-rOPN/NEO和pET-28(-)/NEO质粒被连于CMV启动子上,以便在VSMC中表达。
待培养的VSMCs生长到80%汇合度时,以100μg/ml～1mg/ml的新霉素(neomycin418,G418)浓度进行最低G418浓度筛选。当培养的VSMCs在6孔板内达到80%汇合度时,应用FuGENE6分别向VSMCs内转染pCMV-Egr-1/NEO、pCMV(-)/NEO质粒及pCMV-ET-28-rOPN/NEO、pCMV-ET-28(-)/NEO质粒。细胞转染24小时(hour,h)后,加入400μg/ml G418的培养液进行培养。2周后,采用有限稀释法将细胞传至96孔板中进行单克隆化培养(于含200μg/ml G418的条件培养液中),待长满后进行扩大培养,并检测细胞中Egr-1、OPNmRNA的表达。
3、应用ERK磷酸化抑制剂阻断ERK信号传导通路
当OPN稳定转染的VSMCs生长至80%汇合度时,向细胞培养液中加入10μMERK磷酸化抑制剂PD98059,继续培养24h后,进行Western blot检测。
4、反转录多聚酶链反应(reverse transcriptase polymerase chainreaction,RT-PCR)检测Egr-1、OPNmRNA
用Trizol Reagent提取VSMCs的总RNA。用RNA PCR Kit Ver.3.0扩增目的片段。PCR产物电泳后,用溴化乙啶(ethidium bromide,EB)染色,BioImaging Systems系统成像,用NIH image软件分析产物条带灰度,以Egr-1、OPN产物条带灰度值与甘油醛-3-磷酸脱氢酶(glyceralde-3-phosphate dehydrogena,GAPDH)产物条带灰度值的比值分别作为Egr-1、OPNmRNA的相对表达量。
5、Western blot检测Egr-1、OPN、ERK、P-ERK蛋白
提取细胞总蛋白,上样量为80μg。转印到PVDF膜上后用5%脱脂奶粉封闭,用Egr-1兔抗大鼠多克隆抗体(1:300)、OPN兔抗大鼠多克隆抗体(1:500)、ERK兔抗大鼠多克隆抗体(1:200)、磷酸化细胞外调节蛋白激酶(phosphoextracellularsignal-regulated kinase,P-ERK)兔抗大鼠多克隆抗体(1:200)及兔抗大鼠α-tubulin多克隆抗体(1:500)4℃孵育过夜,分别用相应的二抗37℃孵育2h,ECL发光1分钟,胶片中的蛋白条带经BioImaging Systems采集后进行灰度值测定。目的蛋白灰度值与内对照α-tubulin比值为其相对蛋白定量。
6、ChIP方法检测Egr-1与OPN启动子的结合
分别取2×10~6个对数生长期的稳定转染Egr-1的VSMCs及正常VSMCs,用甲醛交联,0.125M甘氨酸终止交联。超声剪切染色质脱氧核糖核酸(deoxyribonucleicacid,DNA)成500～1000个碱基对(basepair,bp)片段。分别取50μl全细胞抽提物作为实验和阴性对照样本。用ChIP缓冲液10倍稀释后各取其中10μl合并作为输入对照(input),在实验样本中加入5g Egr-1兔抗大鼠多克隆抗体(1:300),将实验样品及阴性对照样本4℃孵育过夜。几次洗涤后,复合物从珠子上洗脱下来。用酚氯仿抽提纯化DNA,最后应用于PCR分析。
7、统计学分析
采用SPSS11.0统计软件进行数据分析。Egr-1与OPN的相关性用双变量相关分析法(Spearman相关分析),不同组间的参数比较用One-way ANOVA分析,两两比较采用LSD方法,P＜0.05有统计学意义。
实验结果
1、RT-PCR结果显示:Egr-1稳定转染组的Egr-1mRNA较正常对照组和空载体组表达明显增强(p＜0.01);OPN稳定转染组OPNmRNA与正常对照组和空载体组比较,表达也明显增强(p＜0.01)。
2、RT-PCR及Western blot方法检测Egr-1和OPN稳定转染VSMCs后OPN和Egr-1的表达水平。结果显示:与正常对照组和空载体组比较,Egr-1转染组的OPNmRNA及蛋白表达均随着Egr-1表达的增强而明显上调(p＜0.01);同样,OPN稳定转染组的Egr-1mRNA及蛋白表达较正常对照组和空载体组明显升高(p＜0.01)。
3、ChIP结果显示:无论正常对照组还是Egr-1稳定转染组的Egr-1抗体免疫沉淀DNA中均可以扩增出含OPN启动子结合位点的基因调控区片段。
4、Western blot结果显示:正常对照组与OPN转染组给予PD98059后,与未处理组比较,P-ERK/ERK表达均明显减少(p＜0.01),同时,Egr-1表达明显下调(p＜0.01)。
结论
1、成功建立了Egr-1、OPN的稳定转染细胞株。
2、Egr-1无论在转录水平还是在翻译水平均可影响OPN的表达,同样,OPN也可以影响Egr-1的表达,二者的表达呈正相关性。
3、VSMC中,OPN启动子区域存在着Egr-1的结合位点,Egr-1参与了对OPN基因的转录调控。
4、OPN可以通过ERK途径反馈上调Egr-1的表达。
5、VSMC中,Egr-1与OPN之间形成一个作用级联放大的正反馈环。
Objective
Vascular remodeling diseases,such as atherosclerosis,restenosis(RS)following reconstructive vascular operation,and hypertension has become an increasingly serious threat to human health.Currently,many studies have demonstrated that migration and proliferation of the vascular smooth muscle cells(VSMCs)in the vascular tunica media is one pathological basis of vascular remodeling.
Studies have found that some transcription factors can regulate the expressions of several VSMC proliferation-associated genes,and thereby regulate VSMC proliferation. At the same time,under the influence of various external stimuli,some extracellular matrix can be used as extracellular signal reaches the nucleus and then through an intracellular signal transduction system to induces the expression of a series of VSMC proliferation-associated transcription factors,and thereby promotes VSMC migration and proliferation.With careful attention to the reciprocal relationship between transcription factors and extracellular matrix,which conduce to finding out the mechanism underlying vascular remodeling.
Early growth response factor-1(Egr-1),a zinc finger transcription factor,regulates the expression of multiple proliferation-associated genes,and its expression level is closely correlated with cell proliferation.Studies found that Egr-1 could mediate the migration and proliferation of VSMCs.Osteopontin(OPN)is a functionally important protein in the extracellular matrix(ECM),and anti-OPN antibodies can inhibit the phenotypic modulation,proliferation and migration of VSMCs.Although OPN and Egr-1 both play a significant role in mediating the process of migration and proliferation of the VSMCs,relatively little is known about the relationship between them.
We used Egr-1 and OPN cDNA to transfect VSMCs and then detected the expression changes of Egr-1 and OPN and observed the relationship between Egr-1 and OPN before and after transfection.Meanwhile,we performed a chromatin immunoprecipitation(CHIP)assay to examine whether Egr-1 could bind to the OPN promoter,and In order to clarify the mechanism of OPN-mediated changes in Egr-1 expression,extracellular signal-regulated kinase(ERK)inhibitor was added to OPN-transfected cells.These results of the interaction between Egr-1 and OPN will likely provide an important theoretical and experimental basis needed to control the inappropriate remodeling of vessel walls.
Materials and Methods
1.Rat A10 aortic VSMCs were cultured
Rat A10 aortic VSMCs were purchased from ATCC.Cells were cultured in Dulbecco's Modified Eagle Medium(DMEM)with 10%fetal bovine serum(FBS)at 37℃in a humid atmosphere of 5%CO_2.
2.Vectors and stable transfection
pCMV-Egr-1/NEO and pCMV(-)/NEO were kindly provided by Dr.Lorraine E. Chalifour,Division of Experimental Medicine,McGill University,Montreal,Canada. pET-28-rOPN/NEO and pET-28(-)/NEO were kindly provided by Dr.Harvey A Goldberg,University of Western Ontario,London,Canada.The plasmids were purified, amplified,identified and the ET-28-rOPN and ET-28(-)cassette were placed under control of a CMV promoter to drive OPN expression by TaKaRa.
To establish stable cell lines,cells were cultured at 80%confluence in 6-well dishes and then transfected with either pCMV-Egr-1/NEO or pCMV(-)/NEO or pCMV-ET-28-rOPN/NEO or pCMV-ET-28(-)/NEO,which both express Geneticin 418(G418)resistance.All transfections were performed using FuGENE6 according to the manufacturer's instructions.One day later,cells were subcultured and grown in the presence of 400μg/ml G418.After 2 weeks,single G418-resistant colonies were obtained by serial dilution in 96-well dishes.Colonies were maintained in a medium containing 200μg/ml of G418 and analyzed individually for expression of Egr-1 or OPN.
3.ERK inhibitor to inhibit the ERK pathway
Cells expressing OPN were cultured at 80%confluence and then treated with 10μMPD98059,the ERK pathway inhibitor.24h after PD98059 treatment,we analyzed Egr-1 expression via Western blot.
4.Reverse transcriptase polymerase chain reaction(RT-PCR)for Egr-1、OPNmRNA
Total RNA was extracted from VSMCs using Trizol Reagent according to the manufacturer's instructions.RT-PCR analysis was performed using TaKaRa RNA PCR Kit(AMV)Ver.3.0 according to the manufacturer's instructions.Products were resolved by 1%agarose gel and bands visualized by ethidium bromide staining. Densitometric analysis of bands was performed using BioImaging Systems.
5.Western blot analysis for Egr-1、OPN、ERK、P-ERK protein
Protein lysate(80μg)from cells was resolved by 12%sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE)and transferred to PVDF membranes. The filters were blocked with TBST buffer containing 5%skim milk,incubated with primary antibodies(anti-Egr-1(1:300),anti-OPN(1:500),anti-ERK(1:200), anti-P-ERK(1:200),andα-tubulin(1:500)overnight at 4℃Samples were then incubated with HRP-IgG secondary antibody and enhanced chemiluminescence(ECL) was used to visualize the bands.Band quantification was performed using Quantity One.Experiments were performed in triplicate.
6.Chromatin immunoprecipitation(ChIP)analysis to observe the Egr-1 binds to the OPN gene promoter.
ChIP assays were performed using the ChIP assay kit according to the manufacturer's instructions.The procedure included DNA-protein cross-linking inchromatin,shearing DNA into smaller fragments,immunoprecipitation with anti-Egr-1 antibody,and PCR identification of associated DNA sequences.Input was used as the positive control.
7.Statistical analyses
All values were expressed as mean±SD.SPSS11.0 software was used for all statistical analysis.The relationship between Egr-1 and OPN was analyzed using bivariate correlation analytical method(Spearman's correlation analysis).Data were analyzed using one-way analysis of variance followed by a least significant difference test(LSD)for multiple comparisons.Differences were considered significant If p＜0.05.
Results
1.RT-PCR results showed that Egr-1 mRNA was more highly expressed in VSMCs which were stably transfected with Egr-1 than in control cells and vector-transfected cells(p＜0.01),similarly,OPN mRNA was more highly expressed in cells stably transfected with OPN cDNA than controls group and vector-transfected group(p＜0.01).
2.We analyzed Egr-1 and OPN expression via RT-PCR and Western blot.The results showed that OPN mRNA and protein expression levels in the Egr-1-transfected VSMCs were significantly increased(p＜0.01),compared to untransfected cells and vector-transfected cells.Similarly,Egr-1 mRNA and protein levels were increased in the OPN-transfected VSMCs(p＜0.01).
3.ChIP results showed that the OPN promoter sequence can be amplified in both control and Egr-1-transfected cells.
4.Western blot results showed that levels of phospho-ERK were decreased relative to ERK in the PD98059 treated group compared to controls(p＜0.01). Furthermore,we found that Egr-1 expression was reduced in PD98059 treated cells compared to control-treated cells(p＜0.01).
Conclusion
1.Egr-1 cDNA or OPN cDNA was successfully transfected into VSMC_S.
2.Egr-1 can affect the expression of OPN,Similarly,OPN can also affect the expression of Egr-1,and Egr-1 and OPN expression were positively correlated.
3.Egr-1 can bind to the OPN promoter,and is likely to directly regulate its transcription.
4.OPN acts through the ERK pathway to upregulate Egr-1.
5.Egr-1 and OPN factors are likely to operate in a positive feedback loop in VSMC.
引文
1 Stula M,Orzechowski HD,Gschwend S,et al.Influence of sustained mechanical stress on Egr-1 mRNA expression in cultured human endothelial cells.Mol Cell Biochem.2000;210:101-108.
2 Thiel G,Cibelli G Regulation of life and death by the zinc finger transcription factor Egr-l.J Cell Phyiol.2002;193:287-292.
3 Khachigian LM.Early growth response-1 in cardiovascular pathobiology.Circ Res.2006;98:186-191.
4 Takahashi Y,Fujioka Y,Takahashi T,et a/.Chylomicron remnants regulate early growth response factor-1 in vascular smooth muscle cells.Life Sci.2005;77:670-682.
5 Fahmy RG,Khachigian LM.et al.Suppression of growth factor expression and human vascular smooth muscle cell growth by small interfering RNA targeting EGR-l.J Cell Biochem.2007;100:1526-1535.
6 Zhang YM,Shi GG,Tang Z,et al.Effects of N-n-butyl haloperidol iodide on myocardial ischemia/reperfusion injury and Egr-1 expression in rat.Acta Biochim Biophys Sin (Shanghai).2006;38:435-441.
7 Fahmy RG,Khachigian LM.Suppression of growth factor expression and human vascular smooth muscle cell growth by small interfering RNA targeting EGR-l.J Cell Biochem.2007;100:1526-1535.
8 Liu GN,Teng YX,Yan W.Transfected synthetic DNA Enzyme Gene specifically inhibits Egr-1 gene expression and reduces Neointimal Hyperplasia following balloon injury in rats.Int J Cardiol.2008;129:118-124.
9 Yunfeng Lin,Xizhe Chen,Zhengbin Yan,et al.Multilineage differentiation of adipose-derived stromal cells from GFP transgenic mice.Mol Cell Biochem.2006;285:69-78.
10 Missihoun C,Zisa D,Shabbir A,et al.Myocardial oxidative stress,osteogenic phenotype,and energy metabolism are differentially involved in the initiation and early progression of delta-sarcoglycan-null cardiomyopathy.Mol Cell Biochem.2009;321:45-52.
11 Wu L,Wu Y,Lin Y,et al.Osteogenic differentiation of adipose derived stem cells promoted by overexpression of osterix.Mol Cell Biochem.2007;301:83-92.
12 Ashkar S,Weber GF,Panoutsakopoulou V,et al.Eta-1 (osteopontin):an early component of type-1 (cell-mediated)immunity.Science.2000;287:860-864.
13 Denhardt DT,Guo X.Osteopontin:a protein with diverse functions.FASEB J.1993;7:1475-1482.
14 Mohler ER 3rd,Adam LP,McClelland P,et al.Detection of osteopontin in calcified human aortic valves.Arterioscler Thromb Vase Biol.1997;17:547-552.
15 Coskun S,Atalar E,Ozturk E,et aLPlasma osteopontin levels are elevated in non-ST-segment elevation acute coronary syndromesJ Natl Med Assoc.2006;98:1746-1750.
16 Singh M,Ananthula S,Milhorn DM,et al.Osteopontin:a novel inflammatory mediator of cardiovascular disease.Front Biosci.2007;12:214-221.
17 Soejima H,Irie A,Fukunaga T,et al.Osteopontin expression of circulating T cells and plasma osteopontin levels are increased in relation to severity of heart failure.Circ J.2007;71:1879-1884.
18 Georgiadou P,Iliodromitis EK,Kolokathis F,et al.Plasma levels of osteopontin before and 24 h after percutaneous coronary intervention.Expert Opin Ther Targets.2008;12:1477-1480.
19 Kato R,Momiyama Y,Ohmori R,et al.Prognostic significance of plasma osteopontin levels in patients undergoing percutaneous coronary intervention.Circ J.2009;73:152-157.
20 Liaw L,Lombardi DM,Almeida MM,et al.Neutralizing antibodies directed against osteopontin inhibit rat carotid neointimal thickening after endothelial denudation.Arterioscler Thromb Vase Biol.1997;17:188-193.
21 Yasumoto H,Kim S,Zhan Y,et al.Dominant negative c-jun gene transfer inhibits vascular smooth muscle cell proliferation and neointimal hyperplasia in rats.Gene Ther.2001;8:1682-1689.
22 Kume M,Komori K,Matsumoto T,et al.Administration of a decoy against the activator protein-1 binding site suppresses neointimal thickening in rabbit balloon-injured arteries.Circulation.2002;105:1226-1232.
23 Hlawaty H,San Juan A,Jacob MP,et al.Inhibition of MMP-2 gene expression with small interfering RNA in rabbit vascular smooth muscle cells.Am J Physiol Heart Circ Physiol.2007;293:3593-3601.
24 Zaltsman AB,Newby AC.Increased secretion of gelatinases A and B from the aortas of cholesterol fed rabbits:relationship to lesion severity.Atherosclerosis.1997;130:61-70.
25 Ohtani K,Egashira K,Usui M,et al.Inhibition of neointimal hyperplasia after balloon injury by element‘decoy'of early growth response gene-1 in hypercholesterolemic rabbits.Gene Ther.2004;11:126-132.
26 Wada Y,Suzuki J,Kawauchi M,et al.Early growth-response factor 1 and basic transcriptional element-binding protein 2 expression in cardiac allografts.J Heart Lung Transplant.2001;20:590-594.
27 Lowe HC,Fahmy RG,Kavurma MM,et al.Catalytic oligodeoxynucleotides define a key regulatory role for early growth response factor-1 in the porcine model of coronary instent restenosis.Circ Res.2001;89:670-677.
28 Rupp J,Hellwig-Burgel T,Wobbe V,et al.Chlamydia pneumoniae infection promotes a proliferative phenotype inthe vasculature through Egr-1 activation in vitro and in vivo.Proc Natl Acad Sci U S A.2005;102:3447-3452.
29 滕雅轩,赵卫华,刘闺男.针对 Egr-1mRNA 的 10-23 脱氧核酶抑制大鼠动脉损伤后内膜增生的研究.中国现代医学杂志.2006;16:1823-1827.
30 赵卫华,滕雅轩,刘闺男.早期生长反应因子-1 的特异脱氧核酶对球囊损伤大鼠颈动脉后转化生长因子-β1 表达的影响.中国组织化学与细胞化学杂志.2006;15:366-371.
31 滕雅轩,刘闺男,周敬.生长反应因子-1 特异的脱氧核酶与大鼠颈动脉损伤后诱生型一氧化氮合酶表达的关系.中国临床康复杂志.2005;9:52-55.
32 滕雅轩,刘闺男,周敬.早期生长反应因子-1 特异的脱氧核酶对大鼠动脉损伤后血管内皮功能的保护及超微结构的影响.中国动脉硬化杂志.2005;13:709-713.
33 周敬,刘闺男,滕雅轩.Egr-1特异的脱氧核酶对大鼠颈总动脉损伤后内膜增生的影响.中国组织化学与细胞化学杂志.2005;14:257-261.
34 Religa P,Bojakowski K,Gaciong Z,et al.Arteriosclerosis in rat aortic allografts:dynamics of cell growth,apoptosis and expression of extracellular matrix proteins.Mol Cell Biochem.2003;249:75-83.
35 Yunfeng Lin,Xizhe Chen,Zhengbin Yan,et al.Multilineage differentiation of adipose-derived stromal cells from GFP transgenic mice.Mol Cell Biochem.2006;285:69-78.
36 Missihoun C,Zisa D,Shabbir A,et al.Myocardial oxidative stress,osteogenic phenotype,and energy metabolism are differentially involved in the initiation and early progression of delta-sarcoglycan-null cardiomyopathy.Mol Cell Biochem.2009;321:45-52.
37 Wu L,Wu Y,Lin Y,et al.Osteogenic differentiation of adipose derived stem cells promoted by overexpression of osterix.Mol Cell Biochem.2007;301:83-92.
38 Ashkar S,Weber GF,Panoutsakopoulou V,et al.Eta-1 (osteopontin):an early component of type-1 (cell-mediated)immunity.Science.2000;287:860-864.
39 Denhardt DT,Guo X.Osteopontin:a protein with diverse functions.FASEB J.1993;7:1475-1482.
40 Mohler ER,Adam LP,Mcclell P,et al.Detection of osteopontin in calcified human aortic valves.Arterioscler Thromb Vase Biol.1997;17:547-552.
41 Liaw L,Lombardi DM,Almeida MM,et al.Neutralizing antibodies directed against osteopontin inhibit rat carotid neointimal thickening after endothelial denudation.Arterioscler Thromb Vase Biol.1997;17:188-193.
42 Okamoto H.Osteopontin and cardiovascular system.Mol Cell Biochem.2007;300:1-7.
43 Jugdutt BI,Palaniyappan A,Uwiera RR,et al.Role of healing-specific-matricellular proteins and matrix metalloproteinases in age-related enhanced early remodeling after reperfused STEMI in dogs.Mol Cell Biochem.2009.322;25-36.
44 Bostrom K.Osteopontin,a missing link in PDGF-induced smooth muscle cell migration.Cardiovasc Res.2007;75:634-635.
45 Jalvy S,Renault MA,Leen LL,et al.Autocrine expression of osteopontin contributes to PDGF-mediated arterial smooth muscle cell migration.Cardiovasc Res.2007;75:738-747.
46 Fu M,Zhu X,Zhang J,et al.Egr-1 target genes in human endothelial cells identified by microarray analysis.Gene.2003;315:33-41.
47 Goetze S,Kintscher U,Kaneshiro K,et al.TNFalpha induces expression of transcription factors c-fos,Egr-l,and Ets-1 in vascular lesions through extracellular signal-regulated kinases l/2.Atherosclerosis.2001;159:93-101.
48 Mechtcheriakova D,Schabbauer G,Lucerna M,et a/.Specificity,diversity,and convergence in VEGF and TNF-alpha signaling events leading to tissue factor up-regulation via EGR-1 in endothelial cells.FASEB J.2001;15:230-242.
49 Gousseva N,Kugathasan K,Chesterman CN,et al.Early growth response factor-1 mediates insulin-inducible vascular endothelial cell proliferation and regrowth after injury.J Cell Biochem.2001;81:523-534.
50 Das A,Chendil D,Dey S,et a/.Ionizing radiation down-regulates p53 protein in primary Egr-1-/-mouse embryonic fibroblast cells causing enhanced resistance to apoptosis.J Biol Chem.2001;276:3279-3286.
51 Guha M,O'Connell MA,Pawlinski R,et al.Lipopolysaccharide activation of the MEK-ERK pathway in human monocytic cells mediates tissue factor and tumor necrosis factor alpha expression by inducing Elk-1 phosphorylation and Egr-1 expression.Blood.2001;98:1429-1439.
52 Lo LW,Cheng JJ,Chiu JJ,et al.Endothelial exposure to hypoxia induces Egr-1 expression involving PKCalpha-mediated Ras/Raf-1/ERK pathway.J Cell Physiol.2001;188:304-312.
53 Day RM,Yang Y,Suzuki YJ,et al.Bleomycin upregulates gene expression of angiotensin--converting enzyme via mitogen-activated protein kinase and early growth response 1 transcription factor.Am J Respir Cell Mol Biol.2001;25:613-619.
54 De Sousa LP,Brasil BS,Silva BM,et al.Plasminogen/plasmin regulates c-fos and egr-1 expression via the MEK/ERK pathway.Biochem Biophys Res Commun.2005;329:237-245.
55 Burgeson RE,Nimni ME.Molecular structure and tissue distribution.Clin Orthop.1992;282:250-272.
56 Eugenia N,Baramovajohn D,et al.Degradation of extracellular matrix proteins by hemorrhagic metalloproteinase.Arch Biochem Biophys.1989;275;63-71.
57 Farb A,Kolodgie FD,Hwang JY,et al.Extracellular matrix changes in stented human coronary arteries.Circulation.2004;110:940-947.
58 Religa P,Bojakowski K,Gaciong Z,et al.Arteriosclerosis in rat aortic allografts:dynamics of cell growth,apoptosis and expression of extracellular matrix proteins.Mol Cell Biochem.2003;249:75-83.
59 Irrcher I,Hood DA.Regulation of Egr-1,SRF,and Sp1 mRNA expression in contracting skeletal muscle cells.J Appl Physiol.2004;97:2207-2213.
60 Fukada T,Tonks NK.The reciprocal role of Egr-1 and Sp family proteins in regulation of the PTP1B promoter in response to the p210 Bcr-Abl oncoprotein-tyrosine kinase.J Biol Chem.2001;276:25512-25519.
61 Scharnhorst V,Menke AL,Attema J,et al.EGR-1 enhances tumor growth and modulates the effect of the Wilms' tumor 1 gene products on tumorigenicity.Oncogene.2000;19:791-800.
62 Chapman NR,Perkins ND.Inhibition of the RelA(p65)NF-kappaB subunit by Egr-1.J Biol Chem.2000;275:4719-4725.
63 Khachigian LM.Catalytic oligonucleotides targeting EGR-1 as potential inhibitors of in-stent restenosis.Ann N Y Acad Sci.2001;947:412-415.
64 Santiago FS,Atkins DG,Khachigian LM.Vascular smooth muscle cell proliferation and regrowth after mechanical injury in vitro are Egr-1/NGFI-A-dependent.Am J Pathol.1999;155:897-905.
65 Zhao X,Johnson JN,Singh K,et al.Impairment of myocardial angiogenic response in the absence of osteopontin.Microcirculation.2007;14:233-240.
66 Yin FY,Sun JH,Han YY.Experimental study on expression of osteopontin in viral myocarditis.Zhonghua Er Ke Za Zhi.2008;46:710-711.
67 Qiu ZB,Wan S.Association between osteopontin and restenosis.Zhonghua Xin Xue Guan Bing Za Zhi.2008;36:470-472.
68 Kato R,Momiyama Y,Ohmori R,et al.High plasma levels of osteopontin in patients with restenosis after percutaneous coronary intervention.Arterioscler Thromb Vasc Biol.2006;26:1-2.
69 孙爱军,高平进,朱鼎良,等.骨桥蛋白在血管外膜中的表达及其对外膜成纤维细胞迁移的影响.中华心血管病杂志.2003;5:38-47.
70 孙爱军,高平进,刘建军,等骨桥蛋白增强自发性高血压大鼠血管外膜成纤维细胞的迁移活性．生理学报．2004：56：21-24．
71 Gonzalez P,Reguero JR,Lozano l,et al.A functional Spl/Egrl-tandem repeat polymorphism in the 5-lipoxygenase gene is not associated with myocardial infarction.Int J Immunogenet.2007;34:127-130.
72 Al-Sarraj A,Day RM,Thiel G.Specificity of transcriptional regulation by the zinc finger transcription factors Spl,Sp3,and Egr-1.J Cell Biochem.2005;94:153-167.
73 Irrcher I,Hood DA.Regulation of Egr-l,SRF,and Spl mRNA expression in contracting skeletal muscle cells.J Appl Physiol.2004;97:2207-2213.
74 Okamoto H.Osteopontin and cardiovascular system.Mol Cell Biochem.2007;300:1-7.
75 Scatena M,Liaw L,Giachelli CM.Osteopontin:a multifunctional molecule regulating chronic inflammation and vascular disease.Arterioscler Thromb Vase Biol.2007;27:2302-2309.
76 Lee JL,Wang MJ,Sudhir PR,et al.Osteopontin promotes integrin activation through outside-in and inside-out mechanisms:OPN-CD44V interaction enhances survival in gastrointestinal cancer cells.Cancer Res.2007;67:2089-2097.
77 Infanger M,Grosse J,Westphal K.Vascular endothelial growth factor induces extracellular matrix proteins and osteopontin in the umbilical artery.Ann Vase Surg.2008;22:273-284.
78 Abe K,Nakashima H,Ishida M,et al.Angiotensin Ⅱ-induced osteopontin expression in vascular smooth muscle cells involves Gq/11,Ras,ERK,Src and Ets-l.Hypertens Res.2008;31:987-998.
79 Jeon YM,Kook SH,Son YO Role of MAPK in mechanical force-induced up-regulation of type Ⅰ collagen and osteopontin in human gingival fibroblasts.Mol Cell Biochem.2009;320:45-52.
80 Zhu B,Suzuki K,Goldberg HA.Osteopontin modulates CD44-dependent chemotaxis of peritoneal macrophages through G-protein-coupled receptors:evidence of a role for an intracellular form of osteopontin.J Cell Physiol.2004;198:155-167.
81 Gordon JA,Hunter GK,Goldberg HA.Activation of the mitogen-activated protein kinase pathway by bone sialoprotein regulates osteoblast differentiation.Cells Tissues Organs.2009;189:138-143.
82 CollissonEA,De A,Suzuki H.Treatolent of metastatic melanoma with an orally available inhibitor of the Ras-Raf-MAPK caseade.CaneerRes.2003;63:5669-5673.
83 Lee JT Jr,Steelman LS,MeCubrey JA.Modulation of Raf/MEK/ERK kinase activity does not after the chemoresistance profile of advanced prostate cancer cells.Int J Oncol.2005;26:1637-1644.
84 Moon Y,Yang H,Kim YB.Up-regulation of early growth response gene 1 (EGR-1)via ERK signals attenuates sulindac sulfide-mediated cytotoxicity in the human intestinal epithelial cells.Toxicol Appl Pharmacol.2007;223:155-163.
85 Shin SY,Lee JH,Min B,et al.The translation inhibitor anisomycin induces Elk-1-mediated transcriptional activation of egr-1 through multiple mitogen-activated protein kinase pathways.Exp Mol Med.2006;38:677-685.
86 Guha M,0'Connell MA,Pawlinski R,et al.Lipopolysaccharide activation of the MEK-ERK pathway in human monocytic cells mediates tissue factor and tumor necrosis factor alpha expression by inducing Elk-1 phosphorylation and Egr-1 expression.Blood.2001;98:1429-1439.
1 Sukhatme VP,Cao XM,Chang LC,et al.A zinc finger-encoding gene coregulated with c-fos during growth and differentiation,and after cellular depolarization.Cell.1988;53:37-43.
2 朱慧,曹传华,谢丛华.Egr-1启动子辐射诱导的基因表达特性.武汉大学学报(医学版).2008;29:186-189.
3 Thiel G,Cibelli G.Regulation of life and death by the zinc finger transcription factor Egr-1.J Cell Physio.2002;193:287-292.
4 A1-Sarraj A,Day RM,Thiel G.Specificity of transcriptional regulation by the zinc finger transcription factors Sp1,Sp3,and Egr-1.J Cell Biochem.2005,94:153-167.
5 Gonz(?)lez P,Reguero JR,Lozano I,et al.A functional Sp1/Egr1-tandem repeat polymorphism in the 5-lipoxygenase gene is not associated with myocardial infarction.Int J Immunogenet.2007;34:127-130.
6 Kostadinova RM,Nawrocki AR,Frey FJ,et al.Tumor necrosis factor alpha and phorbol 12-myristate-13 acetate down-regulate human 11beta-hydroxysteroid dehydrogenase type 2through p50/p50 NF-kappaB homodimers and Egr-1.FASEB J.2005;19:650-652.
7 DUN,Pei XT,Sun JZ,et al.Regulatory Effect of Egr-1 Promoter Sequences Induced by Doxorubicin in Transcriptional Targeting on Expression of GM-CSF Gene.Zhongguo Shi Yan Xue Ye Xue Za Zhi.2008;16:1165-1169.
8 Kumbrink J,Gerlinger M,Johnson JP.Egr-1 induces the expression of its corepressor nab2 by activation of the nab2 promoter thereby establishing a negative feedback loop.J Biol Chem.2005,280:42785-42793.
9 庄楚香,吴名耀.Egr-1在小鼠和人体正常组织中的表达及其生理学功能.汕头大学医学院学报.2001;14:166-168.
10 吴名耀,庄楚香.Egr-1在小鼠和人组织中的表达及其与细胞增殖的关系.中国组织化学与细胞化学杂志.2002;11:72-74.
11 Ogata T.Egr-1 mRNA induction by medium flow involves mRNA stabilization and is enhanced by the p38 inhibitor SB203580 in osteoblast-like cells.Acta Physiol(Oxf).2008;194:177-188.
12 Irrcher I,Hood DA.Regulation of Egr-1,SRF,and Spl mRNA expression in contracting skeletal muscle cells.J Appl Physiol.2004;97:2207-2213.
13 Fukada T,Tonks NK.The reciprocal role of Egr-1 and Sp family proteins in regulation of the PTP1B promoter in response to the p210 Bcr-Abl oncoprotein-tyrosine kinase.J Biol Chem.2001;276:25512-25519.
14 Scharnhorst V,Menke AL,Attema J,et al.EGR-1 enhances tumor growth and modulates the effect of the Wilms' tumor 1 gene products on tumorigenicity.Oncogene.2000;19:791-800.
15 Chapman NR,Perkins ND.Inhibition of the RelA(p65)NF-kappaB subunit by Egr-1.J Biol Chem.2000;275:4719-4725.
16 Goetze S,Kintscher U,Kaneshiro K,et al.TNFalpha induces expression of transcription factors c-fos,Egr-l,and Ets-1 in vascular lesions through extracellular signal-regulated kinases l/2.Atherosclerosis.2001;159:93-101.
17 Mechtcheriakova D,Schabbauer G,Lucerna M,et al.Specificity,diversity,and convergence in VEGF and TNF-alpha signaling events leading to tissue factor up-regulation via EGR-1 in endothelial cells.FASEB J.2001;15:230-242.
18 Gousseva N,Kugathasan K,Chesterman CN,et al.Early growth response factor-1 mediates insulin-inducible vascular endothelial cell proliferation and regrowth after injury.J Cell Biochem.2001;81:523-534.
19 Das A,Chendil D,Dey S,et al.Ionizing radiation down-regulates p53 protein in primary Egr-1-/-mouse embryonic fibroblast cells causing enhanced resistance to apoptosis.J Biol Chem.2001;276:3279-3286.
20 Guha M,O'Connell MA,Pawlinski R,et al.Lipopolysaccharide activation of the MEK-ERK pathway in human monocytic cells mediates tissue factor and tumor necrosis factor alpha expression by inducing Elk-1 phosphorylation and Egr-1 expression.Blood.2001;98:1429-1439.
21 Lo LW,Cheng JJ,Chiu JJ,et al.Endothelial exposure to hypoxia induces Egr-1 expression involving PKCalpha-mediated Ras/Raf-1/ERK pathway.J Cell Physiol.2001;188:304-312.
22 Day RM,Yang Y,Suzuki YJ,et al.Bleomycin upregulates gene expression of angiotensin- -converting enzyme via mitogen-activated protein kinase and early growth response 1 transcription factor.Am J Respir Cell Mol Biol.2001;25:613-619.
23 De Sousa LP,Brasil BS,Silva BM,et al.Plasminogen/plasmin regulates c-fos and egr-1 expression via the MEK/ERK pathway.Biochem Biophys Res Commun.2005;329:237-245.
24 Moon Y,Yang H,Kim YB.Up-regulation of early growth response gene 1 (EGR-1)via ERK signals attenuates sulindac sulfide-mediated cytotoxicity in the human intestinal epithelial cells.Toxicol Appl Pharmacol.2007;223:155-163.
25 Shin SY,Lee JH,Min B,et al.The translation inhibitor anisomycin induces Elk-1-mediated transcriptional activation of egr-1 through multiple mitogen-activated protein kinase pathways.Exp Mol Med.2006;38:677-685.
26 Wu SQ,Minami T,Donovan DJ,et al.The proximal serum response element in the Egr-1 promoter mediates response to thrombin in primary human endothelial cells.Blood.2002;100:4454-4461.
27 Shin SY,Lee JH,Min B,et al.The translation inhibitor anisomycin induces Elk-1-mediated transcriptional activation of egr-1 through multiple mitogen-activated protein kinase pathways.Exp Mol Med.2006;38:677-685.
28 Grimes CA,Jope RS.Cholinergic stimulation of early growth response-1 DNA binding activity requires protein kinase C and mitogen-activated protein kinase kinase activation and is inhibited by sodium valproate in SH-SY5Y cells.J Neurochem.1999;73:1384-1392.
29 Yan SF,Lu J,Zou YS,et al.Protein kinase C-beta and oxygen deprivation.A novel Egr-1--dependent pathway for fibrin deposition in hypoxemic vasculature..! Biol Chem.2000;275:11921-11928.
30 Mechtcheriakova D,Wlachos A,Holzmuller U.,et al.Vascular endothelial cell growth factor-induced tissue factor expression in endothelial cells is mediated by EGR-1.Blood.1999;93:3811-3823.
31 Fang M,Wee SA,Ronski K,et al.Evidence of EGR1 as a differentially expressed gene among proliferative skin diseases.Genomic Med.2007;1:75-85.
32 Du B,Fu C,Kent KC,et al.Elevated Egr-1 in human atherosclerotic cells transcriptionally represses the transforming growth factor-beta type Ⅱ receptor.J Biol Chem.2000;275:39039-39047.
33 Braddock M.The transcription factor Egr-1:a potential drug in wound healing and tissue repair.Ann Med.2001;33:313-318.
34 Liu C,Yao J,Mercola D,et al.The transcription factor EGR-1 directly trans--activates the fibronectin gene and enhances attachment of human glioblastoma cell line U251.J Biol Chem.2000;275:20315-20323.
35 Bryant M,Drew GM,Houston P,et al.Tissue repair with a therapeutic transcription factor.Hum Gene Ther.2000;11:2143-2158.
36 Houston P,Campbell CJ,Svaren J,et al.The transcriptional corepressor NAB2 blocks Egr-1-mediated growth factor activation and angiogenesis.Biochem Biophys Res Commun.2001;283:480-486.
37 Nakamura H,Isaka Y,Tsujie M,et al.Introduction of DNA enzyme for Egr-1 into tubulointerstitial fibroblasts by electroporation reduced interstitial alpha-smooth muscle actin expression and fibrosis in unilateral ureteral obstruction(UUO)rats.Gene Ther.2002;9:495-502.
38 钟守军,姚运红,李飞虹,等.EGR-1 与p53,p16,Cyclin D1在鼻咽癌中的表达及意义.肿瘤研究与临床.2006;18:664-666.
39 Lee SW,Kim EJ,Um SJ.Transcriptional regulation of the p73 gene,a member of the p53 family,by early growth response-1(Egr-1).Biochem Biophys Res Commun.2008;373:455.
40 40.Cibelli G,Policastro V,R(o|¨)ssler OG,et al.Nitric oxide-induced programmed cell death in human neuroblastoma cells is accompanied by the synthesis of Egr-1,a zinc finger transcription factor.J Neurosci Res.2002;67:450-460.
41滕雅轩,刘闺男,周敬.早期反应生长因子1特异的脱氧核酶对大鼠动脉损伤后血管内皮功能的保护及超微结构的影.中国动脉硬化杂志.2005;13:709-713.
42 Eyries M,Agrapart M,Alonso A,et al.Phorbol ester induction of angiotensin-converting enzyme transcription is mediated by Egr-1 and AP-1 in human endothelial cells via ERK pathway.Circ Res.2002;91:899-906.
43 Yan SF,Mackman N,Kisiel W,et al.Hypoxia/Hypoxemia-Induced activation of the procoagulant pathways and the pathogenesis of ischemia-associated thrombosis.Biol.1999;19:2029-2035.
44 Ohtani K,Egashira K,Usui M,et al.Inhibition of neointimal hyperplasia after balloon injury by ciselement 'decoy' of early growth response gene-1 in hypercholesterolemic rabbits.Gene Ther.2004;l 1:126-132.
45 Wada Y,Suzuki J,Kawauchi M,et al.Early growth-response factor 1 and basic transcriptional element-binding protein 2 expression in cardiac allografts.J Heart Lung Transplant.2001;20:590-594.
46 Lowe HC,Fahmy RG,Kavurma MM,et al.Catalytic oligodeoxynucleotides define a key regulatory role for early growth response factor-1 in the porcine model of coronary in-stent restenosis.CircRes.2001;89:670-677.
47 Rupp J,Hellwig-Burgel T,Wobbe V,et al.Chlamydia pneumoniae infection promotes a proliferative phenotype in the vasculature through Egr-1 activation in vitro and in vivo.Proc Natl Acad Sci USA.2005;102:3447-3452.
48 Fahmy RG,Khachigian LM.et al.Suppression of growth factor expression and human vascular smooth muscle cell growth by small interfering RNA targeting EGR-l.J Cell Biochem.2007;100:1526-1535.
49 Zhang YM,Shi GG,Tang Z,et al.Effects of N-n-butyl haloperidol iodide on myocardial ischemia/reperfusion injury and Egr-1 expression in rat.Acta Biochim Biophys Sin (Shanghai).2006;38:435-441.
50 Fahmy RG,Khachigian LM.Suppression of growth factor expression and human vascular smooth muscle cell growth by small interfering RNA targeting EGR-l.J Cell Biochem.2007;100:1526-1535.
51 Khachigian LM.Catalytic oligonucleotides targeting EGR-1 as potential inhibitors of in-stent restenosis.Ann N Y Acad Sci.2001;947:412-415.
52 Santiago FS,Atkins DG,Khachigian LM.Vascular smooth muscle cell proliferation and regrowth after mechanical injury in vitro are Egr-1/NGFI-A-dependent.Am J Pathol.1999;155:897-905.
53 Nagai R,Suzuki T,Aizawa K,et al.Phenotypic modulation of vascular smooth muscle cells:dissection of transcriptional regulatory mechanisms.Ann N Y Acad Sci.2001;947:56-66.
54 Zhu X,Lin Y,Bacanamwo M,et al.Interleukin-1 beta-induced Id2 gene expression is mediated by Egr-1 in vascular smooth muscle cells.Cardiovasc Res.2007;76:141-148.
55 Liang KW,Ting CT,Yin SC,et al.Berberine suppresses MEK/ERK-dependent Egr-1 signaling pathway and inhibits vascular smooth muscle cell regrowth after in vitro mechanical injury.Biochem Pharmacol.2006;71:806-817.
56 Kamimura M,Bea F,Akizawa T,et al.Platelet-derived growth factor induces tissue factor expression in vascular smooth muscle cells via activation of Egr-1.Hypertension.2004;44:944-951.
57 滕雅轩,赵卫华,刘闺男,等.针对 Egr-1mRNA 的10-23脱氧核酶抑制大鼠动脉损伤后内膜增生的研究.中国现代医学杂志.2006;16:1823-1827.
58 吴岩,刘闺男,韩伟,等.Egr-1特异脱氧核酶抑制血管平滑肌细胞增殖的研究.山东医药.2008;48:53-55.
59 刘程伟,胡新华,张雪松,等.Egr-1 在自体移植静脉中的表达及意义.中国普外基础与临床杂志.2006;13:642-646.
60 Patrizio M,Musumeci M,Stati T,et al.Propranolol promotes Egr1 gene expression in cardiomyocytes via beta-adrenoceptors.Eur J Pharmacol.2008;587:85-89.
61 Bhindi R,Khachigian LM,Lowe HC.DNAzymes targeting the transcription factor Egr-1 reduce myocardial infarct size following ischemia-reperfusion in rats.J Thromb Haemost.2006;4:1476-1478.
62 Zhang Y,Shi G,Zheng J,et al.The protective effects ofNbutyl haloperidol iodide on myocardial ischemia-reperfusion injury in rats by inhibiting Egr-1 overexpre--ssion.Cell Physiol Biochem.2007;20:639-648.
63 Wang C,Dostanic S,Servant N,et al.Egr-1 negatively regulates expression of the sodium-calcium exchanger in cardiomyocytes in vitro and in vivo.Cardiovasc Res.2005;65:187-194.
64 Hara S,Arai M,Tomaru K,et al.Prostaglandin F2alpha inhibits SERCA2 gene transcription through an induction of Egr-1 in cultured neonatal rat cardiac myocytes.Int Heart J.2008;49:329-342.
65 初令,李翔,胡永斌,等.大鼠实验性矽肺早期生长反应基因表达及其作用的初步探讨.中华劳动卫生职业病杂志.2004;22:47-50.
66 Yan SF,Fujita T,Lu J,et al.Egr-1,a master switch coordinating upregulation of divergent gene families underlying ischemic stress.Nat Med.2000;6:1355-1361.
67 李海青,石刚刚,高分飞,等.钙拮抗剂对大鼠心肌缺血再灌注早期生长反应基因-1 表达的影响.汕头大学医学院学报.2008;21:7-10.
68 Arai M,Yoguchi A,Takizawa T,et al.Mechanism of doxorubicin-induced inhibition of sarcoplasmic reticulum Ca(2+)-ATPase gene transcription.Circ Res.2000;86:8-14.
69 Okada M,Fujita T,Sakaguchi T,et al.Extinguishing Egr-1-dependent inflammatory and thrombotic cascades after lung transplantation.F ASEB J.2001;15:2757-2759.
70 Okada M,Wang CY,Hwang DW,et al.Transcriptional control of cardiac allograft vasculopathy by early growth response gene-1(Egr-1).Circ Res.2002;91:135-142.
1 傅国香,高平进.骨桥蛋白与血管损伤的研究进展.中华高血压杂志,2006;14:603-605.
2 Helluin O,Chan C,Vilaire G,et al.The activation state of alphav beta 3 regulates platelet and lymphocyte adhesion to intactand thrombin-cleavaged osteopontin.J Biol Chem.2000;275:18337-18343.
3 林全,王良兴.骨桥蛋白与血管重塑的关系研究进展.医学综述.2004;10:731-733.
4 杨淑莉,韩梅,温进坤.骨桥蛋白与心血管疾病.中国动脉粥样硬化杂志.2000;8:371-373.
5 方欣,管思明.骨桥蛋白与高血压血管重构.心脏杂志.2007;19:232-233.
6 Gao C,Guo H,Downey L,et al.Osteopontin-dependent CD44v6 expression and cell adhesion in HepG2 cells.Carcinogenesis.2003;24:1871-1878.
7 Soejima H,Irie A,Fukunaga T,et al.Elevated plasma osteopontin levels were associated with osteopontin expression of CD4+ T cells in patients with unstable angina.Circ J.2006;70:851-856.
8 陈荣新,叶胜龙.骨桥蛋白与肿瘤转移的研究进展.国际外科学杂志.2006;33:16-18.
9 俞文华,张国新.骨桥蛋白与胶质溜研究进展.医学研究杂志.2006;35:91-93.
10 范哲,李鸿雁,龙毅等.骨桥蛋白的研究进展.中国实验诊断学.2005;9:307-309.
11 Pereira RO,Carvalho SN,Stumbo AC,et al.Osteopontin expression in coculture of differentiating rat fetal skeletal fibroblasts and myoblasts.In Vitro Cell Dev Biol Anim.2006;42:4-7.
12 Schoensiegel F,Bekeredjian R,Schrewe A,et al.Atrial natriuretic peptide and osteopontin are useful markers of cardiac disorders in mice.Comp Med.2007;57:546-553.
13 Yamate T,Kohri K,Umekawa T,et al.Interaction between osteopontin on madin darby canine kidney cell membrane and calcium oxalate crystal.Urol Int.1999;62:81-86.
14 Hickey FB,England K,Cotter TG.Bcr-Abl regulates osteopontin transcription via Ras,PI-3K,aPKC,Raf-l,and MEK.J Leukoc Biol.2005;78:289-300.
15 Li G,Oparil S,Kelpke SS,et al.Fibroblast growth factor receptor-1 signaling induces osteopontin expression and vascular smooth muscle cell-dependent adventitial fibroblast migration in vitro.Circulation.2002;106:854-859.
16 Renault MA,Jalvy S,Belloc I,et al.AP-1 is involved in UTP-induced osteopontin expression in arterial smooth muscle cells.Circ Res.2003;93:674-681.
17 周秀霞,温进坤,韩梅.白细胞介素-1β和肿瘤坏死因子-α对血管平滑肌细及基质金属蛋白酶-2骨桥蛋白基因表达的影响.中国动脉硬化杂志.1999;7:292.
18 傅国香,朱鼎良,刘建军等.骨桥蛋白参与血管紧张素Ⅱ诱导的血管平滑肌细胞迁移.高血压杂志.2006;14:277-280.
19 Jalvy S,Renault MA,Lam Shang Leen L,et al.CREB mediates UTP-directed arterial smooth muscle cell migration and expression of the chemotactic protein osteopontin via its interaction with activator protein-1 sites.Circ Res.2007;100:1292-1299.
20 Nakamura H,Honda H,Inada Y,et al.Osteopontin expression in vascular smooth muscle cells in patients with end-stage renal disease.Ther Apher Dial.2006;10:273-277.
21 Li G,Chen YF,Kelpke SS,et al.Estrogen attenuates integrin-beta(3)-dependent adventitial fibroblast migration after inhibition of osteopontin production in vascular smooth muscle cells.Circulation.2000;101:2949-2955.
22 Sugiyama T,Yoshimoto T,Hirono Y,et al.Aldosterone increases osteopontin gene expression in rat endothelial cells.Biochem Biophys Res Commun.2005;336:163-167.
23 Takemoto M,Yokote K,Yamazaki M,et al.Enhanced expression of osteopontin by high glucose in cultured rat aortic smooth muscle cells.Biochem Biophys Res Commun.1999;258:722-726.
24 Isoda K,Kamezawa Y,Ayaori M,et al.Osteopontin transgenic mice fed a high-cholesterol diet develop early fatty-streak lesions.Circulation.2003;107:679-681.
25 Zhu Y,Denhardt DT,Cao H,et al.Hypoxia upregulates osteopontin expression in NIH-3T3 cells via a Ras-activated enhancer.Oncogene.2005;24:6555-6563.
26 俞广进,熊其如.骨桥蛋白在肿瘤转移中的研究进展.临床肿瘤学杂志.2007;12:156-160.
27 Sodhi CP,Phadke SA,Batlle D,et al.Hypoxia stimulates osteopontin expression and proliferation of cultured vascular smooth muscle cells:potentiation by high glucose.Diabetes.2001;50:1482-1490.
28 Fujihara S,Yokozeki M,Oba Y,et al.Function and regulation of osteopontin in response to mechanical stress.J Bone Miner Res.2006;21:956-564.
29 Tanaka N,Momiyama Y,Ohmori R,et al.Effect of atorvastatin on plasma osteopontin levels in patients with hypercholesterolemia.Arterioscler Thromb Vase Biol.2006;26:129-130.
30 Luomala M,P(a|¨)iv(a|¨)H,Thelen K,et al.Osteopontin levels are associated with cholesterol synthesis markers in mildly hypercholesterolaemic patients.Acta Cardiol.2007;62:177-181.
31 Hu T,Li H,Zhang X,et al.Exposure of aortic vascular smooth muscle cells to low-frequency electromagnetic field inhibits osteopontin expression and matrix metalloproteinase activity.Int Heart J.2008;49:597-604.
32 Ashkar S,weber GF,Panoutsakopoulou V,et al.Eta-l(esteopentin):an early component of type-1(cell-mediated)immunity.Science.2000;287:860-864.
33 李秉慧,宋杏丽,周强,等.骨桥蛋白促创面愈合的实验研究.中华实验外科杂志.2004:21:1355-1357.
34 Khan SA,Cook AC,Kappil M,et al.Enhanced cell surface CD44 variant(v6,v9)expression by osteopontin in breast cancer epithelial cells facilitates tumor cell migration:novel post-transcriptional,post-translational regulation.Clin Exp Metastasis.2005;22:663-673.
35 Hikita ST,Vistica BP,Jones HR,et al.Osteopontin is proinflammatory in experimental autoimmune uveitis.Invest Ophthalmol Vis Sci.2006;47:4435-4443.
36 Ariztia E.Osteopontin contributes to hepatocyte growth factor induced tumor growth and metastasis formation.Exp Cell Res.2003;228:257-267.
37 Moye VE,Barraeloush R,West C,et al.Osteopontin expression correlates with adhesive and metastatic potential in metastasis indueing DNA-transfected rat mammary cell lines.Br J Canc.er.2004;90:1796-1802.
38 Irby RB,McCarthy SM,Yeatman TJ,et al.Osteopontin regulates multiple functions contributing to human colon cancer development and progression.Clin Exp Metastasis.2004;21:515-523.
39 Ganmann A,Petrow P,Mentzel T,et al.Osteopontin expression in primary sarcomas of the pulmonary artery.Virchows Arch.2001;439:668-674.
40 付汐,杨竹林,李代强,等.胰腺癌组织中 kail 和骨桥蛋白表达及其临床病理意义.检验与临床.2006;8:1690-1691.
41 张辉,应敏刚,任宁.肝细胞癌患者血桨骨桥蛋白表达水平与术后复发及愈后的关系.中国临 床医学．2007；14：1-6．
42 Pan HW,Ou YH,Peng SY,et al.Overexpression of osteopontin is associated with intrahepatic metastasis,early recurrence,and poorer prognosis of surgically reseeted hepatocellular carcinoma.Cancer.2003;98:119-127.
43 Nakamura H,Honda H,Inada Y,et al.Osteopontin expression in vascular smooth muscle cells in patients with end-stage renal disease.Ther Apher Dial.2006;10:273-277.
44 Nakae MJwamoto I,Fujino T,et al.Preoperative plasma osteopontin level as a biomarker complementary to carbohydrate antigen 125 in predicting ovarian cancer.J Obstet Gynaecol Res.2006;32:309-314.
45 Zhang YL,Zhou SX,Lei i,et ah Blockades of angiotensin and aldosterone reduce osteopontin expression and interstitial fibrosis infiltration in rats with myocardial infarction.Chin Med J (Engl).2008;121:2192-2196.
46 Coskun S,Atalar E,Ozturk E,et al.Plasma osteopontin levels are elevated in non-ST-segment elevation acute coronary syndromes.J Natl Med Assoc.2006;98:1746-1750.
47 Singh M,Ananthula S,Milhorn DM,et al.Osteopontin:a novel inflammatory mediator of cardiovascular disease.Front Biosci.2007;12:214-221.
48 Soejima H,Irie A,Fukunaga T,et al.Osteopontin expression of circulating T cells and plasma osteopontin levels are increased in relation to severity of heart failure.Circ J.2007;71:1879-1884.
49 Georgiadou P,Iliodromitis EK,Kolokathis F,et al.Plasma levels of osteopontin before and 24 h after percutaneous coronary intervention.Expert Opin Ther Targets.2008;12:1477-1480.
50 Kato R,Momiyama Y,Ohmori R,et al.Prognostic significance of plasma osteopontin levels in patients undergoing percutaneous coronary intervention.Circ J.2009;73:152-157.
51 Zhao X,Johnson JN,Singh K,et al.Impairment of myocardial angiogenic response in the absence of osteopontin.Microcirculation.2007;14:233-240.
52 Yin FY,Sun JH,Han YY.Experimental study on expression of osteopontin in viral myocarditis.Zhonghua Er Ke Za Zhi.2008;46:710-711.
53 Qiu ZB,Wan S.Association between osteopontin and restenosis.Zhonghua Xin Xue Guan Bing Za Zhi.2008;36:470-472.
54 Kato R,Momiyama Y,Ohmori R,et al.High plasma levels of osteopontin in patients with restenosis after percutaneous coronary intervention.Arterioscler Thromb Vasc Biol.2006;26:1-2.
55 孙爱军,高平进,朱鼎良,等.骨桥蛋白在血管外膜中的表达及其对外膜成纤维细胞迁移的影响.中华心血管病杂志.2003;5:38-47.
56 孙爱军,高平进,刘建军,等骨桥蛋白增强自发性高血压大鼠血管外膜成纤维细胞的迁移活性.生理学报.2004;56:21-24.
57 Bostr(o|¨)m K.Osteopontin,a missing link in PDGF-induced smooth muscle cell migration.Cardiovasc Res.2007;75:634-635.
58 Jalvy S,Renault MA,Leen LL,et al.Autocrine expression of osteopontin contributes to PDGF-mediated arterial smooth muscle cell migration.Cardiovasc Res.2007;75:738-747.
59 Roy S,Ruest PJ,Hanks SK.FAK regulates tyrosine phosphorylation of CAS,paxillin,and PYK2 in cells expressing v-Src,but is not a critical determinant of v-Src transformation.J Cell Biochem.2002;84:377-388.
60 Yin BL,Hao H,Wang YY,et al.Downregulating osteopontin reduces angiotensin Ⅱ-induced inflammatory activation in vascular smooth muscle cells.Inflamm Res.2009;58:67-73.
61 Mori S,Takemoto M,Yokote K,et al.Hyperglycemia-induced alteration of vascular smooth muscle phenotype.Diabetes Com placations.2002;16:65-68.
62.Jing-Jing Li,Mei Han,Jin-Kun Wen,et al.Osteopontin stimulates vascular smooth muscle cell migration by inducing FAK phosphorylation and ILK dephosphorylation.Biochemical and Biophysical Research Communications.2007;356:13-19.
63 李菁菁,温进坤,韩梅.Src 介导骨桥蛋白诱导的血管平滑肌细胞黏附和迁移过程.细胞生物学杂志.2006;28:757-762.
64 李菁菁,温进坤,韩梅.FAK 和 ILK 介导骨桥蛋白诱导的血管平滑肌细胞黏附和迁移.中国生物化学与分子生物学报.2006;2:477-483.
65 Li JJ,Han M,Wen JK,et al.Osteopontin stimulates vascular smooth muscle cell migration by inducing FAK phosphorylation and ILK dephosphorylation.Biochem Biophys Res Commun. 2007;356:13-19.
66 K han SA,Lopez-Chua CA,Zhang J,et al.Soluble osteopontin inhibits apoptosis of adherent endothelial cells deprived of growth factors.J Cell Biochem.2002;85:728-736.
67 Weintraub AS,Schnapp LM,Lin X,et al.Osteopontin deficiency in rat vascular smooth muscle cells is associated with an inability to adhere to collagen and increased apoptosis.Lab Invest.2000;80:1603-1615.
68 Abel B,Kurrer M,Shamshiev A,et al.The osteopontin-CD44 pathway is superfluous for the development of autoimmune myocarditis.Eur J Immunol.2006;36:494-499.
69 Lin YH,Yang-Yen HF.The osteopontin-CIM4 survival signal involves activation of the phosphatidylinositol 3-kinase/Akt signalingpathway.J Biol Chem.2001;276:46024-46030.
70 Malyankar UM,Seatena M,Suchland KL,et al.osteoprotegerln is an alpha v beta 3-induced.NF-kappa B-dependent survival factor for endothdial cells.J Biol Chem.2001;275:20959-20962.
71 Shin T,Ahn M,Kim K,et al.Increased expression of osteopontin in the heart tissue of Lewis rats with experimental autoimmune myocarditis.J Vet Med Sci.2006;68:379-382.
72 Abe K,Nakashima H,Ishida M,et al.Angiotensin Il-induced osteopontin expression in vascular smooth muscle cells involves Gq/11,Ras,ERK,Src and Ets-l.Hypertens Res.2008;31:987-998.
73 Kurata M,Okura T,Watanabe S,et al.Osteopontin and carotid atherosclerosis in patients with essential hypertension.Clin Sci (Lond).2006;l 11:319-324.
74 Li G,Oparil S,Kelpke SS,et al.Fibroblast growth factor receptor-1 signaling induces osteopontin expression and vascular smooth muscle cell-dependent adventitial fibroblast migration in vitro.Circulation.2002;106:854-859.
75 Roy S,Ruest PJ,Hanks SK.FAK regulates tyrosine phosphorylation of CAS,paxillin,and PYK2 in cells expressing v-Src,but is not a critical determinant of v-Src transformation.J Cell Biochem.2002;84:377-388.
76 Xie B,Zhao J,Kitagawa M,et al.Focal adhesion kinase activates Statlin integrin 2 mediated cell migration and adhesion.J Cel Biochem.2001;76:19512-119523.
77 Tuck AB,Hota C,Wilson SM,et al.Osteopontin-induced migration of human mammary epithelial cells involves activation of EGF receptor and multiple signal transduction pathways.Oncogene.2003;22:1198-1205.
78 Rangaswami H,Bulbule A,Kundu GC.Nuclear factor inducing kinase:a key regulator in osteopontin-induced MAPK/DcappaB kinase dependent NF-kappaB-mediated promatrix metalloproteinase-9 activation.Glycoconj J.2006;23:221-232.
79 Nerurkar SS,01zinski AR,Frazier KS,et al.P38 MAPK inhibitors suppress biomarkers of hypertension end-organ damage,osteopontin and plasminogen activator inhibitor-1.Biomarkers.2007;12:87-112.
80 Tuck AB,Hota C,Wilson SM,et al.Osteopontin-induced migration of human mammary epithelial cells involves activation of EGF receptor and multiple signal transduction pathways.Oncogene.2003;22:1198-1205.
81 Farb A,Kolodgie FD,Hwang JY,et al.Extracellular matrix changes in stented human coronary arteries.Circulation.2004;24,110:940-947.
82 Asa Strom,Ahnders Franzen.Altered vascular remodeling in osteopontin-deficient atherosclerotic mice.J Vase Res.2004;41:314-322.
83 Lai CF,Seshadri V,Hiiang K,et al.An osteopontin-NADPH oxidase signaling cascade promotes pro-matrix metalloproteinase 9 activation in aortic mesenchymal cells.Circ Res.2006;98:1479-1489.
84 Kikuo Isoda,Knichirou Nishikawa,Yashuhiro Kamezawa,et al.Osteopontin plays an important role in the development of medial thickening and neointimal formation.Circ Res.2002;91:77-82.
2 Thiel G,Cibelli G Regulation of life and death by the zinc finger transcription factor Egr-l.J Cell Phyiol.2002;193:287-292.
3 Khachigian LM.Early growth response-1 in cardiovascular pathobiology.Circ Res.2006;98:186-191.
4 Takahashi Y,Fujioka Y,Takahashi T,et a/.Chylomicron remnants regulate early growth response factor-1 in vascular smooth muscle cells.Life Sci.2005;77:670-682.
5 Fahmy RG,Khachigian LM.et al.Suppression of growth factor expression and human vascular smooth muscle cell growth by small interfering RNA targeting EGR-l.J Cell Biochem.2007;100:1526-1535.
6 Zhang YM,Shi GG,Tang Z,et al.Effects of N-n-butyl haloperidol iodide on myocardial ischemia/reperfusion injury and Egr-1 expression in rat.Acta Biochim Biophys Sin (Shanghai).2006;38:435-441.
7 Fahmy RG,Khachigian LM.Suppression of growth factor expression and human vascular smooth muscle cell growth by small interfering RNA targeting EGR-l.J Cell Biochem.2007;100:1526-1535.
8 Liu GN,Teng YX,Yan W.Transfected synthetic DNA Enzyme Gene specifically inhibits Egr-1 gene expression and reduces Neointimal Hyperplasia following balloon injury in rats.Int J Cardiol.2008;129:118-124.
9 Yunfeng Lin,Xizhe Chen,Zhengbin Yan,et al.Multilineage differentiation of adipose-derived stromal cells from GFP transgenic mice.Mol Cell Biochem.2006;285:69-78.
10 Missihoun C,Zisa D,Shabbir A,et al.Myocardial oxidative stress,osteogenic phenotype,and energy metabolism are differentially involved in the initiation and early progression of delta-sarcoglycan-null cardiomyopathy.Mol Cell Biochem.2009;321:45-52.
11 Wu L,Wu Y,Lin Y,et al.Osteogenic differentiation of adipose derived stem cells promoted by overexpression of osterix.Mol Cell Biochem.2007;301:83-92.
12 Ashkar S,Weber GF,Panoutsakopoulou V,et al.Eta-1 (osteopontin):an early component of type-1 (cell-mediated)immunity.Science.2000;287:860-864.
13 Denhardt DT,Guo X.Osteopontin:a protein with diverse functions.FASEB J.1993;7:1475-1482.
14 Mohler ER 3rd,Adam LP,McClelland P,et al.Detection of osteopontin in calcified human aortic valves.Arterioscler Thromb Vase Biol.1997;17:547-552.
15 Coskun S,Atalar E,Ozturk E,et aLPlasma osteopontin levels are elevated in non-ST-segment elevation acute coronary syndromesJ Natl Med Assoc.2006;98:1746-1750.
16 Singh M,Ananthula S,Milhorn DM,et al.Osteopontin:a novel inflammatory mediator of cardiovascular disease.Front Biosci.2007;12:214-221.
17 Soejima H,Irie A,Fukunaga T,et al.Osteopontin expression of circulating T cells and plasma osteopontin levels are increased in relation to severity of heart failure.Circ J.2007;71:1879-1884.
18 Georgiadou P,Iliodromitis EK,Kolokathis F,et al.Plasma levels of osteopontin before and 24 h after percutaneous coronary intervention.Expert Opin Ther Targets.2008;12:1477-1480.
19 Kato R,Momiyama Y,Ohmori R,et al.Prognostic significance of plasma osteopontin levels in patients undergoing percutaneous coronary intervention.Circ J.2009;73:152-157.
20 Liaw L,Lombardi DM,Almeida MM,et al.Neutralizing antibodies directed against osteopontin inhibit rat carotid neointimal thickening after endothelial denudation.Arterioscler Thromb Vase Biol.1997;17:188-193.
21 Yasumoto H,Kim S,Zhan Y,et al.Dominant negative c-jun gene transfer inhibits vascular smooth muscle cell proliferation and neointimal hyperplasia in rats.Gene Ther.2001;8:1682-1689.
22 Kume M,Komori K,Matsumoto T,et al.Administration of a decoy against the activator protein-1 binding site suppresses neointimal thickening in rabbit balloon-injured arteries.Circulation.2002;105:1226-1232.
23 Hlawaty H,San Juan A,Jacob MP,et al.Inhibition of MMP-2 gene expression with small interfering RNA in rabbit vascular smooth muscle cells.Am J Physiol Heart Circ Physiol.2007;293:3593-3601.
24 Zaltsman AB,Newby AC.Increased secretion of gelatinases A and B from the aortas of cholesterol fed rabbits:relationship to lesion severity.Atherosclerosis.1997;130:61-70.
25 Ohtani K,Egashira K,Usui M,et al.Inhibition of neointimal hyperplasia after balloon injury by element‘decoy'of early growth response gene-1 in hypercholesterolemic rabbits.Gene Ther.2004;11:126-132.
26 Wada Y,Suzuki J,Kawauchi M,et al.Early growth-response factor 1 and basic transcriptional element-binding protein 2 expression in cardiac allografts.J Heart Lung Transplant.2001;20:590-594.
27 Lowe HC,Fahmy RG,Kavurma MM,et al.Catalytic oligodeoxynucleotides define a key regulatory role for early growth response factor-1 in the porcine model of coronary instent restenosis.Circ Res.2001;89:670-677.
28 Rupp J,Hellwig-Burgel T,Wobbe V,et al.Chlamydia pneumoniae infection promotes a proliferative phenotype inthe vasculature through Egr-1 activation in vitro and in vivo.Proc Natl Acad Sci U S A.2005;102:3447-3452.
29 滕雅轩,赵卫华,刘闺男.针对 Egr-1mRNA 的 10-23 脱氧核酶抑制大鼠动脉损伤后内膜增生的研究.中国现代医学杂志.2006;16:1823-1827.
30 赵卫华,滕雅轩,刘闺男.早期生长反应因子-1 的特异脱氧核酶对球囊损伤大鼠颈动脉后转化生长因子-β1 表达的影响.中国组织化学与细胞化学杂志.2006;15:366-371.
31 滕雅轩,刘闺男,周敬.生长反应因子-1 特异的脱氧核酶与大鼠颈动脉损伤后诱生型一氧化氮合酶表达的关系.中国临床康复杂志.2005;9:52-55.
32 滕雅轩,刘闺男,周敬.早期生长反应因子-1 特异的脱氧核酶对大鼠动脉损伤后血管内皮功能的保护及超微结构的影响.中国动脉硬化杂志.2005;13:709-713.
33 周敬,刘闺男,滕雅轩.Egr-1特异的脱氧核酶对大鼠颈总动脉损伤后内膜增生的影响.中国组织化学与细胞化学杂志.2005;14:257-261.
34 Religa P,Bojakowski K,Gaciong Z,et al.Arteriosclerosis in rat aortic allografts:dynamics of cell growth,apoptosis and expression of extracellular matrix proteins.Mol Cell Biochem.2003;249:75-83.
35 Yunfeng Lin,Xizhe Chen,Zhengbin Yan,et al.Multilineage differentiation of adipose-derived stromal cells from GFP transgenic mice.Mol Cell Biochem.2006;285:69-78.
36 Missihoun C,Zisa D,Shabbir A,et al.Myocardial oxidative stress,osteogenic phenotype,and energy metabolism are differentially involved in the initiation and early progression of delta-sarcoglycan-null cardiomyopathy.Mol Cell Biochem.2009;321:45-52.
37 Wu L,Wu Y,Lin Y,et al.Osteogenic differentiation of adipose derived stem cells promoted by overexpression of osterix.Mol Cell Biochem.2007;301:83-92.
38 Ashkar S,Weber GF,Panoutsakopoulou V,et al.Eta-1 (osteopontin):an early component of type-1 (cell-mediated)immunity.Science.2000;287:860-864.
39 Denhardt DT,Guo X.Osteopontin:a protein with diverse functions.FASEB J.1993;7:1475-1482.
40 Mohler ER,Adam LP,Mcclell P,et al.Detection of osteopontin in calcified human aortic valves.Arterioscler Thromb Vase Biol.1997;17:547-552.
41 Liaw L,Lombardi DM,Almeida MM,et al.Neutralizing antibodies directed against osteopontin inhibit rat carotid neointimal thickening after endothelial denudation.Arterioscler Thromb Vase Biol.1997;17:188-193.
42 Okamoto H.Osteopontin and cardiovascular system.Mol Cell Biochem.2007;300:1-7.
43 Jugdutt BI,Palaniyappan A,Uwiera RR,et al.Role of healing-specific-matricellular proteins and matrix metalloproteinases in age-related enhanced early remodeling after reperfused STEMI in dogs.Mol Cell Biochem.2009.322;25-36.
44 Bostrom K.Osteopontin,a missing link in PDGF-induced smooth muscle cell migration.Cardiovasc Res.2007;75:634-635.
45 Jalvy S,Renault MA,Leen LL,et al.Autocrine expression of osteopontin contributes to PDGF-mediated arterial smooth muscle cell migration.Cardiovasc Res.2007;75:738-747.
46 Fu M,Zhu X,Zhang J,et al.Egr-1 target genes in human endothelial cells identified by microarray analysis.Gene.2003;315:33-41.
47 Goetze S,Kintscher U,Kaneshiro K,et al.TNFalpha induces expression of transcription factors c-fos,Egr-l,and Ets-1 in vascular lesions through extracellular signal-regulated kinases l/2.Atherosclerosis.2001;159:93-101.
48 Mechtcheriakova D,Schabbauer G,Lucerna M,et a/.Specificity,diversity,and convergence in VEGF and TNF-alpha signaling events leading to tissue factor up-regulation via EGR-1 in endothelial cells.FASEB J.2001;15:230-242.
49 Gousseva N,Kugathasan K,Chesterman CN,et al.Early growth response factor-1 mediates insulin-inducible vascular endothelial cell proliferation and regrowth after injury.J Cell Biochem.2001;81:523-534.
50 Das A,Chendil D,Dey S,et a/.Ionizing radiation down-regulates p53 protein in primary Egr-1-/-mouse embryonic fibroblast cells causing enhanced resistance to apoptosis.J Biol Chem.2001;276:3279-3286.
51 Guha M,O'Connell MA,Pawlinski R,et al.Lipopolysaccharide activation of the MEK-ERK pathway in human monocytic cells mediates tissue factor and tumor necrosis factor alpha expression by inducing Elk-1 phosphorylation and Egr-1 expression.Blood.2001;98:1429-1439.
52 Lo LW,Cheng JJ,Chiu JJ,et al.Endothelial exposure to hypoxia induces Egr-1 expression involving PKCalpha-mediated Ras/Raf-1/ERK pathway.J Cell Physiol.2001;188:304-312.
53 Day RM,Yang Y,Suzuki YJ,et al.Bleomycin upregulates gene expression of angiotensin--converting enzyme via mitogen-activated protein kinase and early growth response 1 transcription factor.Am J Respir Cell Mol Biol.2001;25:613-619.
54 De Sousa LP,Brasil BS,Silva BM,et al.Plasminogen/plasmin regulates c-fos and egr-1 expression via the MEK/ERK pathway.Biochem Biophys Res Commun.2005;329:237-245.
55 Burgeson RE,Nimni ME.Molecular structure and tissue distribution.Clin Orthop.1992;282:250-272.
56 Eugenia N,Baramovajohn D,et al.Degradation of extracellular matrix proteins by hemorrhagic metalloproteinase.Arch Biochem Biophys.1989;275;63-71.
57 Farb A,Kolodgie FD,Hwang JY,et al.Extracellular matrix changes in stented human coronary arteries.Circulation.2004;110:940-947.
58 Religa P,Bojakowski K,Gaciong Z,et al.Arteriosclerosis in rat aortic allografts:dynamics of cell growth,apoptosis and expression of extracellular matrix proteins.Mol Cell Biochem.2003;249:75-83.
59 Irrcher I,Hood DA.Regulation of Egr-1,SRF,and Sp1 mRNA expression in contracting skeletal muscle cells.J Appl Physiol.2004;97:2207-2213.
60 Fukada T,Tonks NK.The reciprocal role of Egr-1 and Sp family proteins in regulation of the PTP1B promoter in response to the p210 Bcr-Abl oncoprotein-tyrosine kinase.J Biol Chem.2001;276:25512-25519.
61 Scharnhorst V,Menke AL,Attema J,et al.EGR-1 enhances tumor growth and modulates the effect of the Wilms' tumor 1 gene products on tumorigenicity.Oncogene.2000;19:791-800.
62 Chapman NR,Perkins ND.Inhibition of the RelA(p65)NF-kappaB subunit by Egr-1.J Biol Chem.2000;275:4719-4725.
63 Khachigian LM.Catalytic oligonucleotides targeting EGR-1 as potential inhibitors of in-stent restenosis.Ann N Y Acad Sci.2001;947:412-415.
64 Santiago FS,Atkins DG,Khachigian LM.Vascular smooth muscle cell proliferation and regrowth after mechanical injury in vitro are Egr-1/NGFI-A-dependent.Am J Pathol.1999;155:897-905.
65 Zhao X,Johnson JN,Singh K,et al.Impairment of myocardial angiogenic response in the absence of osteopontin.Microcirculation.2007;14:233-240.
66 Yin FY,Sun JH,Han YY.Experimental study on expression of osteopontin in viral myocarditis.Zhonghua Er Ke Za Zhi.2008;46:710-711.
67 Qiu ZB,Wan S.Association between osteopontin and restenosis.Zhonghua Xin Xue Guan Bing Za Zhi.2008;36:470-472.
68 Kato R,Momiyama Y,Ohmori R,et al.High plasma levels of osteopontin in patients with restenosis after percutaneous coronary intervention.Arterioscler Thromb Vasc Biol.2006;26:1-2.
69 孙爱军,高平进,朱鼎良,等.骨桥蛋白在血管外膜中的表达及其对外膜成纤维细胞迁移的影响.中华心血管病杂志.2003;5:38-47.
70 孙爱军,高平进,刘建军,等骨桥蛋白增强自发性高血压大鼠血管外膜成纤维细胞的迁移活性．生理学报．2004：56：21-24．
71 Gonzalez P,Reguero JR,Lozano l,et al.A functional Spl/Egrl-tandem repeat polymorphism in the 5-lipoxygenase gene is not associated with myocardial infarction.Int J Immunogenet.2007;34:127-130.
72 Al-Sarraj A,Day RM,Thiel G.Specificity of transcriptional regulation by the zinc finger transcription factors Spl,Sp3,and Egr-1.J Cell Biochem.2005;94:153-167.
73 Irrcher I,Hood DA.Regulation of Egr-l,SRF,and Spl mRNA expression in contracting skeletal muscle cells.J Appl Physiol.2004;97:2207-2213.
74 Okamoto H.Osteopontin and cardiovascular system.Mol Cell Biochem.2007;300:1-7.
75 Scatena M,Liaw L,Giachelli CM.Osteopontin:a multifunctional molecule regulating chronic inflammation and vascular disease.Arterioscler Thromb Vase Biol.2007;27:2302-2309.
76 Lee JL,Wang MJ,Sudhir PR,et al.Osteopontin promotes integrin activation through outside-in and inside-out mechanisms:OPN-CD44V interaction enhances survival in gastrointestinal cancer cells.Cancer Res.2007;67:2089-2097.
77 Infanger M,Grosse J,Westphal K.Vascular endothelial growth factor induces extracellular matrix proteins and osteopontin in the umbilical artery.Ann Vase Surg.2008;22:273-284.
78 Abe K,Nakashima H,Ishida M,et al.Angiotensin Ⅱ-induced osteopontin expression in vascular smooth muscle cells involves Gq/11,Ras,ERK,Src and Ets-l.Hypertens Res.2008;31:987-998.
79 Jeon YM,Kook SH,Son YO Role of MAPK in mechanical force-induced up-regulation of type Ⅰ collagen and osteopontin in human gingival fibroblasts.Mol Cell Biochem.2009;320:45-52.
80 Zhu B,Suzuki K,Goldberg HA.Osteopontin modulates CD44-dependent chemotaxis of peritoneal macrophages through G-protein-coupled receptors:evidence of a role for an intracellular form of osteopontin.J Cell Physiol.2004;198:155-167.
81 Gordon JA,Hunter GK,Goldberg HA.Activation of the mitogen-activated protein kinase pathway by bone sialoprotein regulates osteoblast differentiation.Cells Tissues Organs.2009;189:138-143.
82 CollissonEA,De A,Suzuki H.Treatolent of metastatic melanoma with an orally available inhibitor of the Ras-Raf-MAPK caseade.CaneerRes.2003;63:5669-5673.
83 Lee JT Jr,Steelman LS,MeCubrey JA.Modulation of Raf/MEK/ERK kinase activity does not after the chemoresistance profile of advanced prostate cancer cells.Int J Oncol.2005;26:1637-1644.
84 Moon Y,Yang H,Kim YB.Up-regulation of early growth response gene 1 (EGR-1)via ERK signals attenuates sulindac sulfide-mediated cytotoxicity in the human intestinal epithelial cells.Toxicol Appl Pharmacol.2007;223:155-163.
85 Shin SY,Lee JH,Min B,et al.The translation inhibitor anisomycin induces Elk-1-mediated transcriptional activation of egr-1 through multiple mitogen-activated protein kinase pathways.Exp Mol Med.2006;38:677-685.
86 Guha M,0'Connell MA,Pawlinski R,et al.Lipopolysaccharide activation of the MEK-ERK pathway in human monocytic cells mediates tissue factor and tumor necrosis factor alpha expression by inducing Elk-1 phosphorylation and Egr-1 expression.Blood.2001;98:1429-1439.
1 Sukhatme VP,Cao XM,Chang LC,et al.A zinc finger-encoding gene coregulated with c-fos during growth and differentiation,and after cellular depolarization.Cell.1988;53:37-43.
2 朱慧,曹传华,谢丛华.Egr-1启动子辐射诱导的基因表达特性.武汉大学学报(医学版).2008;29:186-189.
3 Thiel G,Cibelli G.Regulation of life and death by the zinc finger transcription factor Egr-1.J Cell Physio.2002;193:287-292.
4 A1-Sarraj A,Day RM,Thiel G.Specificity of transcriptional regulation by the zinc finger transcription factors Sp1,Sp3,and Egr-1.J Cell Biochem.2005,94:153-167.
5 Gonz(?)lez P,Reguero JR,Lozano I,et al.A functional Sp1/Egr1-tandem repeat polymorphism in the 5-lipoxygenase gene is not associated with myocardial infarction.Int J Immunogenet.2007;34:127-130.
6 Kostadinova RM,Nawrocki AR,Frey FJ,et al.Tumor necrosis factor alpha and phorbol 12-myristate-13 acetate down-regulate human 11beta-hydroxysteroid dehydrogenase type 2through p50/p50 NF-kappaB homodimers and Egr-1.FASEB J.2005;19:650-652.
7 DUN,Pei XT,Sun JZ,et al.Regulatory Effect of Egr-1 Promoter Sequences Induced by Doxorubicin in Transcriptional Targeting on Expression of GM-CSF Gene.Zhongguo Shi Yan Xue Ye Xue Za Zhi.2008;16:1165-1169.
8 Kumbrink J,Gerlinger M,Johnson JP.Egr-1 induces the expression of its corepressor nab2 by activation of the nab2 promoter thereby establishing a negative feedback loop.J Biol Chem.2005,280:42785-42793.
9 庄楚香,吴名耀.Egr-1在小鼠和人体正常组织中的表达及其生理学功能.汕头大学医学院学报.2001;14:166-168.
10 吴名耀,庄楚香.Egr-1在小鼠和人组织中的表达及其与细胞增殖的关系.中国组织化学与细胞化学杂志.2002;11:72-74.
11 Ogata T.Egr-1 mRNA induction by medium flow involves mRNA stabilization and is enhanced by the p38 inhibitor SB203580 in osteoblast-like cells.Acta Physiol(Oxf).2008;194:177-188.
12 Irrcher I,Hood DA.Regulation of Egr-1,SRF,and Spl mRNA expression in contracting skeletal muscle cells.J Appl Physiol.2004;97:2207-2213.
13 Fukada T,Tonks NK.The reciprocal role of Egr-1 and Sp family proteins in regulation of the PTP1B promoter in response to the p210 Bcr-Abl oncoprotein-tyrosine kinase.J Biol Chem.2001;276:25512-25519.
14 Scharnhorst V,Menke AL,Attema J,et al.EGR-1 enhances tumor growth and modulates the effect of the Wilms' tumor 1 gene products on tumorigenicity.Oncogene.2000;19:791-800.
15 Chapman NR,Perkins ND.Inhibition of the RelA(p65)NF-kappaB subunit by Egr-1.J Biol Chem.2000;275:4719-4725.
16 Goetze S,Kintscher U,Kaneshiro K,et al.TNFalpha induces expression of transcription factors c-fos,Egr-l,and Ets-1 in vascular lesions through extracellular signal-regulated kinases l/2.Atherosclerosis.2001;159:93-101.
17 Mechtcheriakova D,Schabbauer G,Lucerna M,et al.Specificity,diversity,and convergence in VEGF and TNF-alpha signaling events leading to tissue factor up-regulation via EGR-1 in endothelial cells.FASEB J.2001;15:230-242.
18 Gousseva N,Kugathasan K,Chesterman CN,et al.Early growth response factor-1 mediates insulin-inducible vascular endothelial cell proliferation and regrowth after injury.J Cell Biochem.2001;81:523-534.
19 Das A,Chendil D,Dey S,et al.Ionizing radiation down-regulates p53 protein in primary Egr-1-/-mouse embryonic fibroblast cells causing enhanced resistance to apoptosis.J Biol Chem.2001;276:3279-3286.
20 Guha M,O'Connell MA,Pawlinski R,et al.Lipopolysaccharide activation of the MEK-ERK pathway in human monocytic cells mediates tissue factor and tumor necrosis factor alpha expression by inducing Elk-1 phosphorylation and Egr-1 expression.Blood.2001;98:1429-1439.
21 Lo LW,Cheng JJ,Chiu JJ,et al.Endothelial exposure to hypoxia induces Egr-1 expression involving PKCalpha-mediated Ras/Raf-1/ERK pathway.J Cell Physiol.2001;188:304-312.
22 Day RM,Yang Y,Suzuki YJ,et al.Bleomycin upregulates gene expression of angiotensin- -converting enzyme via mitogen-activated protein kinase and early growth response 1 transcription factor.Am J Respir Cell Mol Biol.2001;25:613-619.
23 De Sousa LP,Brasil BS,Silva BM,et al.Plasminogen/plasmin regulates c-fos and egr-1 expression via the MEK/ERK pathway.Biochem Biophys Res Commun.2005;329:237-245.
24 Moon Y,Yang H,Kim YB.Up-regulation of early growth response gene 1 (EGR-1)via ERK signals attenuates sulindac sulfide-mediated cytotoxicity in the human intestinal epithelial cells.Toxicol Appl Pharmacol.2007;223:155-163.
25 Shin SY,Lee JH,Min B,et al.The translation inhibitor anisomycin induces Elk-1-mediated transcriptional activation of egr-1 through multiple mitogen-activated protein kinase pathways.Exp Mol Med.2006;38:677-685.
26 Wu SQ,Minami T,Donovan DJ,et al.The proximal serum response element in the Egr-1 promoter mediates response to thrombin in primary human endothelial cells.Blood.2002;100:4454-4461.
27 Shin SY,Lee JH,Min B,et al.The translation inhibitor anisomycin induces Elk-1-mediated transcriptional activation of egr-1 through multiple mitogen-activated protein kinase pathways.Exp Mol Med.2006;38:677-685.
28 Grimes CA,Jope RS.Cholinergic stimulation of early growth response-1 DNA binding activity requires protein kinase C and mitogen-activated protein kinase kinase activation and is inhibited by sodium valproate in SH-SY5Y cells.J Neurochem.1999;73:1384-1392.
29 Yan SF,Lu J,Zou YS,et al.Protein kinase C-beta and oxygen deprivation.A novel Egr-1--dependent pathway for fibrin deposition in hypoxemic vasculature..! Biol Chem.2000;275:11921-11928.
30 Mechtcheriakova D,Wlachos A,Holzmuller U.,et al.Vascular endothelial cell growth factor-induced tissue factor expression in endothelial cells is mediated by EGR-1.Blood.1999;93:3811-3823.
31 Fang M,Wee SA,Ronski K,et al.Evidence of EGR1 as a differentially expressed gene among proliferative skin diseases.Genomic Med.2007;1:75-85.
32 Du B,Fu C,Kent KC,et al.Elevated Egr-1 in human atherosclerotic cells transcriptionally represses the transforming growth factor-beta type Ⅱ receptor.J Biol Chem.2000;275:39039-39047.
33 Braddock M.The transcription factor Egr-1:a potential drug in wound healing and tissue repair.Ann Med.2001;33:313-318.
34 Liu C,Yao J,Mercola D,et al.The transcription factor EGR-1 directly trans--activates the fibronectin gene and enhances attachment of human glioblastoma cell line U251.J Biol Chem.2000;275:20315-20323.
35 Bryant M,Drew GM,Houston P,et al.Tissue repair with a therapeutic transcription factor.Hum Gene Ther.2000;11:2143-2158.
36 Houston P,Campbell CJ,Svaren J,et al.The transcriptional corepressor NAB2 blocks Egr-1-mediated growth factor activation and angiogenesis.Biochem Biophys Res Commun.2001;283:480-486.
37 Nakamura H,Isaka Y,Tsujie M,et al.Introduction of DNA enzyme for Egr-1 into tubulointerstitial fibroblasts by electroporation reduced interstitial alpha-smooth muscle actin expression and fibrosis in unilateral ureteral obstruction(UUO)rats.Gene Ther.2002;9:495-502.
38 钟守军,姚运红,李飞虹,等.EGR-1 与p53,p16,Cyclin D1在鼻咽癌中的表达及意义.肿瘤研究与临床.2006;18:664-666.
39 Lee SW,Kim EJ,Um SJ.Transcriptional regulation of the p73 gene,a member of the p53 family,by early growth response-1(Egr-1).Biochem Biophys Res Commun.2008;373:455.
40 40.Cibelli G,Policastro V,R(o|¨)ssler OG,et al.Nitric oxide-induced programmed cell death in human neuroblastoma cells is accompanied by the synthesis of Egr-1,a zinc finger transcription factor.J Neurosci Res.2002;67:450-460.
41滕雅轩,刘闺男,周敬.早期反应生长因子1特异的脱氧核酶对大鼠动脉损伤后血管内皮功能的保护及超微结构的影.中国动脉硬化杂志.2005;13:709-713.
42 Eyries M,Agrapart M,Alonso A,et al.Phorbol ester induction of angiotensin-converting enzyme transcription is mediated by Egr-1 and AP-1 in human endothelial cells via ERK pathway.Circ Res.2002;91:899-906.
43 Yan SF,Mackman N,Kisiel W,et al.Hypoxia/Hypoxemia-Induced activation of the procoagulant pathways and the pathogenesis of ischemia-associated thrombosis.Biol.1999;19:2029-2035.
44 Ohtani K,Egashira K,Usui M,et al.Inhibition of neointimal hyperplasia after balloon injury by ciselement 'decoy' of early growth response gene-1 in hypercholesterolemic rabbits.Gene Ther.2004;l 1:126-132.
45 Wada Y,Suzuki J,Kawauchi M,et al.Early growth-response factor 1 and basic transcriptional element-binding protein 2 expression in cardiac allografts.J Heart Lung Transplant.2001;20:590-594.
46 Lowe HC,Fahmy RG,Kavurma MM,et al.Catalytic oligodeoxynucleotides define a key regulatory role for early growth response factor-1 in the porcine model of coronary in-stent restenosis.CircRes.2001;89:670-677.
47 Rupp J,Hellwig-Burgel T,Wobbe V,et al.Chlamydia pneumoniae infection promotes a proliferative phenotype in the vasculature through Egr-1 activation in vitro and in vivo.Proc Natl Acad Sci USA.2005;102:3447-3452.
48 Fahmy RG,Khachigian LM.et al.Suppression of growth factor expression and human vascular smooth muscle cell growth by small interfering RNA targeting EGR-l.J Cell Biochem.2007;100:1526-1535.
49 Zhang YM,Shi GG,Tang Z,et al.Effects of N-n-butyl haloperidol iodide on myocardial ischemia/reperfusion injury and Egr-1 expression in rat.Acta Biochim Biophys Sin (Shanghai).2006;38:435-441.
50 Fahmy RG,Khachigian LM.Suppression of growth factor expression and human vascular smooth muscle cell growth by small interfering RNA targeting EGR-l.J Cell Biochem.2007;100:1526-1535.
51 Khachigian LM.Catalytic oligonucleotides targeting EGR-1 as potential inhibitors of in-stent restenosis.Ann N Y Acad Sci.2001;947:412-415.
52 Santiago FS,Atkins DG,Khachigian LM.Vascular smooth muscle cell proliferation and regrowth after mechanical injury in vitro are Egr-1/NGFI-A-dependent.Am J Pathol.1999;155:897-905.
53 Nagai R,Suzuki T,Aizawa K,et al.Phenotypic modulation of vascular smooth muscle cells:dissection of transcriptional regulatory mechanisms.Ann N Y Acad Sci.2001;947:56-66.
54 Zhu X,Lin Y,Bacanamwo M,et al.Interleukin-1 beta-induced Id2 gene expression is mediated by Egr-1 in vascular smooth muscle cells.Cardiovasc Res.2007;76:141-148.
55 Liang KW,Ting CT,Yin SC,et al.Berberine suppresses MEK/ERK-dependent Egr-1 signaling pathway and inhibits vascular smooth muscle cell regrowth after in vitro mechanical injury.Biochem Pharmacol.2006;71:806-817.
56 Kamimura M,Bea F,Akizawa T,et al.Platelet-derived growth factor induces tissue factor expression in vascular smooth muscle cells via activation of Egr-1.Hypertension.2004;44:944-951.
57 滕雅轩,赵卫华,刘闺男,等.针对 Egr-1mRNA 的10-23脱氧核酶抑制大鼠动脉损伤后内膜增生的研究.中国现代医学杂志.2006;16:1823-1827.
58 吴岩,刘闺男,韩伟,等.Egr-1特异脱氧核酶抑制血管平滑肌细胞增殖的研究.山东医药.2008;48:53-55.
59 刘程伟,胡新华,张雪松,等.Egr-1 在自体移植静脉中的表达及意义.中国普外基础与临床杂志.2006;13:642-646.
60 Patrizio M,Musumeci M,Stati T,et al.Propranolol promotes Egr1 gene expression in cardiomyocytes via beta-adrenoceptors.Eur J Pharmacol.2008;587:85-89.
61 Bhindi R,Khachigian LM,Lowe HC.DNAzymes targeting the transcription factor Egr-1 reduce myocardial infarct size following ischemia-reperfusion in rats.J Thromb Haemost.2006;4:1476-1478.
62 Zhang Y,Shi G,Zheng J,et al.The protective effects ofNbutyl haloperidol iodide on myocardial ischemia-reperfusion injury in rats by inhibiting Egr-1 overexpre--ssion.Cell Physiol Biochem.2007;20:639-648.
63 Wang C,Dostanic S,Servant N,et al.Egr-1 negatively regulates expression of the sodium-calcium exchanger in cardiomyocytes in vitro and in vivo.Cardiovasc Res.2005;65:187-194.
64 Hara S,Arai M,Tomaru K,et al.Prostaglandin F2alpha inhibits SERCA2 gene transcription through an induction of Egr-1 in cultured neonatal rat cardiac myocytes.Int Heart J.2008;49:329-342.
65 初令,李翔,胡永斌,等.大鼠实验性矽肺早期生长反应基因表达及其作用的初步探讨.中华劳动卫生职业病杂志.2004;22:47-50.
66 Yan SF,Fujita T,Lu J,et al.Egr-1,a master switch coordinating upregulation of divergent gene families underlying ischemic stress.Nat Med.2000;6:1355-1361.
67 李海青,石刚刚,高分飞,等.钙拮抗剂对大鼠心肌缺血再灌注早期生长反应基因-1 表达的影响.汕头大学医学院学报.2008;21:7-10.
68 Arai M,Yoguchi A,Takizawa T,et al.Mechanism of doxorubicin-induced inhibition of sarcoplasmic reticulum Ca(2+)-ATPase gene transcription.Circ Res.2000;86:8-14.
69 Okada M,Fujita T,Sakaguchi T,et al.Extinguishing Egr-1-dependent inflammatory and thrombotic cascades after lung transplantation.F ASEB J.2001;15:2757-2759.
70 Okada M,Wang CY,Hwang DW,et al.Transcriptional control of cardiac allograft vasculopathy by early growth response gene-1(Egr-1).Circ Res.2002;91:135-142.
1 傅国香,高平进.骨桥蛋白与血管损伤的研究进展.中华高血压杂志,2006;14:603-605.
2 Helluin O,Chan C,Vilaire G,et al.The activation state of alphav beta 3 regulates platelet and lymphocyte adhesion to intactand thrombin-cleavaged osteopontin.J Biol Chem.2000;275:18337-18343.
3 林全,王良兴.骨桥蛋白与血管重塑的关系研究进展.医学综述.2004;10:731-733.
4 杨淑莉,韩梅,温进坤.骨桥蛋白与心血管疾病.中国动脉粥样硬化杂志.2000;8:371-373.
5 方欣,管思明.骨桥蛋白与高血压血管重构.心脏杂志.2007;19:232-233.
6 Gao C,Guo H,Downey L,et al.Osteopontin-dependent CD44v6 expression and cell adhesion in HepG2 cells.Carcinogenesis.2003;24:1871-1878.
7 Soejima H,Irie A,Fukunaga T,et al.Elevated plasma osteopontin levels were associated with osteopontin expression of CD4+ T cells in patients with unstable angina.Circ J.2006;70:851-856.
8 陈荣新,叶胜龙.骨桥蛋白与肿瘤转移的研究进展.国际外科学杂志.2006;33:16-18.
9 俞文华,张国新.骨桥蛋白与胶质溜研究进展.医学研究杂志.2006;35:91-93.
10 范哲,李鸿雁,龙毅等.骨桥蛋白的研究进展.中国实验诊断学.2005;9:307-309.
11 Pereira RO,Carvalho SN,Stumbo AC,et al.Osteopontin expression in coculture of differentiating rat fetal skeletal fibroblasts and myoblasts.In Vitro Cell Dev Biol Anim.2006;42:4-7.
12 Schoensiegel F,Bekeredjian R,Schrewe A,et al.Atrial natriuretic peptide and osteopontin are useful markers of cardiac disorders in mice.Comp Med.2007;57:546-553.
13 Yamate T,Kohri K,Umekawa T,et al.Interaction between osteopontin on madin darby canine kidney cell membrane and calcium oxalate crystal.Urol Int.1999;62:81-86.
14 Hickey FB,England K,Cotter TG.Bcr-Abl regulates osteopontin transcription via Ras,PI-3K,aPKC,Raf-l,and MEK.J Leukoc Biol.2005;78:289-300.
15 Li G,Oparil S,Kelpke SS,et al.Fibroblast growth factor receptor-1 signaling induces osteopontin expression and vascular smooth muscle cell-dependent adventitial fibroblast migration in vitro.Circulation.2002;106:854-859.
16 Renault MA,Jalvy S,Belloc I,et al.AP-1 is involved in UTP-induced osteopontin expression in arterial smooth muscle cells.Circ Res.2003;93:674-681.
17 周秀霞,温进坤,韩梅.白细胞介素-1β和肿瘤坏死因子-α对血管平滑肌细及基质金属蛋白酶-2骨桥蛋白基因表达的影响.中国动脉硬化杂志.1999;7:292.
18 傅国香,朱鼎良,刘建军等.骨桥蛋白参与血管紧张素Ⅱ诱导的血管平滑肌细胞迁移.高血压杂志.2006;14:277-280.
19 Jalvy S,Renault MA,Lam Shang Leen L,et al.CREB mediates UTP-directed arterial smooth muscle cell migration and expression of the chemotactic protein osteopontin via its interaction with activator protein-1 sites.Circ Res.2007;100:1292-1299.
20 Nakamura H,Honda H,Inada Y,et al.Osteopontin expression in vascular smooth muscle cells in patients with end-stage renal disease.Ther Apher Dial.2006;10:273-277.
21 Li G,Chen YF,Kelpke SS,et al.Estrogen attenuates integrin-beta(3)-dependent adventitial fibroblast migration after inhibition of osteopontin production in vascular smooth muscle cells.Circulation.2000;101:2949-2955.
22 Sugiyama T,Yoshimoto T,Hirono Y,et al.Aldosterone increases osteopontin gene expression in rat endothelial cells.Biochem Biophys Res Commun.2005;336:163-167.
23 Takemoto M,Yokote K,Yamazaki M,et al.Enhanced expression of osteopontin by high glucose in cultured rat aortic smooth muscle cells.Biochem Biophys Res Commun.1999;258:722-726.
24 Isoda K,Kamezawa Y,Ayaori M,et al.Osteopontin transgenic mice fed a high-cholesterol diet develop early fatty-streak lesions.Circulation.2003;107:679-681.
25 Zhu Y,Denhardt DT,Cao H,et al.Hypoxia upregulates osteopontin expression in NIH-3T3 cells via a Ras-activated enhancer.Oncogene.2005;24:6555-6563.
26 俞广进,熊其如.骨桥蛋白在肿瘤转移中的研究进展.临床肿瘤学杂志.2007;12:156-160.
27 Sodhi CP,Phadke SA,Batlle D,et al.Hypoxia stimulates osteopontin expression and proliferation of cultured vascular smooth muscle cells:potentiation by high glucose.Diabetes.2001;50:1482-1490.
28 Fujihara S,Yokozeki M,Oba Y,et al.Function and regulation of osteopontin in response to mechanical stress.J Bone Miner Res.2006;21:956-564.
29 Tanaka N,Momiyama Y,Ohmori R,et al.Effect of atorvastatin on plasma osteopontin levels in patients with hypercholesterolemia.Arterioscler Thromb Vase Biol.2006;26:129-130.
30 Luomala M,P(a|¨)iv(a|¨)H,Thelen K,et al.Osteopontin levels are associated with cholesterol synthesis markers in mildly hypercholesterolaemic patients.Acta Cardiol.2007;62:177-181.
31 Hu T,Li H,Zhang X,et al.Exposure of aortic vascular smooth muscle cells to low-frequency electromagnetic field inhibits osteopontin expression and matrix metalloproteinase activity.Int Heart J.2008;49:597-604.
32 Ashkar S,weber GF,Panoutsakopoulou V,et al.Eta-l(esteopentin):an early component of type-1(cell-mediated)immunity.Science.2000;287:860-864.
33 李秉慧,宋杏丽,周强,等.骨桥蛋白促创面愈合的实验研究.中华实验外科杂志.2004:21:1355-1357.
34 Khan SA,Cook AC,Kappil M,et al.Enhanced cell surface CD44 variant(v6,v9)expression by osteopontin in breast cancer epithelial cells facilitates tumor cell migration:novel post-transcriptional,post-translational regulation.Clin Exp Metastasis.2005;22:663-673.
35 Hikita ST,Vistica BP,Jones HR,et al.Osteopontin is proinflammatory in experimental autoimmune uveitis.Invest Ophthalmol Vis Sci.2006;47:4435-4443.
36 Ariztia E.Osteopontin contributes to hepatocyte growth factor induced tumor growth and metastasis formation.Exp Cell Res.2003;228:257-267.
37 Moye VE,Barraeloush R,West C,et al.Osteopontin expression correlates with adhesive and metastatic potential in metastasis indueing DNA-transfected rat mammary cell lines.Br J Canc.er.2004;90:1796-1802.
38 Irby RB,McCarthy SM,Yeatman TJ,et al.Osteopontin regulates multiple functions contributing to human colon cancer development and progression.Clin Exp Metastasis.2004;21:515-523.
39 Ganmann A,Petrow P,Mentzel T,et al.Osteopontin expression in primary sarcomas of the pulmonary artery.Virchows Arch.2001;439:668-674.
40 付汐,杨竹林,李代强,等.胰腺癌组织中 kail 和骨桥蛋白表达及其临床病理意义.检验与临床.2006;8:1690-1691.
41 张辉,应敏刚,任宁.肝细胞癌患者血桨骨桥蛋白表达水平与术后复发及愈后的关系.中国临 床医学．2007；14：1-6．
42 Pan HW,Ou YH,Peng SY,et al.Overexpression of osteopontin is associated with intrahepatic metastasis,early recurrence,and poorer prognosis of surgically reseeted hepatocellular carcinoma.Cancer.2003;98:119-127.
43 Nakamura H,Honda H,Inada Y,et al.Osteopontin expression in vascular smooth muscle cells in patients with end-stage renal disease.Ther Apher Dial.2006;10:273-277.
44 Nakae MJwamoto I,Fujino T,et al.Preoperative plasma osteopontin level as a biomarker complementary to carbohydrate antigen 125 in predicting ovarian cancer.J Obstet Gynaecol Res.2006;32:309-314.
45 Zhang YL,Zhou SX,Lei i,et ah Blockades of angiotensin and aldosterone reduce osteopontin expression and interstitial fibrosis infiltration in rats with myocardial infarction.Chin Med J (Engl).2008;121:2192-2196.
46 Coskun S,Atalar E,Ozturk E,et al.Plasma osteopontin levels are elevated in non-ST-segment elevation acute coronary syndromes.J Natl Med Assoc.2006;98:1746-1750.
47 Singh M,Ananthula S,Milhorn DM,et al.Osteopontin:a novel inflammatory mediator of cardiovascular disease.Front Biosci.2007;12:214-221.
48 Soejima H,Irie A,Fukunaga T,et al.Osteopontin expression of circulating T cells and plasma osteopontin levels are increased in relation to severity of heart failure.Circ J.2007;71:1879-1884.
49 Georgiadou P,Iliodromitis EK,Kolokathis F,et al.Plasma levels of osteopontin before and 24 h after percutaneous coronary intervention.Expert Opin Ther Targets.2008;12:1477-1480.
50 Kato R,Momiyama Y,Ohmori R,et al.Prognostic significance of plasma osteopontin levels in patients undergoing percutaneous coronary intervention.Circ J.2009;73:152-157.
51 Zhao X,Johnson JN,Singh K,et al.Impairment of myocardial angiogenic response in the absence of osteopontin.Microcirculation.2007;14:233-240.
52 Yin FY,Sun JH,Han YY.Experimental study on expression of osteopontin in viral myocarditis.Zhonghua Er Ke Za Zhi.2008;46:710-711.
53 Qiu ZB,Wan S.Association between osteopontin and restenosis.Zhonghua Xin Xue Guan Bing Za Zhi.2008;36:470-472.
54 Kato R,Momiyama Y,Ohmori R,et al.High plasma levels of osteopontin in patients with restenosis after percutaneous coronary intervention.Arterioscler Thromb Vasc Biol.2006;26:1-2.
55 孙爱军,高平进,朱鼎良,等.骨桥蛋白在血管外膜中的表达及其对外膜成纤维细胞迁移的影响.中华心血管病杂志.2003;5:38-47.
56 孙爱军,高平进,刘建军,等骨桥蛋白增强自发性高血压大鼠血管外膜成纤维细胞的迁移活性.生理学报.2004;56:21-24.
57 Bostr(o|¨)m K.Osteopontin,a missing link in PDGF-induced smooth muscle cell migration.Cardiovasc Res.2007;75:634-635.
58 Jalvy S,Renault MA,Leen LL,et al.Autocrine expression of osteopontin contributes to PDGF-mediated arterial smooth muscle cell migration.Cardiovasc Res.2007;75:738-747.
59 Roy S,Ruest PJ,Hanks SK.FAK regulates tyrosine phosphorylation of CAS,paxillin,and PYK2 in cells expressing v-Src,but is not a critical determinant of v-Src transformation.J Cell Biochem.2002;84:377-388.
60 Yin BL,Hao H,Wang YY,et al.Downregulating osteopontin reduces angiotensin Ⅱ-induced inflammatory activation in vascular smooth muscle cells.Inflamm Res.2009;58:67-73.
61 Mori S,Takemoto M,Yokote K,et al.Hyperglycemia-induced alteration of vascular smooth muscle phenotype.Diabetes Com placations.2002;16:65-68.
62.Jing-Jing Li,Mei Han,Jin-Kun Wen,et al.Osteopontin stimulates vascular smooth muscle cell migration by inducing FAK phosphorylation and ILK dephosphorylation.Biochemical and Biophysical Research Communications.2007;356:13-19.
63 李菁菁,温进坤,韩梅.Src 介导骨桥蛋白诱导的血管平滑肌细胞黏附和迁移过程.细胞生物学杂志.2006;28:757-762.
64 李菁菁,温进坤,韩梅.FAK 和 ILK 介导骨桥蛋白诱导的血管平滑肌细胞黏附和迁移.中国生物化学与分子生物学报.2006;2:477-483.
65 Li JJ,Han M,Wen JK,et al.Osteopontin stimulates vascular smooth muscle cell migration by inducing FAK phosphorylation and ILK dephosphorylation.Biochem Biophys Res Commun. 2007;356:13-19.
66 K han SA,Lopez-Chua CA,Zhang J,et al.Soluble osteopontin inhibits apoptosis of adherent endothelial cells deprived of growth factors.J Cell Biochem.2002;85:728-736.
67 Weintraub AS,Schnapp LM,Lin X,et al.Osteopontin deficiency in rat vascular smooth muscle cells is associated with an inability to adhere to collagen and increased apoptosis.Lab Invest.2000;80:1603-1615.
68 Abel B,Kurrer M,Shamshiev A,et al.The osteopontin-CD44 pathway is superfluous for the development of autoimmune myocarditis.Eur J Immunol.2006;36:494-499.
69 Lin YH,Yang-Yen HF.The osteopontin-CIM4 survival signal involves activation of the phosphatidylinositol 3-kinase/Akt signalingpathway.J Biol Chem.2001;276:46024-46030.
70 Malyankar UM,Seatena M,Suchland KL,et al.osteoprotegerln is an alpha v beta 3-induced.NF-kappa B-dependent survival factor for endothdial cells.J Biol Chem.2001;275:20959-20962.
71 Shin T,Ahn M,Kim K,et al.Increased expression of osteopontin in the heart tissue of Lewis rats with experimental autoimmune myocarditis.J Vet Med Sci.2006;68:379-382.
72 Abe K,Nakashima H,Ishida M,et al.Angiotensin Il-induced osteopontin expression in vascular smooth muscle cells involves Gq/11,Ras,ERK,Src and Ets-l.Hypertens Res.2008;31:987-998.
73 Kurata M,Okura T,Watanabe S,et al.Osteopontin and carotid atherosclerosis in patients with essential hypertension.Clin Sci (Lond).2006;l 11:319-324.
74 Li G,Oparil S,Kelpke SS,et al.Fibroblast growth factor receptor-1 signaling induces osteopontin expression and vascular smooth muscle cell-dependent adventitial fibroblast migration in vitro.Circulation.2002;106:854-859.
75 Roy S,Ruest PJ,Hanks SK.FAK regulates tyrosine phosphorylation of CAS,paxillin,and PYK2 in cells expressing v-Src,but is not a critical determinant of v-Src transformation.J Cell Biochem.2002;84:377-388.
76 Xie B,Zhao J,Kitagawa M,et al.Focal adhesion kinase activates Statlin integrin 2 mediated cell migration and adhesion.J Cel Biochem.2001;76:19512-119523.
77 Tuck AB,Hota C,Wilson SM,et al.Osteopontin-induced migration of human mammary epithelial cells involves activation of EGF receptor and multiple signal transduction pathways.Oncogene.2003;22:1198-1205.
78 Rangaswami H,Bulbule A,Kundu GC.Nuclear factor inducing kinase:a key regulator in osteopontin-induced MAPK/DcappaB kinase dependent NF-kappaB-mediated promatrix metalloproteinase-9 activation.Glycoconj J.2006;23:221-232.
79 Nerurkar SS,01zinski AR,Frazier KS,et al.P38 MAPK inhibitors suppress biomarkers of hypertension end-organ damage,osteopontin and plasminogen activator inhibitor-1.Biomarkers.2007;12:87-112.
80 Tuck AB,Hota C,Wilson SM,et al.Osteopontin-induced migration of human mammary epithelial cells involves activation of EGF receptor and multiple signal transduction pathways.Oncogene.2003;22:1198-1205.
81 Farb A,Kolodgie FD,Hwang JY,et al.Extracellular matrix changes in stented human coronary arteries.Circulation.2004;24,110:940-947.
82 Asa Strom,Ahnders Franzen.Altered vascular remodeling in osteopontin-deficient atherosclerotic mice.J Vase Res.2004;41:314-322.
83 Lai CF,Seshadri V,Hiiang K,et al.An osteopontin-NADPH oxidase signaling cascade promotes pro-matrix metalloproteinase 9 activation in aortic mesenchymal cells.Circ Res.2006;98:1479-1489.
84 Kikuo Isoda,Knichirou Nishikawa,Yashuhiro Kamezawa,et al.Osteopontin plays an important role in the development of medial thickening and neointimal formation.Circ Res.2002;91:77-82.