血管内皮生长因子调控胃癌及胆管癌侵袭转移作用的实验研究
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摘要
研究背景和意义
恶性肿瘤的侵袭和转移是一个多步骤、多阶段的复杂过程。癌变的细胞和组织会发生一系列的变化,包括细胞转化、肿瘤生长、血管及淋巴管的诱导生成及癌细胞扩散到循环系统；然后在靶器官中粘附、侵袭和增殖。目前已经有数种细胞因子被证实参与肿瘤的转移；其中,血管内皮生长因子(vascular endothelial growth factor, VEGF)不仅参与肿瘤血管的生成,还参与伤口愈合及其他重要的病理过程。VEGF通过两种机制诱导肿瘤血管的生成,一是主要通过其同源性受体VEGFR2,二是通过非VEGFR途径,其中包括整合素的上调和(或)激活。
整合素是一种异二聚体跨膜蛋白,通过VEGF及其受体的协调,能够诱导细胞粘附于细胞外基质(extracellular matrix,ECM)。整合素αvβ6在正常的上皮细胞几乎不表达,而在生长中的胚胎组织及许多上皮性恶性肿瘤中高表达。前期研究中,我们已经证实了整合素αvβ6可用来作为胃癌预后的指标之一。另外αvβ6在结肠癌中的表达可促进基质金属蛋白-9(matrix metalloprotein-9,MMP-9)分泌的增加。在胃癌组织中,MMP-9可降解细胞外基质,从而促进肿瘤细胞的播散和转移。由此可见,αvβ6通过MMP-9降解ECM屏障和介导肿瘤细胞的侵袭性。
有研究证实整合素与VEGF在细胞的粘附、侵袭和转移过程中具有协同作用。整合素α1β1和α2β1可促进VEGF介导的信号转导和内皮细胞的侵袭作用。VEGF与整合素共同参与多发性骨髓瘤细胞的侵袭。在结肠癌细胞中,α9β1阻断抗体可以特异性的抑制VEGF-A诱导的血管生成。然而,整合素αvβ6在VEGF介导的肿瘤细胞侵袭转移过程中的作用尚未明确。
VEGF-C是VEGF家族的新成员,是一种酪氨酸激酶受体,其主要表达在内皮组织的淋巴系统内。在一些恶性肿瘤,如：乳腺癌、胃癌以及肺癌中呈高表达,并且有研究证实VEGF-C的表达与肿瘤的侵袭以及淋巴转移密切相关。但是很少研究VEGF-C在胆管癌中的表达以及与胆管癌侵袭转移之间的关系。
本次研究的目的是研究整合素αvβ6的表达和VEGF的关系,以及它们在体外对胃癌细胞侵袭性的影响；并且研究VEGF-C在胆管癌侵袭转移过程中所起的作用。
第一部分VEGF及整合素αvβ6在胃癌中的表达
目的研究VEGF以及整合素αvβ6在胃癌组织及细胞中的表达情况
方法收集手术切除的胃癌组织标本,冰冻储存于-80℃低温冰箱中。应用免疫组化的方法检测VEGF以及整合素αvβ6在胃癌组织的表达情况。体外培养胃癌AGS细胞,采用流式细胞技术、免疫荧光以及免疫沉淀检测胃癌细胞中整合素的表达情况。
结果30例胃癌标本的免疫组织化学分析显示,其中26例VEGF染色阳性,13例整合素αvβ6染色阳性。其中VEGF以及整合素αvβ6均表达阳性的有10例标本。流式细胞仪检测胃癌AGS细胞中整合素αvβ6呈现高表达,而且阳性率大于90%；细胞免疫荧光结果显示AGS细胞膜中整合素的表达,而应用免疫沉淀技术检查蛋白水平整合素αvβ6的表达情况。
结论VEGF以及整合素αvβ6在胃癌组织中均有表达；整合素αvβ6在胃癌细胞系AGS细胞中呈现高表达。
第二部分VEGF介导的胃癌细胞侵袭作用与整合素αvβ6相关
目的探讨整合素αvβ6在VEGF促进胃癌细胞侵袭、转移能力的作用,寻求控制胃癌转移的靶点。
方法体外培养AGS细胞,外源性VEGF(30ng／ml)刺激24h,应用流式细胞仪、Real-time PCR、Western blot以及免疫沉淀检测整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达变化。
再应用体外培养的AGS细胞,分别用ERK通路阻断剂U0126以及鼠抗人整合素αvβ6单克隆抗体10D5预处理后,外源性VEGF(30ng/ml)刺激24h,收集细胞并检测整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达变化。
针对人整合素基因序列设计其特异性siRNA序列。用荧光标记的阴性对照siRNA筛选转染条件,按照预实验筛选的转染条件将β6 siRNA转染到无血清培养液处理过的AGS细胞中,48h后收集细胞进行抑制效应分析,荧光定量PCR检测β6 mRNA的表达,以无血清培养基及对照siRNA为对照。AGS细胞转染β6 siRNA成功后,采用外源性VEGF刺激,收集细胞检测整合素β6、P-ERK和MMP-9在mRNA以及蛋白水平的表达。趋化小室试验检测外源性VEGF介导的AGS细胞侵袭能力。β6 siRNA转染的AGS细胞,以及分别用ERK通路阻断剂U0126以及鼠抗人整合素αvβ6单克隆抗体10D5预处理的AGS细胞,加入包被好的小室上层,在有无外源性VEGF介导的情况下,观察转移到小室下层的细胞个数,从而反应其不同条件下的侵袭能力。
结果应用免疫沉淀、流式细胞仪以及细胞免疫荧光,可以发现体外培养的AGS细胞内有整合素αvβ6的表达,其主要表达在细胞膜,而且表达的阳性率在90%以上。AGS细胞在外源性VEGF(30ng/ml)刺激24h后,整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达明显增加,与对照组相比具有统计学意义。而应用ERK通路阻断剂U0126以及鼠抗人整合素αvβ6单克隆抗体10D5预处理后,外源性VEGF(30ng/ml)刺激24h,整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达显著被抑制,与对照组相比具有统计学意义。荧光显微镜观察见筛选后的转染条件下荧光标记siRNA的转染效率较高,超过90%。在β6 siRNA转染的无血清培养液处理过的AGS细胞中,在有无外源性VEGF作用的条件下,整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达得到相同的结论。趋化小室实验检测到在外源性VEGF介导的情况下,转移至小室下层的AGS细胞数明显增加。而应用ERK通路阻断剂U0126、鼠抗人整合素αvβ6单克隆抗体10D5以及β6 siRNA转染预处理后,转移至小室下层的AGS细胞数显著减少,与对照组相比具有统计学意义。
结论胃癌AGS细胞内有整合素αvβ6的表达；外源性VEGF (30ng/ml)作用于AGS细胞后,可以刺激整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达显著增加以及提高AGS细胞的侵袭能力,而且这种作用可以被ERK通路阻断剂U0126、鼠抗人整合素αvβ6单克隆抗体10D5以及β6 siRNA转染预处理显著抑制,与对照组相比具有统计学意义。这说明整合素αvβ6参与了VEGF介导的胃癌侵袭转移作用,并且ERK通路在此过程中起着一定的作用。
第三部分VEGF-C在胆管癌中的表达及侵袭转移中的作用研究
目的研究VEGF-C在胆管癌癌组织中的表达情况及促进胆管癌癌细胞侵袭、转移能力的作用。
方法收集手术切除的胆管癌组织标本,冰冻储存于-80℃低温冰箱中。应用免疫组化的方法检测VEGF-C在胆管癌组织的表达情况,回顾性分析这些病人的临床病例资料,并与免疫组化结果相结合,分析VEGF-C的表达与患者临床资料之间的关系；体外培养胆管癌FRH-0201细胞,并且在有无外源性VEGF-C作用的情况下,采用增殖实验以及趋化小室实验检测胆管癌细胞FRH-0201的侵袭转移能力。
结果VEGF-C在胆管癌组织中呈高表达,表达率约为75.4%；其表达与病人的年龄、性别以及肿瘤大小、肿瘤的临床分期无明显关系,而与肿瘤的淋巴结转移明显相关。胆管癌细胞在外源性VEGF-C的作用的情况下,胆管癌细胞的增殖以及侵袭能力明显被增强,与对照组相比具有统计学意义。
结论VEGF-C在胆管癌组织中呈高表达并且在其侵袭转移过程中起着一定的作用。
Background and signification
Tumor metastasis is a complex and multi-stage process. A number of cellular and tissue changes occur as a cancer spreads, including cellular transformation and tumor growth, angiogenesis and lymphangiogenesis, and release of cancer cells into the circulation; then attachment, invasion, and proliferation within the new target organ. Several cytokines are now known to be involved in the process of tumor metastasis. Among these is vascular endothelial growth factor (VEGF), a factor that is associated not only with the angiogenesis of cancers but also with those involved in wound healing and other important pathologies. There are two mechanisms involved in VEGF-mediated angiogenesis; one is through its cognate receptor VEGFR2, the other is via up-regulation of the expression and/or activation of integrins.
Integrins are heterodimeric transmembrane proteins that can induce cell adhesion to the extracellular matrix (ECM), coordinated by VEGF and its receptors. The avP6 integrin is restricted to epithelial cells and is highly expressed in developing fetal tissues as well as in a number of epithelial carcinomas and cancers. In our previous studies, we demonstrated that integrin avP6 can be used as a prognostic indicator of gastric carcinoma. In addition,αvβ6 expression in colon cancer cells leads to the increase in secretion of matrix metalloprotein-9 (MMP-9). In gastric cancer, degradation of ECM barriers by MMP-9 is important in facilitating tumor cellular dissemination and metastasis. Thus, integrinαvβ6 appears to have roles in promoting ECM degradation and in mediating tumor cell invasiveness via MMP-9.
There are several confirmed collaborative relationships between integrins and VEGF in mediation of cell adhesion, migration, and proliferation. Theα1β1 andα2β2 integrins support VEGF-stimulated signal transduction and endothelial cell migration. Migration of multiple myeloma cells is induced by VEGF and associated withβ1 integrin. In colon cancer cells,α9β1-blocking antibody has been demonstrated to specifically inhibit the angiogenesis induced by VEGF-A. However, a role for integrinαvβ6 in VEGF-induced tumor progression has not yet been established.
VEGF-C is a new member of the VEGF family, a tyrosine kinase receptor that is predominantly expressed in the endothelium of lymphatic vessels. The expression of VEGF-C is high in several types of human malignant tumors, including breast, lung, and gastric carcinomas and some investigators have shown that VEGF-C expression was closely associated with tumor invasion and lymph node metastasis. However, there have been very few reports on the correlation between VEGF-C gene expression and invasive phenotype in cholangiocarcinoma.
The goal of the present study was therefore to investigate the relationship between VEGF and expression of integrinαvβ6 and their effects on invasiveness of gastric carcinoma cells in vitro. And the effect of VEGF-C in invasiveness and metastasis of cholangiocarcinoma.
PARTⅠExpression of VEGF and integrinαvβ6 in gastric carcinoma tissues and cells
Objective To study the expression of VEGF and integrinαvβ6 in gastric carcinoma tissues and cells.
Methods Thirty gastric cancer tissues were collected from patients requiring clinically indicated surgical resection in QiLu Hospital of Shandong University (Jinan, China), with informed patient consent. Frozen gastric cancer tissue samples were stored at -80℃until processing. Immunohistochemical analysis was used to detect the expression of VEGF and integrinαvβ6 in gastric carcinoma tissues. The human gastric cancer cell line AGS was maintained in RPMI 1640 medium supplemented with 10% fetal bovine serum (FBS), 100U/ml penicillin, 10μg/ml streptomycin, and 2mM L-glutamine. Immuno-precipitation, FAC scanning and immunofluorescence staining were used to confirme the presence ofαvβ6 in gastric cancer AGS cells.
Results Immunohistochemical analysis of tumor samples from 30 patients revealed that 26/30 samples stained positively for VEGF and 13/30 samples stained positively for integrinαvβ6. In 10/30 patients tumors were positive for both VEGF and integrinαvβ6. High surface expression ofαvβ6 integrin was detected by flow cytometry analysis, and the positive rate was higher than 90%. Expression ofαvβ6 was detected with immunofluorescent staining at the cell membrane in AGS cells. AGS cells were examined by immunoprecipitation analysis using antibody of 10D5 for expression ofαvβ6 integrin at protein levels.
Conclusions VEGF and integrinαvβ6 are all expressed in gastric carcinoma tissues. And the high surface expression ofαvβ6 integrin is detected in gastric cancer AGS cells.
PARTⅡVEGF enhances gastric carcinoma invasiveness via integrinαvβ6
Objective To investigate the effect of integrinαvβ6 on VEGF regulated invasiveness and metastasis in gastric carcinoma cells, to probe into the key point of regulating metastasis of gastric carcinoma.
Methods The AGS cells were starved for 4h in medium without fetal bovine serum and then stimulated with VEGF (30ng/ml) for 24h at 37℃. The expression at both mRNA and protein levels was also observed for integrinαvβ6, P-ERK and MMP-9 by using the real-time PCR, western blot analysis and immunoprecipitation.
The roles ofαvβ6, P-ERK and MMP-9 at both mRNA and protein levels were detected by using monoclonal antibody for the humanαvβ6 integrin (10D5) and an inhibitor of the ERK signaling pathway (U0126).
Specific P6 siRNA was designed according to the cDNA sequence of integrin subunit beta6 and the BLAST search was used to avoid unintentional silencing and ensure the uniqueness of target. Cells were grown subconfluently onto 24-well plates and transfected with 1μg ofβ6 siRNA, with nonspecific siRNA as control. After 48h of transfection, cells were washed and analyzed further as specifically indicated. The VEGF-stimulated transfected AGS cells were detected to calculate the change of integrinαvβ6, P-ERK and MMP-9 at both mRNA and protein levels.
Migration of AGS cells through Matrigel-coated filters was examined in 24-well transwell inserts. After trypsinization, 1×105 cells in 100μl were incubated with 10D5 or U0126 for 1h and then were placed in the upper wells. Alternatively,β6 siRNA-treated cells were directly added to the transwell inserts. VEGF was added to the lower wells in a final volume of 1 ml. After 4 hours of incubation, while non-migrating cells were scraped away gently and cells on the lower surface were fixed and stained with Crystal Violet. The migrating cells were counted using a microscope.
Results By using immunoprecipitation analysis, flow cytometry analysis and immunofluorescent staining, integrinαvβ6 was detected at the cell membrane in AGS cells and the positive rate was higher than 90%. AGS cells were treated with VEGF (30ng/ml) after separately pretreatment withβ6 siRNA, with an inhibitor of the ERK signaling pathway (U0126), or with a neutralizing antibody toβ6 (10D5). Expression of integrinαvβ6, P-ERK and MMP-9 at both mRNA and protein levels was measured by means of a real time PCR assay and immunoprecipitation analysis. A significant difference were observed (P<0.05) between VEGF-stimulated and un-stimulated cells. VEGF-induced expression of integrinαvβ6, P-ERK and MMP-9 at both mRNA and protein levels was significantly inhibited byβ6 siRNA, U0126, and 10D5 (P<0.05). Inhibition of the VEGF-induced migration of AGS. AGS cells were separately pretreated withβ6 siRNA, U0126, or 10D5 and were examined for their ability to migrate through a Matrigel filter in the absence/presence of VEGF (30ng/ml). Migration of AGS cells without VEGF or with VEGF stimulation was significantly inhibited byβ6 siRNA, U0126, and 10D5 (P<0.05).
Conclusions Expression ofαvβ6 is detected at the cell membrane in AGS cells. The expression of integrinαvβ6, P-ERK and MMP-9 at both mRNA and protein levels and the invasiveness of AGS cells are stimulated by VEGF, but these effects are significantly inhibited by P6 siRNA, U0126, and 10D5 (P<0.05). These data suggest that VEGF is critical to the invasive process in human gastric cancer and that this occurs via up-regulation of integrin avP6 expression and activation of ERK.
PARTⅢVEGF-C Expression and invasiveness in cholangiocarcinoma
Objective To investigate the expression of VEGF-C in cholangiocarcinoma tissues and the VEGF-C induced invasiveness and metastasis in cholangiocarcinoma cells.
Methods The cholangiocarcinoma tissues were collected from patients requiring clinically indicated surgical resection in QiLu Hospital of Shandong University (Jinan, China), with informed patient consent. Frozen cholangiocarcinoma tissue samples were stored at -80℃until processing. Immunohistochemical analysis was used to detect the expression of VEGF-C in cholangiocarcinoma tissues. And the relationships between the expression of VEGF-C and the clinical-pathological features of cholangiocarcinoma tissues were investigated. The human cholangiocarcinoma cell line FRH-0201 was maintained in RPMI 1640 medium. And the proliferation and invasiveness of FRH-0201 cells were detected by the MTT and Transwell experiment in the absence/presence of VEGF-C.
Results The expression of VEGF-C was detected in 78.2% of cholangio-carcinoma tissues, and the expression was not associated with age, sex and the size of tumor, but associated with lymphatic invasion of the tumors. The proliferation and invasiveness of tumor cells were significantly promoted by VEGF-C (P<0.05).
Conclusions Positive VEGF-C expression in cholangiocarcinoma is linked to significantly enhanced invasiveness, and its value as a prognostic marker is significant for early stage tumors.
恶性肿瘤的侵袭和转移是一个多步骤、多阶段的复杂过程。癌变的细胞和组织会发生一系列的变化,包括细胞转化、肿瘤生长、血管及淋巴管的诱导生成及癌细胞扩散到循环系统；然后在靶器官中粘附、侵袭和增殖。目前已经有数种细胞因子被证实参与肿瘤的转移；其中,血管内皮生长因子(vascular endothelial growth factor, VEGF)不仅参与肿瘤血管的生成,还参与伤口愈合及其他重要的病理过程。VEGF通过两种机制诱导肿瘤血管的生成,一是主要通过其同源性受体VEGFR2,二是通过非VEGFR途径,其中包括整合素的上调和(或)激活。
整合素是一种异二聚体跨膜蛋白,通过VEGF及其受体的协调,能够诱导细胞粘附于细胞外基质(extracellular matrix,ECM)。整合素αvβ6在正常的上皮细胞几乎不表达,而在生长中的胚胎组织及许多上皮性恶性肿瘤中高表达。前期研究中,我们已经证实了整合素αvβ6可用来作为胃癌预后的指标之一。另外αvβ6在结肠癌中的表达可促进基质金属蛋白-9(matrix metalloprotein-9,MMP-9)分泌的增加。在胃癌组织中,MMP-9可降解细胞外基质,从而促进肿瘤细胞的播散和转移。由此可见,αvβ6通过MMP-9降解ECM屏障和介导肿瘤细胞的侵袭性。
有研究证实整合素与VEGF在细胞的粘附、侵袭和转移过程中具有协同作用。整合素α1β1和α2β1可促进VEGF介导的信号转导和内皮细胞的侵袭作用。VEGF与整合素共同参与多发性骨髓瘤细胞的侵袭。在结肠癌细胞中,α9β1阻断抗体可以特异性的抑制VEGF-A诱导的血管生成。然而,整合素αvβ6在VEGF介导的肿瘤细胞侵袭转移过程中的作用尚未明确。
VEGF-C是VEGF家族的新成员,是一种酪氨酸激酶受体,其主要表达在内皮组织的淋巴系统内。在一些恶性肿瘤,如：乳腺癌、胃癌以及肺癌中呈高表达,并且有研究证实VEGF-C的表达与肿瘤的侵袭以及淋巴转移密切相关。但是很少研究VEGF-C在胆管癌中的表达以及与胆管癌侵袭转移之间的关系。
本次研究的目的是研究整合素αvβ6的表达和VEGF的关系,以及它们在体外对胃癌细胞侵袭性的影响；并且研究VEGF-C在胆管癌侵袭转移过程中所起的作用。
第一部分VEGF及整合素αvβ6在胃癌中的表达
目的研究VEGF以及整合素αvβ6在胃癌组织及细胞中的表达情况
方法收集手术切除的胃癌组织标本,冰冻储存于-80℃低温冰箱中。应用免疫组化的方法检测VEGF以及整合素αvβ6在胃癌组织的表达情况。体外培养胃癌AGS细胞,采用流式细胞技术、免疫荧光以及免疫沉淀检测胃癌细胞中整合素的表达情况。
结果30例胃癌标本的免疫组织化学分析显示,其中26例VEGF染色阳性,13例整合素αvβ6染色阳性。其中VEGF以及整合素αvβ6均表达阳性的有10例标本。流式细胞仪检测胃癌AGS细胞中整合素αvβ6呈现高表达,而且阳性率大于90%；细胞免疫荧光结果显示AGS细胞膜中整合素的表达,而应用免疫沉淀技术检查蛋白水平整合素αvβ6的表达情况。
结论VEGF以及整合素αvβ6在胃癌组织中均有表达；整合素αvβ6在胃癌细胞系AGS细胞中呈现高表达。
第二部分VEGF介导的胃癌细胞侵袭作用与整合素αvβ6相关
目的探讨整合素αvβ6在VEGF促进胃癌细胞侵袭、转移能力的作用,寻求控制胃癌转移的靶点。
方法体外培养AGS细胞,外源性VEGF(30ng／ml)刺激24h,应用流式细胞仪、Real-time PCR、Western blot以及免疫沉淀检测整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达变化。
再应用体外培养的AGS细胞,分别用ERK通路阻断剂U0126以及鼠抗人整合素αvβ6单克隆抗体10D5预处理后,外源性VEGF(30ng/ml)刺激24h,收集细胞并检测整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达变化。
针对人整合素基因序列设计其特异性siRNA序列。用荧光标记的阴性对照siRNA筛选转染条件,按照预实验筛选的转染条件将β6 siRNA转染到无血清培养液处理过的AGS细胞中,48h后收集细胞进行抑制效应分析,荧光定量PCR检测β6 mRNA的表达,以无血清培养基及对照siRNA为对照。AGS细胞转染β6 siRNA成功后,采用外源性VEGF刺激,收集细胞检测整合素β6、P-ERK和MMP-9在mRNA以及蛋白水平的表达。趋化小室试验检测外源性VEGF介导的AGS细胞侵袭能力。β6 siRNA转染的AGS细胞,以及分别用ERK通路阻断剂U0126以及鼠抗人整合素αvβ6单克隆抗体10D5预处理的AGS细胞,加入包被好的小室上层,在有无外源性VEGF介导的情况下,观察转移到小室下层的细胞个数,从而反应其不同条件下的侵袭能力。
结果应用免疫沉淀、流式细胞仪以及细胞免疫荧光,可以发现体外培养的AGS细胞内有整合素αvβ6的表达,其主要表达在细胞膜,而且表达的阳性率在90%以上。AGS细胞在外源性VEGF(30ng/ml)刺激24h后,整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达明显增加,与对照组相比具有统计学意义。而应用ERK通路阻断剂U0126以及鼠抗人整合素αvβ6单克隆抗体10D5预处理后,外源性VEGF(30ng/ml)刺激24h,整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达显著被抑制,与对照组相比具有统计学意义。荧光显微镜观察见筛选后的转染条件下荧光标记siRNA的转染效率较高,超过90%。在β6 siRNA转染的无血清培养液处理过的AGS细胞中,在有无外源性VEGF作用的条件下,整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达得到相同的结论。趋化小室实验检测到在外源性VEGF介导的情况下,转移至小室下层的AGS细胞数明显增加。而应用ERK通路阻断剂U0126、鼠抗人整合素αvβ6单克隆抗体10D5以及β6 siRNA转染预处理后,转移至小室下层的AGS细胞数显著减少,与对照组相比具有统计学意义。
结论胃癌AGS细胞内有整合素αvβ6的表达；外源性VEGF (30ng/ml)作用于AGS细胞后,可以刺激整合素αvβ6、P-ERK和MMP-9在mRNA以及蛋白水平的表达显著增加以及提高AGS细胞的侵袭能力,而且这种作用可以被ERK通路阻断剂U0126、鼠抗人整合素αvβ6单克隆抗体10D5以及β6 siRNA转染预处理显著抑制,与对照组相比具有统计学意义。这说明整合素αvβ6参与了VEGF介导的胃癌侵袭转移作用,并且ERK通路在此过程中起着一定的作用。
第三部分VEGF-C在胆管癌中的表达及侵袭转移中的作用研究
目的研究VEGF-C在胆管癌癌组织中的表达情况及促进胆管癌癌细胞侵袭、转移能力的作用。
方法收集手术切除的胆管癌组织标本,冰冻储存于-80℃低温冰箱中。应用免疫组化的方法检测VEGF-C在胆管癌组织的表达情况,回顾性分析这些病人的临床病例资料,并与免疫组化结果相结合,分析VEGF-C的表达与患者临床资料之间的关系；体外培养胆管癌FRH-0201细胞,并且在有无外源性VEGF-C作用的情况下,采用增殖实验以及趋化小室实验检测胆管癌细胞FRH-0201的侵袭转移能力。
结果VEGF-C在胆管癌组织中呈高表达,表达率约为75.4%；其表达与病人的年龄、性别以及肿瘤大小、肿瘤的临床分期无明显关系,而与肿瘤的淋巴结转移明显相关。胆管癌细胞在外源性VEGF-C的作用的情况下,胆管癌细胞的增殖以及侵袭能力明显被增强,与对照组相比具有统计学意义。
结论VEGF-C在胆管癌组织中呈高表达并且在其侵袭转移过程中起着一定的作用。
Background and signification
Tumor metastasis is a complex and multi-stage process. A number of cellular and tissue changes occur as a cancer spreads, including cellular transformation and tumor growth, angiogenesis and lymphangiogenesis, and release of cancer cells into the circulation; then attachment, invasion, and proliferation within the new target organ. Several cytokines are now known to be involved in the process of tumor metastasis. Among these is vascular endothelial growth factor (VEGF), a factor that is associated not only with the angiogenesis of cancers but also with those involved in wound healing and other important pathologies. There are two mechanisms involved in VEGF-mediated angiogenesis; one is through its cognate receptor VEGFR2, the other is via up-regulation of the expression and/or activation of integrins.
Integrins are heterodimeric transmembrane proteins that can induce cell adhesion to the extracellular matrix (ECM), coordinated by VEGF and its receptors. The avP6 integrin is restricted to epithelial cells and is highly expressed in developing fetal tissues as well as in a number of epithelial carcinomas and cancers. In our previous studies, we demonstrated that integrin avP6 can be used as a prognostic indicator of gastric carcinoma. In addition,αvβ6 expression in colon cancer cells leads to the increase in secretion of matrix metalloprotein-9 (MMP-9). In gastric cancer, degradation of ECM barriers by MMP-9 is important in facilitating tumor cellular dissemination and metastasis. Thus, integrinαvβ6 appears to have roles in promoting ECM degradation and in mediating tumor cell invasiveness via MMP-9.
There are several confirmed collaborative relationships between integrins and VEGF in mediation of cell adhesion, migration, and proliferation. Theα1β1 andα2β2 integrins support VEGF-stimulated signal transduction and endothelial cell migration. Migration of multiple myeloma cells is induced by VEGF and associated withβ1 integrin. In colon cancer cells,α9β1-blocking antibody has been demonstrated to specifically inhibit the angiogenesis induced by VEGF-A. However, a role for integrinαvβ6 in VEGF-induced tumor progression has not yet been established.
VEGF-C is a new member of the VEGF family, a tyrosine kinase receptor that is predominantly expressed in the endothelium of lymphatic vessels. The expression of VEGF-C is high in several types of human malignant tumors, including breast, lung, and gastric carcinomas and some investigators have shown that VEGF-C expression was closely associated with tumor invasion and lymph node metastasis. However, there have been very few reports on the correlation between VEGF-C gene expression and invasive phenotype in cholangiocarcinoma.
The goal of the present study was therefore to investigate the relationship between VEGF and expression of integrinαvβ6 and their effects on invasiveness of gastric carcinoma cells in vitro. And the effect of VEGF-C in invasiveness and metastasis of cholangiocarcinoma.
PARTⅠExpression of VEGF and integrinαvβ6 in gastric carcinoma tissues and cells
Objective To study the expression of VEGF and integrinαvβ6 in gastric carcinoma tissues and cells.
Methods Thirty gastric cancer tissues were collected from patients requiring clinically indicated surgical resection in QiLu Hospital of Shandong University (Jinan, China), with informed patient consent. Frozen gastric cancer tissue samples were stored at -80℃until processing. Immunohistochemical analysis was used to detect the expression of VEGF and integrinαvβ6 in gastric carcinoma tissues. The human gastric cancer cell line AGS was maintained in RPMI 1640 medium supplemented with 10% fetal bovine serum (FBS), 100U/ml penicillin, 10μg/ml streptomycin, and 2mM L-glutamine. Immuno-precipitation, FAC scanning and immunofluorescence staining were used to confirme the presence ofαvβ6 in gastric cancer AGS cells.
Results Immunohistochemical analysis of tumor samples from 30 patients revealed that 26/30 samples stained positively for VEGF and 13/30 samples stained positively for integrinαvβ6. In 10/30 patients tumors were positive for both VEGF and integrinαvβ6. High surface expression ofαvβ6 integrin was detected by flow cytometry analysis, and the positive rate was higher than 90%. Expression ofαvβ6 was detected with immunofluorescent staining at the cell membrane in AGS cells. AGS cells were examined by immunoprecipitation analysis using antibody of 10D5 for expression ofαvβ6 integrin at protein levels.
Conclusions VEGF and integrinαvβ6 are all expressed in gastric carcinoma tissues. And the high surface expression ofαvβ6 integrin is detected in gastric cancer AGS cells.
PARTⅡVEGF enhances gastric carcinoma invasiveness via integrinαvβ6
Objective To investigate the effect of integrinαvβ6 on VEGF regulated invasiveness and metastasis in gastric carcinoma cells, to probe into the key point of regulating metastasis of gastric carcinoma.
Methods The AGS cells were starved for 4h in medium without fetal bovine serum and then stimulated with VEGF (30ng/ml) for 24h at 37℃. The expression at both mRNA and protein levels was also observed for integrinαvβ6, P-ERK and MMP-9 by using the real-time PCR, western blot analysis and immunoprecipitation.
The roles ofαvβ6, P-ERK and MMP-9 at both mRNA and protein levels were detected by using monoclonal antibody for the humanαvβ6 integrin (10D5) and an inhibitor of the ERK signaling pathway (U0126).
Specific P6 siRNA was designed according to the cDNA sequence of integrin subunit beta6 and the BLAST search was used to avoid unintentional silencing and ensure the uniqueness of target. Cells were grown subconfluently onto 24-well plates and transfected with 1μg ofβ6 siRNA, with nonspecific siRNA as control. After 48h of transfection, cells were washed and analyzed further as specifically indicated. The VEGF-stimulated transfected AGS cells were detected to calculate the change of integrinαvβ6, P-ERK and MMP-9 at both mRNA and protein levels.
Migration of AGS cells through Matrigel-coated filters was examined in 24-well transwell inserts. After trypsinization, 1×105 cells in 100μl were incubated with 10D5 or U0126 for 1h and then were placed in the upper wells. Alternatively,β6 siRNA-treated cells were directly added to the transwell inserts. VEGF was added to the lower wells in a final volume of 1 ml. After 4 hours of incubation, while non-migrating cells were scraped away gently and cells on the lower surface were fixed and stained with Crystal Violet. The migrating cells were counted using a microscope.
Results By using immunoprecipitation analysis, flow cytometry analysis and immunofluorescent staining, integrinαvβ6 was detected at the cell membrane in AGS cells and the positive rate was higher than 90%. AGS cells were treated with VEGF (30ng/ml) after separately pretreatment withβ6 siRNA, with an inhibitor of the ERK signaling pathway (U0126), or with a neutralizing antibody toβ6 (10D5). Expression of integrinαvβ6, P-ERK and MMP-9 at both mRNA and protein levels was measured by means of a real time PCR assay and immunoprecipitation analysis. A significant difference were observed (P<0.05) between VEGF-stimulated and un-stimulated cells. VEGF-induced expression of integrinαvβ6, P-ERK and MMP-9 at both mRNA and protein levels was significantly inhibited byβ6 siRNA, U0126, and 10D5 (P<0.05). Inhibition of the VEGF-induced migration of AGS. AGS cells were separately pretreated withβ6 siRNA, U0126, or 10D5 and were examined for their ability to migrate through a Matrigel filter in the absence/presence of VEGF (30ng/ml). Migration of AGS cells without VEGF or with VEGF stimulation was significantly inhibited byβ6 siRNA, U0126, and 10D5 (P<0.05).
Conclusions Expression ofαvβ6 is detected at the cell membrane in AGS cells. The expression of integrinαvβ6, P-ERK and MMP-9 at both mRNA and protein levels and the invasiveness of AGS cells are stimulated by VEGF, but these effects are significantly inhibited by P6 siRNA, U0126, and 10D5 (P<0.05). These data suggest that VEGF is critical to the invasive process in human gastric cancer and that this occurs via up-regulation of integrin avP6 expression and activation of ERK.
PARTⅢVEGF-C Expression and invasiveness in cholangiocarcinoma
Objective To investigate the expression of VEGF-C in cholangiocarcinoma tissues and the VEGF-C induced invasiveness and metastasis in cholangiocarcinoma cells.
Methods The cholangiocarcinoma tissues were collected from patients requiring clinically indicated surgical resection in QiLu Hospital of Shandong University (Jinan, China), with informed patient consent. Frozen cholangiocarcinoma tissue samples were stored at -80℃until processing. Immunohistochemical analysis was used to detect the expression of VEGF-C in cholangiocarcinoma tissues. And the relationships between the expression of VEGF-C and the clinical-pathological features of cholangiocarcinoma tissues were investigated. The human cholangiocarcinoma cell line FRH-0201 was maintained in RPMI 1640 medium. And the proliferation and invasiveness of FRH-0201 cells were detected by the MTT and Transwell experiment in the absence/presence of VEGF-C.
Results The expression of VEGF-C was detected in 78.2% of cholangio-carcinoma tissues, and the expression was not associated with age, sex and the size of tumor, but associated with lymphatic invasion of the tumors. The proliferation and invasiveness of tumor cells were significantly promoted by VEGF-C (P<0.05).
Conclusions Positive VEGF-C expression in cholangiocarcinoma is linked to significantly enhanced invasiveness, and its value as a prognostic marker is significant for early stage tumors.
引文
[1]Ferlay J, Bray F, Pisani P, Parkin DM. GLOBOCAN 2002 cancer incidence, mortality and prevalence worldwide. IARC cancerbase no.5, version 2.0. Lyon: IARC Press; 2004.
[2]Correa P, Chen VW. Gastric cancer. Cancer Surv.1994; 19(20):55-76.
[3]Aragones N, Pollan M, Rodero I, Lopez-Abente G. Gastric cancer inthe European Union (1968-1992):mortality trends and cohort effect. Ann Epidemiol.1997; 7: 294-303.
[4]Ferretti S, Gafa L. Upper gastrointestinal tract cancers:oesophagus, stomach, liver, gallbladder and biliary ducts, pancreas. Epidemiol Prev.2004; 28:34-42.
[5]Kelley JR, Duggan JM. Gastric cancer epidemiology and risk factors. J Clin Epidemiol.2003; 56:1-9.
[6]Devesa SS, Blot WJ, Fraumeni Jr JF. Changing patterns in the incidence of esophageal and gastric carcinoma in the United States. Cancer.1998; 83:2049-2053.
[7]Powell J, McConkey CC. Increasing incidence of adenocarcinoma of the gastric cardia and adjacent sites. Br J Cancer.1990; 62:440-443.
[8]Van't Veer LJ, Weigelt B. Road map to metastasis. Nat. Med.2003; 9:999-1000.
[9]Hanahan D, Weinberg RA. The hallmarks of cancer. Cell.2000; 100:57-70.
[10]Ferrara N. The role of vascular endothelial growth factor in pathological angio-genesis. Breast Cancer Res. Treat.1995; 36:127-137.
[11]Dvorak HF, Brown LF, Detmar M, Dvorak AM. Vascular permeability factor/vascular endothelial growth factor, microvas-cular hyperpermeability, and angiogenesis. Am. J. Pathol.1995; 146:1029-1039.
[12]Hood JD, Frausto R, Kiosses WB, Schwartz MA, Cheresh DA. Differential alphav integrin-mediated Ras-ERK signaling during twopathways of angiogene-sis, J Cell Biol.2003; 162:933-943.
[13]Smyth SS, Patterson C. Tiny dancers:the integrin-growth factor nexus in angiogenic signaling. J Cell Biol.2002; 158:17-21.
[14]Schietroma C, Cianfarani F, Lacal PM,et al. Vascular endothelial growth factor-C expression correlates with lymph node localizat-ion of human melanoma metastases. Cancer.2003; 98(4):789-797.
[15]Hung CJ, Ginzinger DG, Zarnegar R, et al. Expression of vascular endothelial growth factor-C in benign and malignant thyroid tumors. J Clin Endocrinol Metab.2003; 88(8):3694-3699.
[16]Liu F, Zhang YJ. Roles of VEGF-C and its receptor Flt-4 in proliferation and metastasis of primary breast cancer. Ai Zheng.2003; 22(10):1053-1056.
[17]Kimura Y, Watanabe M, Ohga T, et al. Vascular endothelial growth factor-C expression correlates with lymphatic involvement and poor prognosis in patients with esophageal squamous cell carcinoma. Oncol Rep.2003; 10(6):1747-1751.
[18]Maeda K, Yashiro M, Nishihara T, et al. Correlation between vascular endothelial growth factor-C expression and lymph node metastasis in T1 carcinoma of the colon and rectum. Surg Today.2003; 33(10):736-739.
[19]Tamura M, Ohta Y. Serum vascular endothelial growth factor-C level in patients with primary nonsmall cell lung carcinoma:a possible diagnostic tool for lymph node metastasis. Cancer.2003; 98(6):1217-1222.
[20]Sedivy R, Mannagetta J, Haverkampf C, et al. Expression of vascular endothelial growth factor-C correlates with the lymphatic microvessel density and the nodal status in oral squamous cell cancer. J Oral Pathol Med.2003; 32(8):455-460.
[21]Futagami S, Tatsuguchi A, Hiratsuka T, et al. Monocyte chemoat-tractant protein-1 and CD40 ligation have a synergistic effect on vascular endothelial growth factor production through cyclooxy-genase upregulation in gastric cancer. J Gastroenterol.2008; 43(3):216-224.
[22]Sheppard D, Rozzo C, Starr L, Quaranta V, Erle DJ, Pytela R. Complete amino acid sequence of a novel integrin beta subunit (beta6) identified in epithelial cells using the polymerase chain reaction. J Biol Chem.1990; 265:11502-11507.
[23]Breuss JM, Gallo J, DeLisser HM, Klimanskaya IV, Folkesson HG, Pittet JF, Nishimura SL, Aldape K, Landers DV, Carpenter W, et al. Expression of the beta6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling. J Cell Sci.1995; 108:2241-2251.
[24]Zhang ZY, Xu KS, He QS, Niu WB, Wang JY, Mi YT, Wang JS, Wang GQ, Yang GY, Niu J. Signaling and regulatory mechanisms of integrin alphavbeta6 on the apoptosis of colon cancer cells. Cancer Lett.2008; 266:209-215.
[25]Mattern J, Koomagi R, Volm M. Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumor cell proliferation in human epidermoid lung carcinoma. Br J Cancer.1996; 73:931-934.
[26]吴胤瑛,李恩孝.VEGF在恶性肿瘤中的研究进展.现代肿瘤医学.2005,13(5)：18-21.
[27]Carmeliet P, Jain RK. Angiogenesis in cancer and other diseases. Nature.2000, 407(6801):249-257.
[28]Poon RT, Fan ST, Wong J. Clinical implications of circulating angiogenic factors in cancer patients. J Clin Oncol.2001,19(4):1207-1225.
[29]Frish SM, Ruoslahti E.Integrin and anoikis. Curr Opin Cell Biol.1997; 9:701-706.
[30]Frish SM, Screaton RA. Anoikis mechanisms. Curr Opin Cell Biol.2001; 13:555-562.
[31]Hynes RO. Integrins:bidirectional, allosteric signaling machines. Cell.2002; 110: 673-687.
[32]Hynes RO. Integrins:versatility, modulation and signaling in cell ahhesion. Cell.1992; 69:11-25.
[33]Akiyama SK, Olden K, Yamada KM. Fibronectin and integrins in invasion and metastasis. Cancer Metastasis Rev.1995; 14:173-189.
[34]Ferrara N, Henzel WJ. Pituitary follicular cells secrete a novel heparin-binding growth factor specific for vascular endothelial cells. Biochem Biophys Res Commun.1989,15;161(2):851-858.
[35]Petrova TV, Makinen T, Alitalo K. Signaling via vascular endothe-lial growth factor receptors. Exp Cell Res.1999; 253(1):117-130.
[36]Sasaki T, Tanno S, Shibukawa K, Osanai S, Kawabe J, Ohsaki Y. Administration of VEGF receptor tyrosine kinase inhibitor increases VEGF production causing angiogenesis in human small-cell lung cancer xenografts. Int J Oncol.2008 Sep; 33(3):525-532.
[37]Los M, Roodhart JM, Voest EE. Target practice:lessons from phase III trials with bevacizumab and vatalanib in the treatment of advanced colorectal cancer. Oncologist.2007; 12:443-450
[38]Marbeuf-Gueye C, Salerno M, Quidu P, Garnier-Suillerot A. Inhibition of the P-glycoprotein and multidrug resistance protein-mediated efflux of anthracyclines and calceinacetoxymethyl ester by PAK-104P. Eur J Pharmacol. 2000 Mar 17; 391(3):207-216.
[39]Motoji T, Motomura S, Wang YH. Multidrug resistance of acute leukemia and a strategy to overcome it. Int J Hematol.2000 Dec; 72(4):418-424.
[40]Poirson-Bichat F, Goncalves RA, Miccoli L, Dutrillaux B, Poupon MF. Methionine depletion enhances the antitumoral efficacy of cytotoxic agents in drug-resistant human tumor xenografts. Clin Cancer Res.2000 Feb; 6(2):643-653.
[41]Mistry P, Stewart AJ, Dangerfield W, et al. In vitro and in vivo characterization of XR11576, a novel, orally active, dual inhibitor of topoisomerase Ⅰ and Ⅱ. Anticancer Drugs.2002 Jan; 13(1):15-28.
[42]Van Hattum AH, Pinedo HM, Schluper HM, Hausheer FH, Boven E. New highly lipophilic camptothecin BNP1350 is an effective drug in experimental human cancer. Int J Cancer.2000 Oct 15; 88(2):260-266.
[43]Giancotti FG, Ruoslahti E.Integrin signaling. Science.1999 Aug 13; 285 (5430): 1028-1032.
[44]Crowe DL, Shuler CF. Regulation of tumor cell invasion by extrac ellular matrix. Histol Histopathol.1999 Apr; 14(2):665-671.
[45]De Melker AA, Sonnenberg A. Integrins:alternative splicing as a mechanism to regulate ligand binding and integrin signaling events. Bioessays.1999 Jun; 21(6): 499-509.
[46]Hynes RO. Integrins:versatility, modulation and spreading in cell adhesion. Cell.1992; 69:11-25.
[47]Albelad SM. Biology of disease:role of integrins and other cell adhesion molecules in tumor progression and metastasis. Lab Invest.1993; 68:4-17.
[48]Cheresh D. Integrins structure, function and biological properties. Adv Mol Cell Biol.1993; 6:225-228.
[49]Albelda SM, Buck CA. Integrins and other cell adhesion molecules. FASEB J. 1990; 4:2868-2880.
[50]Liotta LA, Rao CN. Tumor invasion and the extracellular matrix. Lab Invest. 1983;49(6):636-649.
[51]郑世曦,陈汉琴,杨军等.层粘连蛋白在人原发性肝细胞癌转移中的意义.华人消化杂志.1998；6：507-509.
[52]Meredith JE Jr, Winitz S, Lewis JM, Hess S, Ren XD, Renshaw MW, Schwartz MA. The regulation of growth and intracellular signaling by integrins. Endocr Rev.1996 Jun; 17(3):207-220.
[53]Sjaastad MD, Nelson WJ. Integrin-mediated calcium signaling and regulation of cell adhesion by intracellular calcium. Bioessays.1997 Jan; 19(1):47-55.
[54]Shimizu Y, Hunt SW. Regulating integrin-mediated adhesion:one more function for PI 3-kinase? Immunol Today.1996 Dec; 17(12):565-573.
[55]Hanks SK, Polte TR. Signaling through focal adhesion kinase. Bioessays.1997 Feb; 19(2):137-145.
[56]Hannigan GE, Dedhar S. Protein kinase mediators of integrin signal transduction. J Mol Med.1997 Jan; 75(1):35-44.
[57]Dedhar S, Williams B, Hannigan G. Integrin-linked kinase (ILK):a regulator of integrin and growth-factor signalling. Trends Cell Biol.1999 Aug; 9(8):319-323.
[58]Huang Y, Wu C. Integrin-linked kinase and associated proteins (review). Int J Mol Med.1999 Jun; 3(6):563-572.
[59]Petty HR, Todd RF. Integrins as promiscuous signal transduction devices. Immunol Today.1996 May; 17(5):209-212.
[60]Todd RF, Petty HR. Beta2 (CD 11/CD 18) integrins can serve as signaling partners for other leukocyte receptors. J Lab Clin Med.1997 May; 129(5):492- 498.
[61]Paulus W, Baur I, Schuppan D et al. Characterization of integrin receptors in normal and neoplastic human brain. Am J Pathol.1993; 143(1):154-163.
[62]Mrrjans F, Meyer T, Frrrschen C et al. In vivo therapy of malignant melanoma by means of antagonists of αv integrins. Int J Cancer.2000; 87:716-723.
[63]Liapis H, Adler LM, Wick MR et al. Expression of alpha(v)beta3 integrin is less frequent in ovarian epithelial tumors of low malignant potential in contrast to ovarian carcinomas. Hum Pathol.1997; 28(4):443-449.
[64]Rhim JS, Tsai WP, Chen ZQ et al. A human vascular endothelial cell model to study angiogenesis and tumorigenesis. Carcinogene- sis.1998; 19(4):673-681.
[65]Erdreich EA, Shimada H, Groshen S et al. Integrins alpha(v)beta3 and alpha(v) beta5 are expressed by endothelium of highrisk neuroblastoma and their inhibition is associated with increased endogenous ceramide. Cancer Res.2000; 60(3):712-721.
[66]Takano S, Tsuboi K, Tomono Y et al. Tissue factor, osteopontin, alphavbeta3 integrin expression in microvasculature of glioma associated with vascular endothelial growth factor expression. Br J Cancer.2000; 82(12):1967-1973.
[67]Max R, Gerritsen RR, Nooijen PT et al. Immunohistochemical analysis of integrin alphavbeta3 expression on tumor-associated vessels of human carcinomas. Int J Cancer.1997; 71(3):320-324.
[68]Pozzi A, Moberg PE, Miles LA et al. Elevated matrix metallo- protease and angiostatin levels in integrin alpha1 knockout mice cause reduced tumor vascularization. Proc Natl Acad Sci USA.2000; 97(5):22022207.
[69]Kim S, Bell K, Mousa SA et al. Regulation of angiogenesis in vivo by ligation of integrin alpha5betal with the central cell-binding domain of fibronectin. Am J Pathol.2000; 156:1345-1362.
[70]Breuss JM, Sheppard D. Expression of the β6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling. J Cell Sci. 1995; 108:2241-2251.
[71]Xiaowu Li, Brian L. Schmidt, Amha Atakilit, Bing Chen, Duncan Ellis and Daniel M. αvβ6-Fyn Signaling Promotes Oral Cancer Progression. J Biol Chem. 2003; 278:(43) 41646-41653.
[72]Agrez MV, Chen A, Cone RI, Pytela R and Sheppard D. The αvβ6 integrin promotes proliferation of colon carcinoma cells through a unique region of the β6 cytoplasmic domain. J Cell Biol.1994; 127:547-556.
[73]吴小华,李冬秀,李利等.整合素对顺铂诱导的卵巢癌细胞凋亡的影响.中华妇产科杂志.2004；39(2)：112-115.
[74]Arihiro K, Kaneko M, Fujii S, Inai K and Yokosaki Y. Significance of alpha9betal and alphavbeta6 integrin expression in breast carcinoma. Breast Cancer.2000;7:19-26.
[75]Zhang ZY, Xu KS, Wang JS, et al. Integrin alphanvbeta6 acts as a prognostic indicator in gastric carcinoma. Clin Oncol (R Coll Radiol).2008 Feb; 20(1):61-66.
[1]Allum WH, Powell DJ, McConkey CC, Fielding JW. Gastric cancer:a 25-year review. Br J Surg.1989; 76:535-540.
[2]Wanebo HJ, Kennedy BJ, Chmiel J, et al. Cancer of the stomach. A patient care study by the American College of Surgeons. Ann Surg.1993; 218:583-92.
[3]Akoh JA, Macintyre IM. Improving survival in gastric cancer:review of 5-year survival rates in English language publications from 1970. Br J Surg.1992; 79:293-9.
[4]Hermanek P, Maruyama K, Sobin LH. Gastric cancer. In:Hermanek P, Gospodarowicz MK, Henson DE, Hutter RVP, Sobin LH, editors. Prognostic factors in cancer. Geneve:International Union Against Cancer (UICC); 1995
[5]Wartenberg M, Ling FC, Schallenberg M, et al. Down-regulation of intrinsic P-glycoprotein expression in multicellular prostate tumor spheroids by reactive oxygen species. J Biol Chem.2001; 276 (20):17420-17428.
[6]Ferrara N. The role of vascular endothelial growth factor in pathological angiogenesis. Breast Cancer Res. Treat.1995; 36:127-137.
[7]Dvorak HF, Brown LF, Detmar M, Dvorak AM. Vascular permeability factor/vascular endothelial growth factor, microvas- cular hyperpermeability, and angiogenesis. Am. J. Pathol.1995; 146:1029-1039.
[8]Hood JD, Frausto R, Kiosses WB, Schwartz MA, Cheresh DA. Differential alphav integrin-mediated Ras-ERK signaling during twopathways of angiogenesis. J Cell Biol.2003; 162:933-943.
[9]Smyth SS, Patterson C. Tiny dancers:the integrin-growth factor nexus in angiogenic signaling. J Cell Biol.2002; 158:17-21.
[10]Senger DR, Perruzzi CA, Streit M, Koteliansky VE, Fougerolles AR, Detmar M. The alpha(1)beta(1) and alpha(2)beta(1) integrins provide critical support for vascular endothelial growth factor signaling, endothelial cell migration, and tumor angiogenesis. Am J Pathol.2002; 160:195-204.
[11]Eliceiri BP, Puente XS, Hood JD, Stupack DG, Schlaepfer DD, Huang XZ, Sheppard D, Cheresh DA. Src-mediated coupling of focal adhesion kinase to integrin alpha(v)beta5 in vascular endothe- lial growth factor signaling. J Cell Biol.2002;157:149-159.
[12]Lee TH, Seng S, Li H, Kennel SJ, Avraham HK, Avraham S. Integrin regulation by vascular endothelial growth factor in human brain microvascular endothelial cells:role of alpha6betal integrin in angiogenesis. J Microbiol.2006; 44:502-507.
[13]Sheppard D, Rozzo C, Starr L, Quaranta V, Erle DJ, Pytela R. Complete amino acid sequence of a novel integrin beta subunit (beta6) identified in epithelial cells using the polymerase chain reaction. J Biol Chem.1990; 265:11502-11507.
[14]Breuss JM, Gallo J, DeLisser HM, Klimanskaya IV, Folkesson HG, Pittet JF, Nishimura SL, Aldape K, Landers DV, Carpenter W, et al. Expression of the beta6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling. J Cell Sci.1995; 108:2241-2251.
[15]Zhang ZY, Xu KS, He QS, Niu WB, Wang JY, Mi YT, Wang JS, Wang GQ, Yang GY, Niu J. Signaling and regulatory mechanisms of integrin alphavbeta6 on the apoptosis of colon cancer cells, Cancer Lett.2008; 266:209-215.
[16]Gu X, Niu J, Dorahy DJ, Scott R, Agrez MV. Integrin alpha(v) beta6-associated ERK2 mediates MMP-9 secretion in colon cancer cells. Brit J Cancer.2002;87: 348-351.
[17]Hynes, RO. Integrins:versatility, modulation and signaling in cell adhesion. Cell. 1992; 69:11-25.
[18]Sawhney RS, Sharma B, Humphrey LE, Brattain MG. Integrin alpha2 and extracellular signal-regulated kinase are functionally linked in highly malignant autocrine transforming growth factor-alpha-driven colon cancer cells. J Biol Chem.2003; 278(22):19861-19869.
[19]吴小华,李冬秀,李利等.整合素对顺铂诱导的卵巢癌细胞凋亡的影响.中华妇产科杂志.2004；39(2)：112-115.
[20]Arihiro K, Kaneko M, Fujii S, Inai K and Yokosaki Y. Significance of alpha9betal and alphavbeta6 integrin expression in breast carcinoma. Breast Cancer.2000; 7:19-26.
[21]Regezi JA, Ramos DM, Pytela R, Dekker NP and Jordan RCK. Tenascin and β6 integrin are overexpressed in floor of mouth in situ carcinomas and invasive squamous cell carcinomas. Oral Oncol.2002; 38:332-336.
[22]Ahmed N, Niu J, Dorahy DJ, Gu X, Andrews S, Meldrum CJ, Scott RJ, Baker MS, Macreadie IG, Agrez MV. Direct integrin alphavbeta6-ERK binding: implications for tumour growth. Oncogene.2002 Feb 21; 21(9):1370-1380.
[23]Yang GY, Xu KS, Pan ZQ, Zhang ZY, Mi YT, Wang JS, Chen R, Niu J. Integrin alphavbeta6 mediates the potential for colon cancer cells to colonize in and metastasize to the liver. Cancer Sci.2008 May; 99(5):879-887
[24]张朝阳,徐克森,杨广运等.反义整合素β6基因对结肠癌细胞作用的研究.中华医学杂志.2007；87(37)：2645-2649.
[25]Zhang ZY, Xu KS, Wang JS, Yang GY, Wang W, Wang JY, Niu WB, Liu EY, Mi YT, Niu J. Integrin alphanubeta6 Acts as a Prognostic Indicator in Gastric Carcinoma. Clin Oncol (R Coll Radiol).2008 Feb; 20(1):61-66.
[26]Xiaowu Li, Brian L. Schmidt, Amha Atakilit, Bing Chen, Duncan Ellis and Daniel M. Ramos. αvβ6-Fyn Signaling Promotes Oral Cancer Progression. J Biol Chem.2003; 278:(43) 41646-41653.
[27]Brockbank EC, Bridges J, Marshall CJ, Sahai E. Integrin betal is required for the invasive behaviour but not proliferation of squam-ous cell carcinoma cells in vivo. Br J Cancer.2005,17; 92(1):102-112.
[28]Niu J, share 1st author with Ahmed N, Dorahy DJ, Gu X, Andrews S, Meldrum CJ, Scott RJ, Baker MS, Macreadie IG, Agrez MV. Direct integrin alphavbeta 6-ERK binding:implicate- ons for tumour growth. Oncogene. 2002 Feb 21; 21(9):1370-1380.
[29]J. Wang, Z. Zhang, K. Xu, X. Sun, G. Yang, W. Niu, E. Liu, C. Peng, P. Lin, J. Wang, R. Chen, M. Agrez, J. Niu. Suppression of integrin alphaupsilonbeta6 by RNA interference in colon cancer cells inhibits extracellular matrix degradation through the MAPK pathway. Int J Cancer.2008; 123:1311-1317.
[30]Kabashima A, Maehara Y, Kakeji Y, Baba H, Koga T, Sugimachi K. Clinicopathological features and overexpression of matrix metallo- proteinases in intramucosal gastric carcinoma with lymph node metastasis. Clin Cancer Res. 2000; 9:3581-3584.
[31]Torii A, Kodera Y, Uesaka K, Hirai T, Yasui K, Morimoto T, Yamamura Y, Kato T, Hayakawa T, Fujimoto N, Kito T, Plasma concentration of matrix metalloproteinase 9 in gastric cancer. Brit J Surg.1997; 84:133-136.
[32]Petrova TV, Makinen T, Alitalo K. Signaling via vascular endothelial growth factor receptors. Exp Cell Res.1999; 253(1):117-130.
[33]刘杰,孙正义,曹蕾.血管内皮生长因子对生物衍生骨复合骨髓基质细胞体内成骨的影响.中国临床康复.2003；7(11)：1622-1623.
[34]Shibuya M. Vascular endothelial growth factor receptor-2:its unique signaling and specific ligand, VEGF-E. Cancer Sci.2003; 94:751-756.
[35]Shinkai A, Ito M, Anazawa H, Yamaguchi S, Shitara K, Shibuya M. Mapping of the sites involved in ligand association and dissociation at the extracellular domain of the kinase insert domaincontaining domaincontaining receptor for
vascular endothelial growth factor. J Biol Chem.1998; 273:31283-31288.
[36]Li B, Fuh G, Meng G, Xin X, Gerritsen ME, Cunningham B, de Vos AM. Receptor-selective variants of human vascular endothelial growth factor. Generation and characterization. J Biol Chem.2000; 275:29823-29828
[37]Gille H, Kowalski J, Li B, LeCouter J, Moffat B, Zioncheck TF, Pelletier N, Ferrara N. Analysis of biological effects and signaling properties of Flt-1 (VEGFR-1) and KDR (VEGFR-2). A reassessment using novel receptor-specific vascular endothelial growth factor mutants. J Biol Chem.2001; 276:3222-3230
[38]Larrivee B, Karsan A. Signaling pathways induced by vascular endothelial growth factor. Int J Mol Med.2000; 5:447-456.
[39]Brekken RA, Overholser JP, Stastny VA, el al. Selective inhibition of vascula endothelial growth factor(VEGF) receptor2(KDR/FIk-1) activity by a monocle-nal anti-VEGF antibody blocks tumor growth in mice. Cancer Res.2000; 60(18): 5117-5124.
[40]Pasqualini R, Koivunen E, Ruoslahti E. av Integrins as receptors for tumor targeting by circulating ligands. Nat Biotech.1997; 15:542-546.
[41]Varner JA. The role of vascular cell integrins αvβ3 and αvβ5 in angiogenesis. In: Goldberg ID, Rosen EM. Regulation of angiogenesis. Basel:Brkhauser Vrlag. 1997; 361-390.
[42]Shen JR, Yen MH, Kan YC et al. Inhibition of angiogenesis in vitro and in vivo: comparision of the relative activities of triflavin, an Arg-Gly-Asp-containing peptide and anti-alpha(v)beta3 integrin monoclonal antibody. Biochem Biophys Acta.1997; 1336(3):445-454.
[43]Herskowitz I. MAP kinase pathways in yeast:for mating and more. Cell.1995; 80(2):187-197.
[44]Widmann C, Gibson S, Jarpe M.B, et al. Mitogen-activated protein kinase: conservation of a three-kinase module from yeast to human. Physiol Rev.1999; 79(1):143-180.
[45]Chen Z, Gibson TB, Robinson F, et al. MAP kinases. Chem Rev.2001; 101(8):2449-2476.
[46]Westermarck J, Kahari VM. Regulation of matrix metalloproteinase expression in tumor invasion. FASEB J.1999 May; 13(8):781-792.
[47]EgebladM, Werb Z. New functions for the matrix metalloproteina-ses in cancer progression. Nature Rev Cancer.2002; 2(3):161-174.
[48]Johansson N, Ala-aho R, Uitto V, et al. Expression of collagenase-3 (MMP-13) and collagenase-1(MMP-1) by transformed keratinocy- tes is dependent on the activity of p38 mitogen-activated protein kinase. J Cell Sci.2000 Jan;113 Pt 2:227-235.
[49]Simon C, Hicks MJ, Nemechek AJ, Mehta R, O'Malley BW Jr, Goepfert H, Flaitz CM, Boyd D. PD 098059, an inhibitor of ERK1 activation, attenuates the in vivo invasiveness of head and neck squamous cell carcinoma. Br J Cancer. 1999 Jul; 80(9):1412-1419.
[50]Herrera R. Modulation of hepatocyte growth factor-induced scattering of HT29 colon carcinoma cells. Involvement of the MAPK pathway. J Cell Sci.1998 Apr; 111 (Pt 8):1039-1049.
[51]Badache A, Hynes NE. Interleukin 6 inhibits proliferation and, in cooperation with an epidermal growth factor receptor autocrine loop, increases migration of T47D breast cancer cells. Cancer Res.2001 Jan 1; 61(1):383-391.
[52]Tomanek RJ, Schatteman GC. Angiogenesis:new insights and ther- apeutic potential. Anat Rec.2000; 261:126-135.
[53]Bancroft CC, Chen Z, Dong G, Sunwoo JB, Yeh N, Park C, Van Waes C. Coexpression of proangiogenic factors IL-8 and VEGF by human head and neck squamous cell carcinoma involves coactive- tion by MEK-MAPK and IKK-NF-kappaB signal pathways. Clin Cancer Res.2001 Feb; 7(2):435-442.
[54]Zhou JN, Ljungdahl S, Shoshan MC, Swedenborg J, Linder S. Acti vation of tissue-factor gene expression in breast carcinoma cells by stimulation of the RAF-ERK signaling pathway. Mol Carcinog.1998 Apr; 21(4):234-43.
[55]Hulboy DL, Rudolph LA, Matrisian LM. Matrix metalloproteinases as mediators of reporductive function. Molecular Human Reprodu-ction.1997; 3:27-45.
[56]Birkedal-Hansen H, Moore WGL, Bodden MK, Windor LJ, Birke- dal- Hansen
B, DeCarl A, et al. Matrix metalloproteinase:a review. Crit Rev Oral Biol Med. 1993; 4:197-250.
[57]Shapiro SD. Matrix metalloproteinase degradation of extracelluar matrix:biolog-ical consequence. Curr Opin Cell Biol.1998; 10:602-603.
[58]Nagase H. Activation mechanisms of matrix metalloproteinase. Biol Chem Hoppe Seyler.1997; 378:151-160.
[59]Nagase H,Woessner JF. Matrix metalloproteinase. J Boil Chem 1999; 274: 21491-21494.
[60]Vu TH, Werb Z. Matrix metalloproteinase:effectors of development and normal physiology. Gences Dev.2000; 14:2123-2133.
[61]Dis S, et al.IL-12 regulates VEGF and MMPs in a murine breast cancer model. Int J Cancer.1998 Oct 29;78(3):361-365.
[62]Gum R, Lengyel E, Juarez J, Chen JH, Sato H, Seiki M, Boyd D. Stimulation of 92-kDa gelatinase B promoter activity by ras is mitogen-activated protein kinase kinase 1-independent and requires multiple transcription factor binding sites including closely spaced PEA3/ets and AP-1 sequences. J Biol Chem.1996 May 3; 271(18):10672-10680.
[63]Van Wart H, Birkedal-Hansen H. The cystein switch:a principle of regulation of metalloproteinase activity with potential applicability to the entire matrix metalloproteinase gene family. Proc Natl Acad Sci USA.1990; 87:5578-5582.
[64]Mignatti P, Rifkin DB. Plasminogen activators and matrix metallo-proteinases in angiogenesis. Enzyme Proein.1996; 49:117-137.
[65]DeCLerck YA, Laug WE. Plasminogen activators and their inhibit- tors: cooperation between matrix metalloproteinases and the plasminogen activator-plasmin system in tumor progression. Enzyme Protein.1996; 49:72-84
[66]Rosenthal EL, Johson TM, Allen ED, Apel IJ, Punturieri A, Weiss SJ. Role of plasminogen activator and matrix metalloproteinase systems in epidermal growth factor and scatter factor-stimulated invasion of carcinoma cells. Cancer Res. 1998; 58:5221-5230.
[67]Deryugina EI, Zijlstra A, Partridge JJ, Kupriyanova TA, Madsen MA,
Papagiannakopoulos T, Quigley JP. Unexpected effect of matrix metallo-proteinase down-regulation on vascular intravasation and metastasis of human fibrosarcoma cells selected in vivo for high rates of dissemination. Cancer Res. 2005 Dec 1;65(23):10959-10969.
[68]Cox G, O'Byrne KJ. Matrix metalloproteinases and cancer. Anticancer Research. 2001; 21:4207-4220.
[69]Liotta L. Isolation of a protein that stimulates blood vessel growth. Nature,1985, 318(6041):14.
[70]Ricca A, Biroccio A, Bufalo DD, et al. bcl-2 over-expression enhances NF-kappaB activity and induces mmp-9 transcription in human MCF7(ADR) breast-cancer cells. Int J Cancer.2000; 86(2):188-196.
[71]Shalinsky DR, Brekken J, Zou H, et al. Broad Antitumor and Antiangiogenic Activities of AG3340, a Potent and Selective MMP Inhibitor Undergoing Advanced Oncology Clinical Trials. Ann NY Acad Sci.1999; 878:236-270.
[72]Shalinsky DR, Brekken J, Zou H, et al. Marked Antiangiogenic and Antitumor Efficacy of AG3340 in Chemoresistant Human Non-Small Cell Lung Cancer Tumors:Single Agent and Combination Chemotherapy Studies. Clin Cancer Res. 1999; 5:1905-1917.
[73]Patterson BC, Amy Sang QX. Angiostatin-converting Enzyme Activities of Human Matrilysin (MMP-7) and Gelatinase B/Type IV Collagenase (MMP-9). J Biol Chem.1997; 272:28823-28825.
[74]Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC. Potent and specific genetic interference by double-strand RNA in Caenorhabditis elegans Nature.1998; 391(6669):744-745.
[75]McManus MT, Sharp PA. Gene silencing in mammals by small interfering RNAs. Nature.2002; 3(10):737-747.
[76]Hannon GJ. RNA interference. Nature.2002; 418 (6894):244-251.
[77]Zamore PD, Tuschl T, Sharp PA, Bartel DP. RNAi:double-strand RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell. 2000; 101:25-33.
[78]Caplen NJ, Parrish S, Imani F, Fire A, Morgan RA. Specific inhibition of gene expression by small double-stranded RNAs in invertebrate and vertebrate systems. Proc Natl Acad Sci USA.2001; 98(17):9742-9747.
[79]Nykanen A, Haley B, Zamore PD. ATP requirements and small interferings RNA structure in the RNA interference pathway. Cell.2001; 107(3):309-321.
[80]Gil J, Esteban M. Induction of apoptosis by the dsRNA dependent protein kinase (PKR):mechanism of action. Apoptosis.2000; 5 (2):107-114.
[1]黄志强,周宁新,黄晓强.肝门部胆管癌的外科治疗.中华消化外科杂志.2003；4：
[2]周宁新,黄志强,刘永雄.肝外胆道癌全国调查1098例分析.中华外科杂志.1990；28：516-520.
[3]黄志强.肝门部胆管癌的外科治疗.中华外科杂志.1990；28：522-524.
[4]Gebbia V, Majello E, Testa A, et al.Treatment of advanced adenocar- cinoma of the exocrine pancreas and the gallbladder with 5-Fu, high dose levofolinic acid and oral hydroxyurea on weekly schedule, Results of a multicenter study of the southern Italy Incology Group (G.O.I.M). Cancer.1996;78:1300-1307.
[5]黄志强.肝门部胆管癌外科治疗的现状与展望.中国普外基础与临床杂志.2005：12：317-320.
[6]Volm M, Koomagi R, Mattern J, et al. PD-ECGF, b-FGF, and VEGF expression in non small cell lung carcinomas and their association with lymph node metastasis. Anticancer Res.1999;19:65-112.
[7]Maehara Y, Kabashima A, Koga T, et al. Vascular invasion and potential for tumor angiogenesis and metastasis in gastric carcinoma. Surgery.2000;128:408.
[8]Fujisaki K, Mitsuyama K, Toyonaga A, et al. Circulating vascular endothelial growth factor in patients with colorectal cancer. Am J Gastroenterol.1998; 93: 249.
[9]Jeffrey E, Isaish J. Targeting Lymphatic Metastasis. Science.2002; 296: 1811-1812.
[10]Lillemoe KD, Cameron JL. Surgery for hilar cholangiocarcinoma:the Johns Hopkins approach. J Hepatobiliary Pancreat Surg.2000; 7:115-121.
[11]周宁新.肝门部胆管癌的外科治疗新进展.中国医师进修杂志.2006；29：6-7.
[12]Imai M, Hoshi T, Ogawa K. K-ras codon 12 mutations biliary tract tumors detected by polymerase chain reaction denaturing gradient gel electrophoresis. Cancer.2001;73:2724-2733.
[13]Joukov V, Pajusola K, Kaipainen A, et al. A novel vascular endothelial growth factor, VEGF-C, is a ligand for the Flt4 (VEGFR-3) and KDR (VEGFR-2) receptor tyrosine kinases. EMBO J.1996;15:290-298.
[14]Li XR, Eriksson U. Novel VEGF family members:VEGF-B, VEGF-C and VEGF-D. Int J Biochem Cell Biol.2001; 33:421-426.
[15]Pan J, Hua C, Wen Y. Cloning and sequencing of human vascular endothelial growth factor-C encoded cDNA. Hua Xi Kou Qiang Yi Xue Za Zhi.2003; 21(4): 318-320.
[16]Dixelius J, Makinen T, Wirzenius M, et al. Ligand induced vascular endothelial growth factor receptor-3 (VEGFR-3) hetero-dimerization with VEGFR-2 in primary lymphatic endothelial cells regulates tyrosine phosphorylation sites. J Biol Chem.2003; 78 (42):40973-40979.
[17]Meyer RD, Rahimi N. Comparative structure function analysis of VEGFR-1 and VEGFR-2:What have we learned from chimeric systems. Ann N Y Acad Sci. 2003; (995):200-207.
[18]Ryden L, Linderholm B, Nielsen NH, et al.Tumor specific VEGF-A and VEGFR2/KDR protein are coexpressed in breast cancer. Breast Cancer Res Treat. 2003; 82(3):147-154.
[19]Nisato RE, Tille JC, Pepper MS. Lymphangiogenesis and tumor metastasis. Thromb Haemost.2003; 90(4):591-597.
[20]Skobe M, Hamberg LM, Hawighorst T, et al. Concurrent induction of lymphan-giogenesis, angiogenesis, and maemphage recruitment by vascular endothelial growth factor-C in melanoma. Am J Pathol.2001; 159:893-903.
[21]Mandriota SJ, Jussila L, Jeltsch M, et al. Vascular endothelial growth factor-C mediated lymphangiogenesis promotes tumor metastasis. EMBO J.2001; 20:672-682.
[22]Stacker SA, Caesar C, Baldwin ME, et al. VEGF-D promotes the metastatic spmad of tumor cells via the lymphatics. Nat Med.2001; 7:186-191.
[23]Von Marsehall Z, Scholz A, Stacker SA, et al. Vascular endothelial growth factor-D induces lymphangiogenesis and lymphatic-metastasis in models of ductal pancreatic cancer. Int J Oncol.2005; 27:669-679.
[24]Schietroma C, Cianfarani F, Lacal PM,et al. Vascular endothelial growth factor-C expression correlates with lymph node localizat-ion of human melanoma metastases. Cancer.2003; 98(4):789-797.
[25]Hung CJ, Ginzinger DG, Zarnegar R, et al. Expression of vascular endothelial growth factor-C in benign and malignant thyroid tumors. J Clin Endocrinol Metab.2003; 88(8):3694-3699.
[26]Liu F, Zhang YJ. Roles of VEGF-C and its receptor Flt-4 in proliferation and metastasis of primary breast cancer. Ai Zheng.2003; 22(10):1053-1056.
[27]Kimura Y, Watanabe M, Ohga T, et al. Vascular endothelial growth factor-C expression correlates with lymphatic involvement and poor prognosis in patients with esophageal squamous cell carcinoma. Oncol Rep.2003; 10(6):1747-1751.
[28]Maeda K, Yashiro M, Nishihara T, et al. Correlation between vascular endothelial growth factor-C expression and lymph node metastasis in Tl carcinoma of the colon and rectum. Surg Today.2003; 33(10):736-739.
[29]Tamura M, Ohta Y. Serum vascular endothelial growth factor-C level in patients with primary nonsmall cell lung carcinoma:a possible diagnostic tool for lymph node metastasis. Cancer.2003; 98(6):1217-1222.
[30]Sedivy R, Mannagetta J, Haverkampf C, et al. Expression of vascular endothelial growth factor-C correlates with the lymphatic microvessel density and the nodal status in oral squamous cell cancer. J Oral Pathol Med.2003; 32(8):455-460.
[31]Ogawa E, Takenaka K, Yanagihara K, et al. Clinical significance of VEGF-C status in tumour cells and stromal macrophages in non-small cell lung cancer patients. Br J Cancer.2004; 91:498-503.
[32]Skobe M, Hawighorst T, Jackson DG, et al. Induction of tumor lymphan-giogenesis by VEGF-C promotes breast cancer metastasis. Nat Med.2001; 7(2):192-198.
[33]Partanen TA, Paavonen K. Lymphatic versus blood vascular endothelial growth factors and receptors in humans. Microsc Res Tech.2001; 55(2):108-121.
[34]Karpanen T, Egeblad M, Karkkainen MJ, et al. Vascular endothelial growth factor-C promotestumor lymphangiogenesis and intralym-phatic tumor growth. Cancer Res.2001; 61(5):1786-1790.
[35]张萌,彭利,王顺祥等.VEGF与VEGF-C蛋白在胆管癌中的表达及意义.山东医药.2007；6：17-18.
[36]Karpanen T, Alitalo K. Lymphatic vessels as targets of tumor therapy. J Exp Med. 2001; 194(6):37-42.
[37]Hamada K, Oike Y, Takakura N, et al. VEGF-C signaling pathways through VEGFR-2 and VEGFR-3 in vasculoangiogenesis and hematopoiesis. Blood.
2000; 96(12):3793-3800.
[38]Valtola R, Salven P, Heikkila P. VEGFR-3 and its ligand VEGF-C are associated with angiogenesis in breast cancer. Am J Pathol.1999; 154(5):1381-1390.
[39]Zachary I. Signaling mechanisms mediating vascular protective actions of vascular endothelial growth factor. Am J Physiol Cell Physiol.2001; 280(6): 1375-1386.
[40]Nathanson SD. Insights into the mechanism of lymph node metastasis. Cancer. 2003;98(2):413-423.
[41]Jeltsch M, Kaipainen A, Joukov V, et al. Hyperplasia of lymphatic vessels in VEGF-C transgenic mice. Science.1997; 276 (5317):1423-1425.
[42]Ueda M, Teraj Y, Yamashita Y, et al. Correlation between vascular endothelial growth factor-C expression and invasion phenotype in cervical carcinomas. Int J Cancer.2002; 98:335-343.
[43]George ML, Tutton MG, Janssen F, et al. VEGF-A, VEGF-C and VEGF-D in colorectal cancer progression. Neoplasia.2001; 3:420-427.
[44]常实,汤恢焕,周军等.肝外胆管癌nm23和VEGF的表达及其临床意义.中国普通外科杂志.2003；12(18)：625-627.
[45]Benckert C, Sven Jonas, Cramer T, et al. Transforming growth factor beta-1 stimulates vascular endothelial growth factor gene transcription in human cholangiocellular carcinoma cells. Cancer Research.2003; 63(5):1083-1092.
[1]L.J. Van't Veer, B. Weigelt, Road map to metastasis, Nat. Med.9 (2003) 999-1000.
[2]D. Hanahan, R.A. Weinberg, The hallmarks of cancer, Cell 100 (2000) 57-70.
[3]N. Ferrara, The role of vascular endothelial growth factor in pathological angiogenesis, Breast. Cancer. Res. Treat.36 (1995) 127-137.
[4]H.F. Dvorak, L.F. Brown, M. Detmar, A.M. Dvorak, Vascular permeability factor/vascular endothelial growth factor, microvascular hyperpermeability, and angiogenesis, Am. J. Pathol.146 (1995) 1029-1039.
[5]J.D. Hood, R. Frausto, W.B. Kiosses, M.A. Schwartz, D.A. Cheresh, Differential alphav integrin-mediated Ras-ERK signaling during two pathways of angiogenesis, J. Cell Biol.162 (2003) 933-943.
[6]S.S. Smyth, C. Patterson, Tiny dancers:the integrin-growth factor nexus in angiogenic signaling, J. Cell Biol.158 (2002) 17-21.
[7]R.O. Hynes, Integrins:versatility, modulation, and signaling in cell adhesion, Cell 69(1992)11-25.
[8]R.O. Hynes, Integrins:bidirectional, allosteric signaling machines, Cell 110 (2002) 673-687.
[9]J.M. Breuss, N. Gillett, L. Lu, D. Sheppard and R. Pyetala, Restricted distribution of integrin beta 6 mRNA in primate epithelial tissues, J. Histochem. Cytochem. 41 (1993)1521-1527.
[10]Z.Y. Zhang, K.S. Xu, J.S. Wang, GY. Yang, W. Wang, J.Y. Wang, W.B. Niu, E.Y. Liu, Y.T. Mi, J. Niu, Integrin alphanvbeta6 acts as a prognostic indicator in gastric carcinoma, Clin. Oncol.20 (2008) 61-66.
[11]G.Y. Yang, K.S. Xu, Z.Q. Pan, Z.Y. Zhang, Y.T. Mi, J.S. Wang, R. Chen, J. Niu, Integrin alphavbeta6 mediates the potential for colon cancer cells to colonize in and metastasize to the liver, Cancer. Sci.99 (2008) 879-887.
[12]J. Wang, Z. Zhang, K. Xu, X. Sun, G. Yang, W. Niu, E. Liu, C. Peng, P. Lin, J. Wang, R. Chen, M. Agrez, J. Niu, Suppression of integrin alphaupsilonbeta6 by RNA interference in colon cancer cells inhibits extracellular matrix degradation through the MAPK pathway, Int. J. Cancer.123 (2008) 1311-1317.
[13]A. Kabashima, Y. Maehara, Y. Kakeji, H. Baba, T. Koga, K. Sugimachi, Clinicopathological features and overexpression of matrix metalloproteinases in intramucosal gastric carcinoma with lymph node metastasis, Clin. Cancer. Res. 9(2000) 3581-3584.
[14]A. Torii, Y. Kodera, K. Uesaka, T. Hirai, K. Yasui, T. Morimoto, Y. Yamamura, T. Kato, T. Hayakawa, N. Fujimoto, T. Kito, Plasma concentration of matrix metalloproteinase 9 in gastric cancer, Br. J. Surg.84(1997) 133-136.
[15]D.R. Senger, C.A. Perruzzi, M. Streit, V.E. Koteliansky, A.R. Fougerolles, M. Detmar, The alpha(1)beta(1) and alpha(2)beta(1) integrins provide critical support for vascular endothelial growth factor signaling, endothelial cell migration, and tumor angiogenesis, Am. J. Pathol.160 (2002) 195-204.
[16]K. Podar, Y.T. Tai, B.K. Lin, R.P. Narsimhan, M. Sattler, T. Kijima, R. Salgia, D. Gupta, D. Chauhan, K.C. Anderson, Vascular endothelial growth factor-induced migration of multiple myeloma cells is associated with betal integrin-and phosphatidylinositol 3-kinase-dependent PKC alpha activation, J. Biol. Chem. 277(2002)7875-7881.
[17]N.E. Vlahakis, B.A. Young, A. Atakilit, A.E. Hawkridge, R.B. Issaka, N. Boudreau, D. Sheppard, Integrin alpha9betal directly binds to vascular endothelial growth factor (VEGF)-A and contributes to VEGF-A-induced angiogenesis, J. Biol. Chem.282 (2007) 15187-15196.
[18]D. Sheppard, C. Rozzo, L. Starr, V. Quaranta, D.J. Erle, R. Pytela, Complete amino acid sequence of a novel integrin beta subunit (beta 6) identified in epithelial cells using the polymerase chain reaction, J. Biol. Chem.265 (1990) 11502-11507.
[19]J.M. Breuss, J. Gallo, H.M. DeLisser, I.V. Klimanskaya, H.G. Folkesson, J.F. Pittet, S.L. Nishimura, K. Aldape, D.V. Landers, W. Carpenter, et al, Expression of the beta 6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling, J. Cell. Sci.108 (1995) 2241-2251.
[20]Z.Y. Zhang, K.S. Xu, Q.S. He, W.B. Niu, J.Y. Wang, Y.T. Mi, J.S. Wang, G.Q. Wang, G.Y. Yang, J. Niu, Signaling and regulatory mechanisms of integrin alphavbeta6 on the apoptosis of colon cancer cells, Cancer. Lett.266 (2008) 209-215.
[21]X. Gu, J. Niu, D.J. Dorahy, R. Scott, M.V. Agrez, Integrin alpha(v)beta6-associated ERK2 mediates MMP-9 secretion in colon cancer cells, Br. J. Cancer. 87(2002)348-351.
[22]B.P. Eliceiri, X.S. Puente, J.D. Hood, D.G. Stupack, D.D. Schlaepfer, X.Z. Huang, D. Sheppard, D. A. Cheresh, Src-mediated coupling of focal adhesion kinase to integrin alpha(v)beta5 in vascular endothelial growth factor signaling, J. Cell. Biol.157(2002)149-159.
[23]T.H. Lee, S. Seng, H. Li, S.J. Kennel, H.K. Avraham, S. Avraham, Integrin regulation by vascular endothelial growth factor in human brain microvascular endothelial cells:role of alpha6betal integrin in angiogenesis, J. Microbiol.44 (2006) 502-507.
[24]K. Milde-Langosch, A.M. Bamberger, G. Rieck, D. Grund, G. Hemminger, V. Muller, T. Loning, Expression and prognostic relevance of activated extracellular-regulated kinases (ERK1/2) in breast cancer, Br. J. Cancer.92 (2005)2206-2215.
[25]A.A. Adjei, The role of mitogen-activated ERK-kinase inhibitors in lung cancer therapy, Clin. Lung. Cancer.7 (2005) 221-223.
[26]N. Ahmed, C. Riley, G.E. Rice, M.A. Quinn, M.S. Baker. Alpha(v)beta(6) integrin-A marker for the malignant potential of epithelial ovarian cancer, J. Histochem. Cytochem.50 (2002) 1371-1380.
[27]J.S. Krueger, V.G. Keshamouni, N. Atanaskova, K.B. Reddy, Temporal and quantitative regulation of mitogen-activated protein kinase (MAPK) modulates cell motility and invasion, Oncogene 20 (2001) 4209-4218.
[28]N. Ahmed, F. Pansino, M. Baker, G. Rice, M. Quinn, Association between alphavbeta6 integrin expression, elevated p42/44 kDa MAPK, and plasminogen-dependent matrix degradation in ovarian cancer, J. Cell. Biochem. 84 (2002) 675-686.
[29]N. Ahmed, J. Niu, D.J. Dorahy, X. Gu, S. Andrews, C.J. Meldrum, R.J. Scott, M.S. Baker, I.G. Macreadie, M.V. Agrez, Direct integrin alphavbeta6-ERK binding:implications for tumor growth, Oncogene 21(2002) 1370-1380.
[30]J. Niu, D.J. Dorahy, X. Gu, R.J. Scott, B. Draganic, N. Ahmed, M.V. Agrez, Integrin expression in colon cancer cells is regulated by the cytoplasmic domain of the beta6 integrin subunit, Int. J. Cancer.99 (2002) 529-537.
[1]Patel T. Increasing incidence and mortality of primary intrahepatic cholangiocar-cinoma in the United States. Hepatology 2001; 33:1353-1357.
[2]Gores GJ. Cholangiocarcinoma:current concepts and insights. Hepatology 2003; 37:961-969.
[3]N. Ferrara. The role of vascular endothelial growth factor in pathological angio-genesis. Breast Cancer Res Treat.1995; 36:127-137.
[4]H.F. Dvorak, L.F. Brown, M. Detmar, A.M. Dvorak, Vascular permeability factor/vascular endothelial growth factor, microvascular hyperpermeability, and angiogenesis. Am J Pathol.1995; 146:1029-1039.
[5]Salven P, Lymboussaki A, Heikkila P. Vascular endothelial growth factors VEGF-B and VEGF-C are expressed in human tumors. Am J Pathol 1998; 153:103-108.
[6]O-charoenrat P, Rhys-Evans P, Eccles SA. Expression of vascular endothelial growth factor family members in head and neck squamous cell carcinoma correlates with lymph node metastasis. Cancer.2001; 92:556-65.
[7]Tsurusaki T, Kanda S, Sakai H. Vascular endothelial growth factor-C expression in human prostatic carcinoma and its relationship to lymph node metastasis. Br J Cancer.1999; 80:309-313.
[8]George ML, Tutton MG, Janssen F, Arnaout A, Abulafi AM, Eccles SA, et al. VEGF-A, VEGF-C and VEGF-D in colorectal cancer progression. Neoplasia. 2001; 3:420-7.
[9]Makoto I, Joji K, Shinsuke K and Hirokazu N. Expression of Vascular Endothelial Growth Factor C and D (VEGF-C and -D) is an Important Risk Factor for Lymphatic Metastasis in Undifferentiated Early Gastric Carcinoma. Jpn J Clin Oncol.2003; 33(1):21-27.
[10]Yonemura Y, Endo Y, Fujita H, Fushida S et al. Role of vascular endothelial growth factor C expression in the development of lymph node metastasis in gastric cancer. Clin Cancer Res.1999; 5:1823-1829.
[11]Zhao R, Liu XQ, Wu XP, et al. Vascular endothelial growth factor (VEGF) enhances gastric carcinoma invasiveness via integrin alpha(v)beta6. Cancer Lett. 2010 Jan 28; 287(2):150-156.
[12]Chang Y, Wu XY. JNK1/2 siRNA inhibits transforming-growth factor-betal-induced connective tissue growth factor expression and fibrotic function in THSFs. Mol Cell Biochem.2010 Feb; 335(1-2):83-89.
[13]Karkkainen MJ, Petrova TV. Vascular endothelial growth factor receptors in the regulation of angiogenesis and lymphangiogenesis. Oncogene.2000; 19:5598-5605.
[14]Enholm B, Paavonen K, Ristimaki A. Comparison of VEGF, VEGF-B, VEGF-C and Ang-1 mRNA regulation by serum, growth factors, oncoproteins and hypoxia. Oncogene.1997; 14:2475-2783.
[15]Joukov V, Pajusola K, Kaipainen A, Chilov D, Lahtinen I, Kukk E, et al. A novel vascular endothelial growth factor, VEGF-C, is a ligand for the Flt4 (VEGFR-3)
and KDR (VEGFR-2) receptor tyrosine kinases [erratum]. EMBO J.1996; 15: 1751.
[16]Skobe M, Hawighorst T, Jackson DG, et al.Induction of tumor lymphangio-genesis by VEGF-C promotes breast cancer metastasis. Nature Med.2001; 7: 192-198.
[17]Masatsugu Ueda, Yao-Ching Hung, YoshitoTerai,et al.Vascular Endothelial Growth Factor-CExpression and Invasive Phenotype in Ovarian Carcinomas. Clin Cancer Res.2005; 11(9):3225-3232.
[18]VanTrappen PO, Ryan A, CarrollM, et al. A model for co-expression pattern analysis of genes implicated in angiogenesis and tumour cell invasion in cervical cancer. Br J Cancer.2002; 87:537-544.
[2]Correa P, Chen VW. Gastric cancer. Cancer Surv.1994; 19(20):55-76.
[3]Aragones N, Pollan M, Rodero I, Lopez-Abente G. Gastric cancer inthe European Union (1968-1992):mortality trends and cohort effect. Ann Epidemiol.1997; 7: 294-303.
[4]Ferretti S, Gafa L. Upper gastrointestinal tract cancers:oesophagus, stomach, liver, gallbladder and biliary ducts, pancreas. Epidemiol Prev.2004; 28:34-42.
[5]Kelley JR, Duggan JM. Gastric cancer epidemiology and risk factors. J Clin Epidemiol.2003; 56:1-9.
[6]Devesa SS, Blot WJ, Fraumeni Jr JF. Changing patterns in the incidence of esophageal and gastric carcinoma in the United States. Cancer.1998; 83:2049-2053.
[7]Powell J, McConkey CC. Increasing incidence of adenocarcinoma of the gastric cardia and adjacent sites. Br J Cancer.1990; 62:440-443.
[8]Van't Veer LJ, Weigelt B. Road map to metastasis. Nat. Med.2003; 9:999-1000.
[9]Hanahan D, Weinberg RA. The hallmarks of cancer. Cell.2000; 100:57-70.
[10]Ferrara N. The role of vascular endothelial growth factor in pathological angio-genesis. Breast Cancer Res. Treat.1995; 36:127-137.
[11]Dvorak HF, Brown LF, Detmar M, Dvorak AM. Vascular permeability factor/vascular endothelial growth factor, microvas-cular hyperpermeability, and angiogenesis. Am. J. Pathol.1995; 146:1029-1039.
[12]Hood JD, Frausto R, Kiosses WB, Schwartz MA, Cheresh DA. Differential alphav integrin-mediated Ras-ERK signaling during twopathways of angiogene-sis, J Cell Biol.2003; 162:933-943.
[13]Smyth SS, Patterson C. Tiny dancers:the integrin-growth factor nexus in angiogenic signaling. J Cell Biol.2002; 158:17-21.
[14]Schietroma C, Cianfarani F, Lacal PM,et al. Vascular endothelial growth factor-C expression correlates with lymph node localizat-ion of human melanoma metastases. Cancer.2003; 98(4):789-797.
[15]Hung CJ, Ginzinger DG, Zarnegar R, et al. Expression of vascular endothelial growth factor-C in benign and malignant thyroid tumors. J Clin Endocrinol Metab.2003; 88(8):3694-3699.
[16]Liu F, Zhang YJ. Roles of VEGF-C and its receptor Flt-4 in proliferation and metastasis of primary breast cancer. Ai Zheng.2003; 22(10):1053-1056.
[17]Kimura Y, Watanabe M, Ohga T, et al. Vascular endothelial growth factor-C expression correlates with lymphatic involvement and poor prognosis in patients with esophageal squamous cell carcinoma. Oncol Rep.2003; 10(6):1747-1751.
[18]Maeda K, Yashiro M, Nishihara T, et al. Correlation between vascular endothelial growth factor-C expression and lymph node metastasis in T1 carcinoma of the colon and rectum. Surg Today.2003; 33(10):736-739.
[19]Tamura M, Ohta Y. Serum vascular endothelial growth factor-C level in patients with primary nonsmall cell lung carcinoma:a possible diagnostic tool for lymph node metastasis. Cancer.2003; 98(6):1217-1222.
[20]Sedivy R, Mannagetta J, Haverkampf C, et al. Expression of vascular endothelial growth factor-C correlates with the lymphatic microvessel density and the nodal status in oral squamous cell cancer. J Oral Pathol Med.2003; 32(8):455-460.
[21]Futagami S, Tatsuguchi A, Hiratsuka T, et al. Monocyte chemoat-tractant protein-1 and CD40 ligation have a synergistic effect on vascular endothelial growth factor production through cyclooxy-genase upregulation in gastric cancer. J Gastroenterol.2008; 43(3):216-224.
[22]Sheppard D, Rozzo C, Starr L, Quaranta V, Erle DJ, Pytela R. Complete amino acid sequence of a novel integrin beta subunit (beta6) identified in epithelial cells using the polymerase chain reaction. J Biol Chem.1990; 265:11502-11507.
[23]Breuss JM, Gallo J, DeLisser HM, Klimanskaya IV, Folkesson HG, Pittet JF, Nishimura SL, Aldape K, Landers DV, Carpenter W, et al. Expression of the beta6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling. J Cell Sci.1995; 108:2241-2251.
[24]Zhang ZY, Xu KS, He QS, Niu WB, Wang JY, Mi YT, Wang JS, Wang GQ, Yang GY, Niu J. Signaling and regulatory mechanisms of integrin alphavbeta6 on the apoptosis of colon cancer cells. Cancer Lett.2008; 266:209-215.
[25]Mattern J, Koomagi R, Volm M. Association of vascular endothelial growth factor expression with intratumoral microvessel density and tumor cell proliferation in human epidermoid lung carcinoma. Br J Cancer.1996; 73:931-934.
[26]吴胤瑛,李恩孝.VEGF在恶性肿瘤中的研究进展.现代肿瘤医学.2005,13(5)：18-21.
[27]Carmeliet P, Jain RK. Angiogenesis in cancer and other diseases. Nature.2000, 407(6801):249-257.
[28]Poon RT, Fan ST, Wong J. Clinical implications of circulating angiogenic factors in cancer patients. J Clin Oncol.2001,19(4):1207-1225.
[29]Frish SM, Ruoslahti E.Integrin and anoikis. Curr Opin Cell Biol.1997; 9:701-706.
[30]Frish SM, Screaton RA. Anoikis mechanisms. Curr Opin Cell Biol.2001; 13:555-562.
[31]Hynes RO. Integrins:bidirectional, allosteric signaling machines. Cell.2002; 110: 673-687.
[32]Hynes RO. Integrins:versatility, modulation and signaling in cell ahhesion. Cell.1992; 69:11-25.
[33]Akiyama SK, Olden K, Yamada KM. Fibronectin and integrins in invasion and metastasis. Cancer Metastasis Rev.1995; 14:173-189.
[34]Ferrara N, Henzel WJ. Pituitary follicular cells secrete a novel heparin-binding growth factor specific for vascular endothelial cells. Biochem Biophys Res Commun.1989,15;161(2):851-858.
[35]Petrova TV, Makinen T, Alitalo K. Signaling via vascular endothe-lial growth factor receptors. Exp Cell Res.1999; 253(1):117-130.
[36]Sasaki T, Tanno S, Shibukawa K, Osanai S, Kawabe J, Ohsaki Y. Administration of VEGF receptor tyrosine kinase inhibitor increases VEGF production causing angiogenesis in human small-cell lung cancer xenografts. Int J Oncol.2008 Sep; 33(3):525-532.
[37]Los M, Roodhart JM, Voest EE. Target practice:lessons from phase III trials with bevacizumab and vatalanib in the treatment of advanced colorectal cancer. Oncologist.2007; 12:443-450
[38]Marbeuf-Gueye C, Salerno M, Quidu P, Garnier-Suillerot A. Inhibition of the P-glycoprotein and multidrug resistance protein-mediated efflux of anthracyclines and calceinacetoxymethyl ester by PAK-104P. Eur J Pharmacol. 2000 Mar 17; 391(3):207-216.
[39]Motoji T, Motomura S, Wang YH. Multidrug resistance of acute leukemia and a strategy to overcome it. Int J Hematol.2000 Dec; 72(4):418-424.
[40]Poirson-Bichat F, Goncalves RA, Miccoli L, Dutrillaux B, Poupon MF. Methionine depletion enhances the antitumoral efficacy of cytotoxic agents in drug-resistant human tumor xenografts. Clin Cancer Res.2000 Feb; 6(2):643-653.
[41]Mistry P, Stewart AJ, Dangerfield W, et al. In vitro and in vivo characterization of XR11576, a novel, orally active, dual inhibitor of topoisomerase Ⅰ and Ⅱ. Anticancer Drugs.2002 Jan; 13(1):15-28.
[42]Van Hattum AH, Pinedo HM, Schluper HM, Hausheer FH, Boven E. New highly lipophilic camptothecin BNP1350 is an effective drug in experimental human cancer. Int J Cancer.2000 Oct 15; 88(2):260-266.
[43]Giancotti FG, Ruoslahti E.Integrin signaling. Science.1999 Aug 13; 285 (5430): 1028-1032.
[44]Crowe DL, Shuler CF. Regulation of tumor cell invasion by extrac ellular matrix. Histol Histopathol.1999 Apr; 14(2):665-671.
[45]De Melker AA, Sonnenberg A. Integrins:alternative splicing as a mechanism to regulate ligand binding and integrin signaling events. Bioessays.1999 Jun; 21(6): 499-509.
[46]Hynes RO. Integrins:versatility, modulation and spreading in cell adhesion. Cell.1992; 69:11-25.
[47]Albelad SM. Biology of disease:role of integrins and other cell adhesion molecules in tumor progression and metastasis. Lab Invest.1993; 68:4-17.
[48]Cheresh D. Integrins structure, function and biological properties. Adv Mol Cell Biol.1993; 6:225-228.
[49]Albelda SM, Buck CA. Integrins and other cell adhesion molecules. FASEB J. 1990; 4:2868-2880.
[50]Liotta LA, Rao CN. Tumor invasion and the extracellular matrix. Lab Invest. 1983;49(6):636-649.
[51]郑世曦,陈汉琴,杨军等.层粘连蛋白在人原发性肝细胞癌转移中的意义.华人消化杂志.1998；6：507-509.
[52]Meredith JE Jr, Winitz S, Lewis JM, Hess S, Ren XD, Renshaw MW, Schwartz MA. The regulation of growth and intracellular signaling by integrins. Endocr Rev.1996 Jun; 17(3):207-220.
[53]Sjaastad MD, Nelson WJ. Integrin-mediated calcium signaling and regulation of cell adhesion by intracellular calcium. Bioessays.1997 Jan; 19(1):47-55.
[54]Shimizu Y, Hunt SW. Regulating integrin-mediated adhesion:one more function for PI 3-kinase? Immunol Today.1996 Dec; 17(12):565-573.
[55]Hanks SK, Polte TR. Signaling through focal adhesion kinase. Bioessays.1997 Feb; 19(2):137-145.
[56]Hannigan GE, Dedhar S. Protein kinase mediators of integrin signal transduction. J Mol Med.1997 Jan; 75(1):35-44.
[57]Dedhar S, Williams B, Hannigan G. Integrin-linked kinase (ILK):a regulator of integrin and growth-factor signalling. Trends Cell Biol.1999 Aug; 9(8):319-323.
[58]Huang Y, Wu C. Integrin-linked kinase and associated proteins (review). Int J Mol Med.1999 Jun; 3(6):563-572.
[59]Petty HR, Todd RF. Integrins as promiscuous signal transduction devices. Immunol Today.1996 May; 17(5):209-212.
[60]Todd RF, Petty HR. Beta2 (CD 11/CD 18) integrins can serve as signaling partners for other leukocyte receptors. J Lab Clin Med.1997 May; 129(5):492- 498.
[61]Paulus W, Baur I, Schuppan D et al. Characterization of integrin receptors in normal and neoplastic human brain. Am J Pathol.1993; 143(1):154-163.
[62]Mrrjans F, Meyer T, Frrrschen C et al. In vivo therapy of malignant melanoma by means of antagonists of αv integrins. Int J Cancer.2000; 87:716-723.
[63]Liapis H, Adler LM, Wick MR et al. Expression of alpha(v)beta3 integrin is less frequent in ovarian epithelial tumors of low malignant potential in contrast to ovarian carcinomas. Hum Pathol.1997; 28(4):443-449.
[64]Rhim JS, Tsai WP, Chen ZQ et al. A human vascular endothelial cell model to study angiogenesis and tumorigenesis. Carcinogene- sis.1998; 19(4):673-681.
[65]Erdreich EA, Shimada H, Groshen S et al. Integrins alpha(v)beta3 and alpha(v) beta5 are expressed by endothelium of highrisk neuroblastoma and their inhibition is associated with increased endogenous ceramide. Cancer Res.2000; 60(3):712-721.
[66]Takano S, Tsuboi K, Tomono Y et al. Tissue factor, osteopontin, alphavbeta3 integrin expression in microvasculature of glioma associated with vascular endothelial growth factor expression. Br J Cancer.2000; 82(12):1967-1973.
[67]Max R, Gerritsen RR, Nooijen PT et al. Immunohistochemical analysis of integrin alphavbeta3 expression on tumor-associated vessels of human carcinomas. Int J Cancer.1997; 71(3):320-324.
[68]Pozzi A, Moberg PE, Miles LA et al. Elevated matrix metallo- protease and angiostatin levels in integrin alpha1 knockout mice cause reduced tumor vascularization. Proc Natl Acad Sci USA.2000; 97(5):22022207.
[69]Kim S, Bell K, Mousa SA et al. Regulation of angiogenesis in vivo by ligation of integrin alpha5betal with the central cell-binding domain of fibronectin. Am J Pathol.2000; 156:1345-1362.
[70]Breuss JM, Sheppard D. Expression of the β6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling. J Cell Sci. 1995; 108:2241-2251.
[71]Xiaowu Li, Brian L. Schmidt, Amha Atakilit, Bing Chen, Duncan Ellis and Daniel M. αvβ6-Fyn Signaling Promotes Oral Cancer Progression. J Biol Chem. 2003; 278:(43) 41646-41653.
[72]Agrez MV, Chen A, Cone RI, Pytela R and Sheppard D. The αvβ6 integrin promotes proliferation of colon carcinoma cells through a unique region of the β6 cytoplasmic domain. J Cell Biol.1994; 127:547-556.
[73]吴小华,李冬秀,李利等.整合素对顺铂诱导的卵巢癌细胞凋亡的影响.中华妇产科杂志.2004；39(2)：112-115.
[74]Arihiro K, Kaneko M, Fujii S, Inai K and Yokosaki Y. Significance of alpha9betal and alphavbeta6 integrin expression in breast carcinoma. Breast Cancer.2000;7:19-26.
[75]Zhang ZY, Xu KS, Wang JS, et al. Integrin alphanvbeta6 acts as a prognostic indicator in gastric carcinoma. Clin Oncol (R Coll Radiol).2008 Feb; 20(1):61-66.
[1]Allum WH, Powell DJ, McConkey CC, Fielding JW. Gastric cancer:a 25-year review. Br J Surg.1989; 76:535-540.
[2]Wanebo HJ, Kennedy BJ, Chmiel J, et al. Cancer of the stomach. A patient care study by the American College of Surgeons. Ann Surg.1993; 218:583-92.
[3]Akoh JA, Macintyre IM. Improving survival in gastric cancer:review of 5-year survival rates in English language publications from 1970. Br J Surg.1992; 79:293-9.
[4]Hermanek P, Maruyama K, Sobin LH. Gastric cancer. In:Hermanek P, Gospodarowicz MK, Henson DE, Hutter RVP, Sobin LH, editors. Prognostic factors in cancer. Geneve:International Union Against Cancer (UICC); 1995
[5]Wartenberg M, Ling FC, Schallenberg M, et al. Down-regulation of intrinsic P-glycoprotein expression in multicellular prostate tumor spheroids by reactive oxygen species. J Biol Chem.2001; 276 (20):17420-17428.
[6]Ferrara N. The role of vascular endothelial growth factor in pathological angiogenesis. Breast Cancer Res. Treat.1995; 36:127-137.
[7]Dvorak HF, Brown LF, Detmar M, Dvorak AM. Vascular permeability factor/vascular endothelial growth factor, microvas- cular hyperpermeability, and angiogenesis. Am. J. Pathol.1995; 146:1029-1039.
[8]Hood JD, Frausto R, Kiosses WB, Schwartz MA, Cheresh DA. Differential alphav integrin-mediated Ras-ERK signaling during twopathways of angiogenesis. J Cell Biol.2003; 162:933-943.
[9]Smyth SS, Patterson C. Tiny dancers:the integrin-growth factor nexus in angiogenic signaling. J Cell Biol.2002; 158:17-21.
[10]Senger DR, Perruzzi CA, Streit M, Koteliansky VE, Fougerolles AR, Detmar M. The alpha(1)beta(1) and alpha(2)beta(1) integrins provide critical support for vascular endothelial growth factor signaling, endothelial cell migration, and tumor angiogenesis. Am J Pathol.2002; 160:195-204.
[11]Eliceiri BP, Puente XS, Hood JD, Stupack DG, Schlaepfer DD, Huang XZ, Sheppard D, Cheresh DA. Src-mediated coupling of focal adhesion kinase to integrin alpha(v)beta5 in vascular endothe- lial growth factor signaling. J Cell Biol.2002;157:149-159.
[12]Lee TH, Seng S, Li H, Kennel SJ, Avraham HK, Avraham S. Integrin regulation by vascular endothelial growth factor in human brain microvascular endothelial cells:role of alpha6betal integrin in angiogenesis. J Microbiol.2006; 44:502-507.
[13]Sheppard D, Rozzo C, Starr L, Quaranta V, Erle DJ, Pytela R. Complete amino acid sequence of a novel integrin beta subunit (beta6) identified in epithelial cells using the polymerase chain reaction. J Biol Chem.1990; 265:11502-11507.
[14]Breuss JM, Gallo J, DeLisser HM, Klimanskaya IV, Folkesson HG, Pittet JF, Nishimura SL, Aldape K, Landers DV, Carpenter W, et al. Expression of the beta6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling. J Cell Sci.1995; 108:2241-2251.
[15]Zhang ZY, Xu KS, He QS, Niu WB, Wang JY, Mi YT, Wang JS, Wang GQ, Yang GY, Niu J. Signaling and regulatory mechanisms of integrin alphavbeta6 on the apoptosis of colon cancer cells, Cancer Lett.2008; 266:209-215.
[16]Gu X, Niu J, Dorahy DJ, Scott R, Agrez MV. Integrin alpha(v) beta6-associated ERK2 mediates MMP-9 secretion in colon cancer cells. Brit J Cancer.2002;87: 348-351.
[17]Hynes, RO. Integrins:versatility, modulation and signaling in cell adhesion. Cell. 1992; 69:11-25.
[18]Sawhney RS, Sharma B, Humphrey LE, Brattain MG. Integrin alpha2 and extracellular signal-regulated kinase are functionally linked in highly malignant autocrine transforming growth factor-alpha-driven colon cancer cells. J Biol Chem.2003; 278(22):19861-19869.
[19]吴小华,李冬秀,李利等.整合素对顺铂诱导的卵巢癌细胞凋亡的影响.中华妇产科杂志.2004；39(2)：112-115.
[20]Arihiro K, Kaneko M, Fujii S, Inai K and Yokosaki Y. Significance of alpha9betal and alphavbeta6 integrin expression in breast carcinoma. Breast Cancer.2000; 7:19-26.
[21]Regezi JA, Ramos DM, Pytela R, Dekker NP and Jordan RCK. Tenascin and β6 integrin are overexpressed in floor of mouth in situ carcinomas and invasive squamous cell carcinomas. Oral Oncol.2002; 38:332-336.
[22]Ahmed N, Niu J, Dorahy DJ, Gu X, Andrews S, Meldrum CJ, Scott RJ, Baker MS, Macreadie IG, Agrez MV. Direct integrin alphavbeta6-ERK binding: implications for tumour growth. Oncogene.2002 Feb 21; 21(9):1370-1380.
[23]Yang GY, Xu KS, Pan ZQ, Zhang ZY, Mi YT, Wang JS, Chen R, Niu J. Integrin alphavbeta6 mediates the potential for colon cancer cells to colonize in and metastasize to the liver. Cancer Sci.2008 May; 99(5):879-887
[24]张朝阳,徐克森,杨广运等.反义整合素β6基因对结肠癌细胞作用的研究.中华医学杂志.2007；87(37)：2645-2649.
[25]Zhang ZY, Xu KS, Wang JS, Yang GY, Wang W, Wang JY, Niu WB, Liu EY, Mi YT, Niu J. Integrin alphanubeta6 Acts as a Prognostic Indicator in Gastric Carcinoma. Clin Oncol (R Coll Radiol).2008 Feb; 20(1):61-66.
[26]Xiaowu Li, Brian L. Schmidt, Amha Atakilit, Bing Chen, Duncan Ellis and Daniel M. Ramos. αvβ6-Fyn Signaling Promotes Oral Cancer Progression. J Biol Chem.2003; 278:(43) 41646-41653.
[27]Brockbank EC, Bridges J, Marshall CJ, Sahai E. Integrin betal is required for the invasive behaviour but not proliferation of squam-ous cell carcinoma cells in vivo. Br J Cancer.2005,17; 92(1):102-112.
[28]Niu J, share 1st author with Ahmed N, Dorahy DJ, Gu X, Andrews S, Meldrum CJ, Scott RJ, Baker MS, Macreadie IG, Agrez MV. Direct integrin alphavbeta 6-ERK binding:implicate- ons for tumour growth. Oncogene. 2002 Feb 21; 21(9):1370-1380.
[29]J. Wang, Z. Zhang, K. Xu, X. Sun, G. Yang, W. Niu, E. Liu, C. Peng, P. Lin, J. Wang, R. Chen, M. Agrez, J. Niu. Suppression of integrin alphaupsilonbeta6 by RNA interference in colon cancer cells inhibits extracellular matrix degradation through the MAPK pathway. Int J Cancer.2008; 123:1311-1317.
[30]Kabashima A, Maehara Y, Kakeji Y, Baba H, Koga T, Sugimachi K. Clinicopathological features and overexpression of matrix metallo- proteinases in intramucosal gastric carcinoma with lymph node metastasis. Clin Cancer Res. 2000; 9:3581-3584.
[31]Torii A, Kodera Y, Uesaka K, Hirai T, Yasui K, Morimoto T, Yamamura Y, Kato T, Hayakawa T, Fujimoto N, Kito T, Plasma concentration of matrix metalloproteinase 9 in gastric cancer. Brit J Surg.1997; 84:133-136.
[32]Petrova TV, Makinen T, Alitalo K. Signaling via vascular endothelial growth factor receptors. Exp Cell Res.1999; 253(1):117-130.
[33]刘杰,孙正义,曹蕾.血管内皮生长因子对生物衍生骨复合骨髓基质细胞体内成骨的影响.中国临床康复.2003；7(11)：1622-1623.
[34]Shibuya M. Vascular endothelial growth factor receptor-2:its unique signaling and specific ligand, VEGF-E. Cancer Sci.2003; 94:751-756.
[35]Shinkai A, Ito M, Anazawa H, Yamaguchi S, Shitara K, Shibuya M. Mapping of the sites involved in ligand association and dissociation at the extracellular domain of the kinase insert domaincontaining domaincontaining receptor for
vascular endothelial growth factor. J Biol Chem.1998; 273:31283-31288.
[36]Li B, Fuh G, Meng G, Xin X, Gerritsen ME, Cunningham B, de Vos AM. Receptor-selective variants of human vascular endothelial growth factor. Generation and characterization. J Biol Chem.2000; 275:29823-29828
[37]Gille H, Kowalski J, Li B, LeCouter J, Moffat B, Zioncheck TF, Pelletier N, Ferrara N. Analysis of biological effects and signaling properties of Flt-1 (VEGFR-1) and KDR (VEGFR-2). A reassessment using novel receptor-specific vascular endothelial growth factor mutants. J Biol Chem.2001; 276:3222-3230
[38]Larrivee B, Karsan A. Signaling pathways induced by vascular endothelial growth factor. Int J Mol Med.2000; 5:447-456.
[39]Brekken RA, Overholser JP, Stastny VA, el al. Selective inhibition of vascula endothelial growth factor(VEGF) receptor2(KDR/FIk-1) activity by a monocle-nal anti-VEGF antibody blocks tumor growth in mice. Cancer Res.2000; 60(18): 5117-5124.
[40]Pasqualini R, Koivunen E, Ruoslahti E. av Integrins as receptors for tumor targeting by circulating ligands. Nat Biotech.1997; 15:542-546.
[41]Varner JA. The role of vascular cell integrins αvβ3 and αvβ5 in angiogenesis. In: Goldberg ID, Rosen EM. Regulation of angiogenesis. Basel:Brkhauser Vrlag. 1997; 361-390.
[42]Shen JR, Yen MH, Kan YC et al. Inhibition of angiogenesis in vitro and in vivo: comparision of the relative activities of triflavin, an Arg-Gly-Asp-containing peptide and anti-alpha(v)beta3 integrin monoclonal antibody. Biochem Biophys Acta.1997; 1336(3):445-454.
[43]Herskowitz I. MAP kinase pathways in yeast:for mating and more. Cell.1995; 80(2):187-197.
[44]Widmann C, Gibson S, Jarpe M.B, et al. Mitogen-activated protein kinase: conservation of a three-kinase module from yeast to human. Physiol Rev.1999; 79(1):143-180.
[45]Chen Z, Gibson TB, Robinson F, et al. MAP kinases. Chem Rev.2001; 101(8):2449-2476.
[46]Westermarck J, Kahari VM. Regulation of matrix metalloproteinase expression in tumor invasion. FASEB J.1999 May; 13(8):781-792.
[47]EgebladM, Werb Z. New functions for the matrix metalloproteina-ses in cancer progression. Nature Rev Cancer.2002; 2(3):161-174.
[48]Johansson N, Ala-aho R, Uitto V, et al. Expression of collagenase-3 (MMP-13) and collagenase-1(MMP-1) by transformed keratinocy- tes is dependent on the activity of p38 mitogen-activated protein kinase. J Cell Sci.2000 Jan;113 Pt 2:227-235.
[49]Simon C, Hicks MJ, Nemechek AJ, Mehta R, O'Malley BW Jr, Goepfert H, Flaitz CM, Boyd D. PD 098059, an inhibitor of ERK1 activation, attenuates the in vivo invasiveness of head and neck squamous cell carcinoma. Br J Cancer. 1999 Jul; 80(9):1412-1419.
[50]Herrera R. Modulation of hepatocyte growth factor-induced scattering of HT29 colon carcinoma cells. Involvement of the MAPK pathway. J Cell Sci.1998 Apr; 111 (Pt 8):1039-1049.
[51]Badache A, Hynes NE. Interleukin 6 inhibits proliferation and, in cooperation with an epidermal growth factor receptor autocrine loop, increases migration of T47D breast cancer cells. Cancer Res.2001 Jan 1; 61(1):383-391.
[52]Tomanek RJ, Schatteman GC. Angiogenesis:new insights and ther- apeutic potential. Anat Rec.2000; 261:126-135.
[53]Bancroft CC, Chen Z, Dong G, Sunwoo JB, Yeh N, Park C, Van Waes C. Coexpression of proangiogenic factors IL-8 and VEGF by human head and neck squamous cell carcinoma involves coactive- tion by MEK-MAPK and IKK-NF-kappaB signal pathways. Clin Cancer Res.2001 Feb; 7(2):435-442.
[54]Zhou JN, Ljungdahl S, Shoshan MC, Swedenborg J, Linder S. Acti vation of tissue-factor gene expression in breast carcinoma cells by stimulation of the RAF-ERK signaling pathway. Mol Carcinog.1998 Apr; 21(4):234-43.
[55]Hulboy DL, Rudolph LA, Matrisian LM. Matrix metalloproteinases as mediators of reporductive function. Molecular Human Reprodu-ction.1997; 3:27-45.
[56]Birkedal-Hansen H, Moore WGL, Bodden MK, Windor LJ, Birke- dal- Hansen
B, DeCarl A, et al. Matrix metalloproteinase:a review. Crit Rev Oral Biol Med. 1993; 4:197-250.
[57]Shapiro SD. Matrix metalloproteinase degradation of extracelluar matrix:biolog-ical consequence. Curr Opin Cell Biol.1998; 10:602-603.
[58]Nagase H. Activation mechanisms of matrix metalloproteinase. Biol Chem Hoppe Seyler.1997; 378:151-160.
[59]Nagase H,Woessner JF. Matrix metalloproteinase. J Boil Chem 1999; 274: 21491-21494.
[60]Vu TH, Werb Z. Matrix metalloproteinase:effectors of development and normal physiology. Gences Dev.2000; 14:2123-2133.
[61]Dis S, et al.IL-12 regulates VEGF and MMPs in a murine breast cancer model. Int J Cancer.1998 Oct 29;78(3):361-365.
[62]Gum R, Lengyel E, Juarez J, Chen JH, Sato H, Seiki M, Boyd D. Stimulation of 92-kDa gelatinase B promoter activity by ras is mitogen-activated protein kinase kinase 1-independent and requires multiple transcription factor binding sites including closely spaced PEA3/ets and AP-1 sequences. J Biol Chem.1996 May 3; 271(18):10672-10680.
[63]Van Wart H, Birkedal-Hansen H. The cystein switch:a principle of regulation of metalloproteinase activity with potential applicability to the entire matrix metalloproteinase gene family. Proc Natl Acad Sci USA.1990; 87:5578-5582.
[64]Mignatti P, Rifkin DB. Plasminogen activators and matrix metallo-proteinases in angiogenesis. Enzyme Proein.1996; 49:117-137.
[65]DeCLerck YA, Laug WE. Plasminogen activators and their inhibit- tors: cooperation between matrix metalloproteinases and the plasminogen activator-plasmin system in tumor progression. Enzyme Protein.1996; 49:72-84
[66]Rosenthal EL, Johson TM, Allen ED, Apel IJ, Punturieri A, Weiss SJ. Role of plasminogen activator and matrix metalloproteinase systems in epidermal growth factor and scatter factor-stimulated invasion of carcinoma cells. Cancer Res. 1998; 58:5221-5230.
[67]Deryugina EI, Zijlstra A, Partridge JJ, Kupriyanova TA, Madsen MA,
Papagiannakopoulos T, Quigley JP. Unexpected effect of matrix metallo-proteinase down-regulation on vascular intravasation and metastasis of human fibrosarcoma cells selected in vivo for high rates of dissemination. Cancer Res. 2005 Dec 1;65(23):10959-10969.
[68]Cox G, O'Byrne KJ. Matrix metalloproteinases and cancer. Anticancer Research. 2001; 21:4207-4220.
[69]Liotta L. Isolation of a protein that stimulates blood vessel growth. Nature,1985, 318(6041):14.
[70]Ricca A, Biroccio A, Bufalo DD, et al. bcl-2 over-expression enhances NF-kappaB activity and induces mmp-9 transcription in human MCF7(ADR) breast-cancer cells. Int J Cancer.2000; 86(2):188-196.
[71]Shalinsky DR, Brekken J, Zou H, et al. Broad Antitumor and Antiangiogenic Activities of AG3340, a Potent and Selective MMP Inhibitor Undergoing Advanced Oncology Clinical Trials. Ann NY Acad Sci.1999; 878:236-270.
[72]Shalinsky DR, Brekken J, Zou H, et al. Marked Antiangiogenic and Antitumor Efficacy of AG3340 in Chemoresistant Human Non-Small Cell Lung Cancer Tumors:Single Agent and Combination Chemotherapy Studies. Clin Cancer Res. 1999; 5:1905-1917.
[73]Patterson BC, Amy Sang QX. Angiostatin-converting Enzyme Activities of Human Matrilysin (MMP-7) and Gelatinase B/Type IV Collagenase (MMP-9). J Biol Chem.1997; 272:28823-28825.
[74]Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC. Potent and specific genetic interference by double-strand RNA in Caenorhabditis elegans Nature.1998; 391(6669):744-745.
[75]McManus MT, Sharp PA. Gene silencing in mammals by small interfering RNAs. Nature.2002; 3(10):737-747.
[76]Hannon GJ. RNA interference. Nature.2002; 418 (6894):244-251.
[77]Zamore PD, Tuschl T, Sharp PA, Bartel DP. RNAi:double-strand RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell. 2000; 101:25-33.
[78]Caplen NJ, Parrish S, Imani F, Fire A, Morgan RA. Specific inhibition of gene expression by small double-stranded RNAs in invertebrate and vertebrate systems. Proc Natl Acad Sci USA.2001; 98(17):9742-9747.
[79]Nykanen A, Haley B, Zamore PD. ATP requirements and small interferings RNA structure in the RNA interference pathway. Cell.2001; 107(3):309-321.
[80]Gil J, Esteban M. Induction of apoptosis by the dsRNA dependent protein kinase (PKR):mechanism of action. Apoptosis.2000; 5 (2):107-114.
[1]黄志强,周宁新,黄晓强.肝门部胆管癌的外科治疗.中华消化外科杂志.2003；4：
[2]周宁新,黄志强,刘永雄.肝外胆道癌全国调查1098例分析.中华外科杂志.1990；28：516-520.
[3]黄志强.肝门部胆管癌的外科治疗.中华外科杂志.1990；28：522-524.
[4]Gebbia V, Majello E, Testa A, et al.Treatment of advanced adenocar- cinoma of the exocrine pancreas and the gallbladder with 5-Fu, high dose levofolinic acid and oral hydroxyurea on weekly schedule, Results of a multicenter study of the southern Italy Incology Group (G.O.I.M). Cancer.1996;78:1300-1307.
[5]黄志强.肝门部胆管癌外科治疗的现状与展望.中国普外基础与临床杂志.2005：12：317-320.
[6]Volm M, Koomagi R, Mattern J, et al. PD-ECGF, b-FGF, and VEGF expression in non small cell lung carcinomas and their association with lymph node metastasis. Anticancer Res.1999;19:65-112.
[7]Maehara Y, Kabashima A, Koga T, et al. Vascular invasion and potential for tumor angiogenesis and metastasis in gastric carcinoma. Surgery.2000;128:408.
[8]Fujisaki K, Mitsuyama K, Toyonaga A, et al. Circulating vascular endothelial growth factor in patients with colorectal cancer. Am J Gastroenterol.1998; 93: 249.
[9]Jeffrey E, Isaish J. Targeting Lymphatic Metastasis. Science.2002; 296: 1811-1812.
[10]Lillemoe KD, Cameron JL. Surgery for hilar cholangiocarcinoma:the Johns Hopkins approach. J Hepatobiliary Pancreat Surg.2000; 7:115-121.
[11]周宁新.肝门部胆管癌的外科治疗新进展.中国医师进修杂志.2006；29：6-7.
[12]Imai M, Hoshi T, Ogawa K. K-ras codon 12 mutations biliary tract tumors detected by polymerase chain reaction denaturing gradient gel electrophoresis. Cancer.2001;73:2724-2733.
[13]Joukov V, Pajusola K, Kaipainen A, et al. A novel vascular endothelial growth factor, VEGF-C, is a ligand for the Flt4 (VEGFR-3) and KDR (VEGFR-2) receptor tyrosine kinases. EMBO J.1996;15:290-298.
[14]Li XR, Eriksson U. Novel VEGF family members:VEGF-B, VEGF-C and VEGF-D. Int J Biochem Cell Biol.2001; 33:421-426.
[15]Pan J, Hua C, Wen Y. Cloning and sequencing of human vascular endothelial growth factor-C encoded cDNA. Hua Xi Kou Qiang Yi Xue Za Zhi.2003; 21(4): 318-320.
[16]Dixelius J, Makinen T, Wirzenius M, et al. Ligand induced vascular endothelial growth factor receptor-3 (VEGFR-3) hetero-dimerization with VEGFR-2 in primary lymphatic endothelial cells regulates tyrosine phosphorylation sites. J Biol Chem.2003; 78 (42):40973-40979.
[17]Meyer RD, Rahimi N. Comparative structure function analysis of VEGFR-1 and VEGFR-2:What have we learned from chimeric systems. Ann N Y Acad Sci. 2003; (995):200-207.
[18]Ryden L, Linderholm B, Nielsen NH, et al.Tumor specific VEGF-A and VEGFR2/KDR protein are coexpressed in breast cancer. Breast Cancer Res Treat. 2003; 82(3):147-154.
[19]Nisato RE, Tille JC, Pepper MS. Lymphangiogenesis and tumor metastasis. Thromb Haemost.2003; 90(4):591-597.
[20]Skobe M, Hamberg LM, Hawighorst T, et al. Concurrent induction of lymphan-giogenesis, angiogenesis, and maemphage recruitment by vascular endothelial growth factor-C in melanoma. Am J Pathol.2001; 159:893-903.
[21]Mandriota SJ, Jussila L, Jeltsch M, et al. Vascular endothelial growth factor-C mediated lymphangiogenesis promotes tumor metastasis. EMBO J.2001; 20:672-682.
[22]Stacker SA, Caesar C, Baldwin ME, et al. VEGF-D promotes the metastatic spmad of tumor cells via the lymphatics. Nat Med.2001; 7:186-191.
[23]Von Marsehall Z, Scholz A, Stacker SA, et al. Vascular endothelial growth factor-D induces lymphangiogenesis and lymphatic-metastasis in models of ductal pancreatic cancer. Int J Oncol.2005; 27:669-679.
[24]Schietroma C, Cianfarani F, Lacal PM,et al. Vascular endothelial growth factor-C expression correlates with lymph node localizat-ion of human melanoma metastases. Cancer.2003; 98(4):789-797.
[25]Hung CJ, Ginzinger DG, Zarnegar R, et al. Expression of vascular endothelial growth factor-C in benign and malignant thyroid tumors. J Clin Endocrinol Metab.2003; 88(8):3694-3699.
[26]Liu F, Zhang YJ. Roles of VEGF-C and its receptor Flt-4 in proliferation and metastasis of primary breast cancer. Ai Zheng.2003; 22(10):1053-1056.
[27]Kimura Y, Watanabe M, Ohga T, et al. Vascular endothelial growth factor-C expression correlates with lymphatic involvement and poor prognosis in patients with esophageal squamous cell carcinoma. Oncol Rep.2003; 10(6):1747-1751.
[28]Maeda K, Yashiro M, Nishihara T, et al. Correlation between vascular endothelial growth factor-C expression and lymph node metastasis in Tl carcinoma of the colon and rectum. Surg Today.2003; 33(10):736-739.
[29]Tamura M, Ohta Y. Serum vascular endothelial growth factor-C level in patients with primary nonsmall cell lung carcinoma:a possible diagnostic tool for lymph node metastasis. Cancer.2003; 98(6):1217-1222.
[30]Sedivy R, Mannagetta J, Haverkampf C, et al. Expression of vascular endothelial growth factor-C correlates with the lymphatic microvessel density and the nodal status in oral squamous cell cancer. J Oral Pathol Med.2003; 32(8):455-460.
[31]Ogawa E, Takenaka K, Yanagihara K, et al. Clinical significance of VEGF-C status in tumour cells and stromal macrophages in non-small cell lung cancer patients. Br J Cancer.2004; 91:498-503.
[32]Skobe M, Hawighorst T, Jackson DG, et al. Induction of tumor lymphan-giogenesis by VEGF-C promotes breast cancer metastasis. Nat Med.2001; 7(2):192-198.
[33]Partanen TA, Paavonen K. Lymphatic versus blood vascular endothelial growth factors and receptors in humans. Microsc Res Tech.2001; 55(2):108-121.
[34]Karpanen T, Egeblad M, Karkkainen MJ, et al. Vascular endothelial growth factor-C promotestumor lymphangiogenesis and intralym-phatic tumor growth. Cancer Res.2001; 61(5):1786-1790.
[35]张萌,彭利,王顺祥等.VEGF与VEGF-C蛋白在胆管癌中的表达及意义.山东医药.2007；6：17-18.
[36]Karpanen T, Alitalo K. Lymphatic vessels as targets of tumor therapy. J Exp Med. 2001; 194(6):37-42.
[37]Hamada K, Oike Y, Takakura N, et al. VEGF-C signaling pathways through VEGFR-2 and VEGFR-3 in vasculoangiogenesis and hematopoiesis. Blood.
2000; 96(12):3793-3800.
[38]Valtola R, Salven P, Heikkila P. VEGFR-3 and its ligand VEGF-C are associated with angiogenesis in breast cancer. Am J Pathol.1999; 154(5):1381-1390.
[39]Zachary I. Signaling mechanisms mediating vascular protective actions of vascular endothelial growth factor. Am J Physiol Cell Physiol.2001; 280(6): 1375-1386.
[40]Nathanson SD. Insights into the mechanism of lymph node metastasis. Cancer. 2003;98(2):413-423.
[41]Jeltsch M, Kaipainen A, Joukov V, et al. Hyperplasia of lymphatic vessels in VEGF-C transgenic mice. Science.1997; 276 (5317):1423-1425.
[42]Ueda M, Teraj Y, Yamashita Y, et al. Correlation between vascular endothelial growth factor-C expression and invasion phenotype in cervical carcinomas. Int J Cancer.2002; 98:335-343.
[43]George ML, Tutton MG, Janssen F, et al. VEGF-A, VEGF-C and VEGF-D in colorectal cancer progression. Neoplasia.2001; 3:420-427.
[44]常实,汤恢焕,周军等.肝外胆管癌nm23和VEGF的表达及其临床意义.中国普通外科杂志.2003；12(18)：625-627.
[45]Benckert C, Sven Jonas, Cramer T, et al. Transforming growth factor beta-1 stimulates vascular endothelial growth factor gene transcription in human cholangiocellular carcinoma cells. Cancer Research.2003; 63(5):1083-1092.
[1]L.J. Van't Veer, B. Weigelt, Road map to metastasis, Nat. Med.9 (2003) 999-1000.
[2]D. Hanahan, R.A. Weinberg, The hallmarks of cancer, Cell 100 (2000) 57-70.
[3]N. Ferrara, The role of vascular endothelial growth factor in pathological angiogenesis, Breast. Cancer. Res. Treat.36 (1995) 127-137.
[4]H.F. Dvorak, L.F. Brown, M. Detmar, A.M. Dvorak, Vascular permeability factor/vascular endothelial growth factor, microvascular hyperpermeability, and angiogenesis, Am. J. Pathol.146 (1995) 1029-1039.
[5]J.D. Hood, R. Frausto, W.B. Kiosses, M.A. Schwartz, D.A. Cheresh, Differential alphav integrin-mediated Ras-ERK signaling during two pathways of angiogenesis, J. Cell Biol.162 (2003) 933-943.
[6]S.S. Smyth, C. Patterson, Tiny dancers:the integrin-growth factor nexus in angiogenic signaling, J. Cell Biol.158 (2002) 17-21.
[7]R.O. Hynes, Integrins:versatility, modulation, and signaling in cell adhesion, Cell 69(1992)11-25.
[8]R.O. Hynes, Integrins:bidirectional, allosteric signaling machines, Cell 110 (2002) 673-687.
[9]J.M. Breuss, N. Gillett, L. Lu, D. Sheppard and R. Pyetala, Restricted distribution of integrin beta 6 mRNA in primate epithelial tissues, J. Histochem. Cytochem. 41 (1993)1521-1527.
[10]Z.Y. Zhang, K.S. Xu, J.S. Wang, GY. Yang, W. Wang, J.Y. Wang, W.B. Niu, E.Y. Liu, Y.T. Mi, J. Niu, Integrin alphanvbeta6 acts as a prognostic indicator in gastric carcinoma, Clin. Oncol.20 (2008) 61-66.
[11]G.Y. Yang, K.S. Xu, Z.Q. Pan, Z.Y. Zhang, Y.T. Mi, J.S. Wang, R. Chen, J. Niu, Integrin alphavbeta6 mediates the potential for colon cancer cells to colonize in and metastasize to the liver, Cancer. Sci.99 (2008) 879-887.
[12]J. Wang, Z. Zhang, K. Xu, X. Sun, G. Yang, W. Niu, E. Liu, C. Peng, P. Lin, J. Wang, R. Chen, M. Agrez, J. Niu, Suppression of integrin alphaupsilonbeta6 by RNA interference in colon cancer cells inhibits extracellular matrix degradation through the MAPK pathway, Int. J. Cancer.123 (2008) 1311-1317.
[13]A. Kabashima, Y. Maehara, Y. Kakeji, H. Baba, T. Koga, K. Sugimachi, Clinicopathological features and overexpression of matrix metalloproteinases in intramucosal gastric carcinoma with lymph node metastasis, Clin. Cancer. Res. 9(2000) 3581-3584.
[14]A. Torii, Y. Kodera, K. Uesaka, T. Hirai, K. Yasui, T. Morimoto, Y. Yamamura, T. Kato, T. Hayakawa, N. Fujimoto, T. Kito, Plasma concentration of matrix metalloproteinase 9 in gastric cancer, Br. J. Surg.84(1997) 133-136.
[15]D.R. Senger, C.A. Perruzzi, M. Streit, V.E. Koteliansky, A.R. Fougerolles, M. Detmar, The alpha(1)beta(1) and alpha(2)beta(1) integrins provide critical support for vascular endothelial growth factor signaling, endothelial cell migration, and tumor angiogenesis, Am. J. Pathol.160 (2002) 195-204.
[16]K. Podar, Y.T. Tai, B.K. Lin, R.P. Narsimhan, M. Sattler, T. Kijima, R. Salgia, D. Gupta, D. Chauhan, K.C. Anderson, Vascular endothelial growth factor-induced migration of multiple myeloma cells is associated with betal integrin-and phosphatidylinositol 3-kinase-dependent PKC alpha activation, J. Biol. Chem. 277(2002)7875-7881.
[17]N.E. Vlahakis, B.A. Young, A. Atakilit, A.E. Hawkridge, R.B. Issaka, N. Boudreau, D. Sheppard, Integrin alpha9betal directly binds to vascular endothelial growth factor (VEGF)-A and contributes to VEGF-A-induced angiogenesis, J. Biol. Chem.282 (2007) 15187-15196.
[18]D. Sheppard, C. Rozzo, L. Starr, V. Quaranta, D.J. Erle, R. Pytela, Complete amino acid sequence of a novel integrin beta subunit (beta 6) identified in epithelial cells using the polymerase chain reaction, J. Biol. Chem.265 (1990) 11502-11507.
[19]J.M. Breuss, J. Gallo, H.M. DeLisser, I.V. Klimanskaya, H.G. Folkesson, J.F. Pittet, S.L. Nishimura, K. Aldape, D.V. Landers, W. Carpenter, et al, Expression of the beta 6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling, J. Cell. Sci.108 (1995) 2241-2251.
[20]Z.Y. Zhang, K.S. Xu, Q.S. He, W.B. Niu, J.Y. Wang, Y.T. Mi, J.S. Wang, G.Q. Wang, G.Y. Yang, J. Niu, Signaling and regulatory mechanisms of integrin alphavbeta6 on the apoptosis of colon cancer cells, Cancer. Lett.266 (2008) 209-215.
[21]X. Gu, J. Niu, D.J. Dorahy, R. Scott, M.V. Agrez, Integrin alpha(v)beta6-associated ERK2 mediates MMP-9 secretion in colon cancer cells, Br. J. Cancer. 87(2002)348-351.
[22]B.P. Eliceiri, X.S. Puente, J.D. Hood, D.G. Stupack, D.D. Schlaepfer, X.Z. Huang, D. Sheppard, D. A. Cheresh, Src-mediated coupling of focal adhesion kinase to integrin alpha(v)beta5 in vascular endothelial growth factor signaling, J. Cell. Biol.157(2002)149-159.
[23]T.H. Lee, S. Seng, H. Li, S.J. Kennel, H.K. Avraham, S. Avraham, Integrin regulation by vascular endothelial growth factor in human brain microvascular endothelial cells:role of alpha6betal integrin in angiogenesis, J. Microbiol.44 (2006) 502-507.
[24]K. Milde-Langosch, A.M. Bamberger, G. Rieck, D. Grund, G. Hemminger, V. Muller, T. Loning, Expression and prognostic relevance of activated extracellular-regulated kinases (ERK1/2) in breast cancer, Br. J. Cancer.92 (2005)2206-2215.
[25]A.A. Adjei, The role of mitogen-activated ERK-kinase inhibitors in lung cancer therapy, Clin. Lung. Cancer.7 (2005) 221-223.
[26]N. Ahmed, C. Riley, G.E. Rice, M.A. Quinn, M.S. Baker. Alpha(v)beta(6) integrin-A marker for the malignant potential of epithelial ovarian cancer, J. Histochem. Cytochem.50 (2002) 1371-1380.
[27]J.S. Krueger, V.G. Keshamouni, N. Atanaskova, K.B. Reddy, Temporal and quantitative regulation of mitogen-activated protein kinase (MAPK) modulates cell motility and invasion, Oncogene 20 (2001) 4209-4218.
[28]N. Ahmed, F. Pansino, M. Baker, G. Rice, M. Quinn, Association between alphavbeta6 integrin expression, elevated p42/44 kDa MAPK, and plasminogen-dependent matrix degradation in ovarian cancer, J. Cell. Biochem. 84 (2002) 675-686.
[29]N. Ahmed, J. Niu, D.J. Dorahy, X. Gu, S. Andrews, C.J. Meldrum, R.J. Scott, M.S. Baker, I.G. Macreadie, M.V. Agrez, Direct integrin alphavbeta6-ERK binding:implications for tumor growth, Oncogene 21(2002) 1370-1380.
[30]J. Niu, D.J. Dorahy, X. Gu, R.J. Scott, B. Draganic, N. Ahmed, M.V. Agrez, Integrin expression in colon cancer cells is regulated by the cytoplasmic domain of the beta6 integrin subunit, Int. J. Cancer.99 (2002) 529-537.
[1]Patel T. Increasing incidence and mortality of primary intrahepatic cholangiocar-cinoma in the United States. Hepatology 2001; 33:1353-1357.
[2]Gores GJ. Cholangiocarcinoma:current concepts and insights. Hepatology 2003; 37:961-969.
[3]N. Ferrara. The role of vascular endothelial growth factor in pathological angio-genesis. Breast Cancer Res Treat.1995; 36:127-137.
[4]H.F. Dvorak, L.F. Brown, M. Detmar, A.M. Dvorak, Vascular permeability factor/vascular endothelial growth factor, microvascular hyperpermeability, and angiogenesis. Am J Pathol.1995; 146:1029-1039.
[5]Salven P, Lymboussaki A, Heikkila P. Vascular endothelial growth factors VEGF-B and VEGF-C are expressed in human tumors. Am J Pathol 1998; 153:103-108.
[6]O-charoenrat P, Rhys-Evans P, Eccles SA. Expression of vascular endothelial growth factor family members in head and neck squamous cell carcinoma correlates with lymph node metastasis. Cancer.2001; 92:556-65.
[7]Tsurusaki T, Kanda S, Sakai H. Vascular endothelial growth factor-C expression in human prostatic carcinoma and its relationship to lymph node metastasis. Br J Cancer.1999; 80:309-313.
[8]George ML, Tutton MG, Janssen F, Arnaout A, Abulafi AM, Eccles SA, et al. VEGF-A, VEGF-C and VEGF-D in colorectal cancer progression. Neoplasia. 2001; 3:420-7.
[9]Makoto I, Joji K, Shinsuke K and Hirokazu N. Expression of Vascular Endothelial Growth Factor C and D (VEGF-C and -D) is an Important Risk Factor for Lymphatic Metastasis in Undifferentiated Early Gastric Carcinoma. Jpn J Clin Oncol.2003; 33(1):21-27.
[10]Yonemura Y, Endo Y, Fujita H, Fushida S et al. Role of vascular endothelial growth factor C expression in the development of lymph node metastasis in gastric cancer. Clin Cancer Res.1999; 5:1823-1829.
[11]Zhao R, Liu XQ, Wu XP, et al. Vascular endothelial growth factor (VEGF) enhances gastric carcinoma invasiveness via integrin alpha(v)beta6. Cancer Lett. 2010 Jan 28; 287(2):150-156.
[12]Chang Y, Wu XY. JNK1/2 siRNA inhibits transforming-growth factor-betal-induced connective tissue growth factor expression and fibrotic function in THSFs. Mol Cell Biochem.2010 Feb; 335(1-2):83-89.
[13]Karkkainen MJ, Petrova TV. Vascular endothelial growth factor receptors in the regulation of angiogenesis and lymphangiogenesis. Oncogene.2000; 19:5598-5605.
[14]Enholm B, Paavonen K, Ristimaki A. Comparison of VEGF, VEGF-B, VEGF-C and Ang-1 mRNA regulation by serum, growth factors, oncoproteins and hypoxia. Oncogene.1997; 14:2475-2783.
[15]Joukov V, Pajusola K, Kaipainen A, Chilov D, Lahtinen I, Kukk E, et al. A novel vascular endothelial growth factor, VEGF-C, is a ligand for the Flt4 (VEGFR-3)
and KDR (VEGFR-2) receptor tyrosine kinases [erratum]. EMBO J.1996; 15: 1751.
[16]Skobe M, Hawighorst T, Jackson DG, et al.Induction of tumor lymphangio-genesis by VEGF-C promotes breast cancer metastasis. Nature Med.2001; 7: 192-198.
[17]Masatsugu Ueda, Yao-Ching Hung, YoshitoTerai,et al.Vascular Endothelial Growth Factor-CExpression and Invasive Phenotype in Ovarian Carcinomas. Clin Cancer Res.2005; 11(9):3225-3232.
[18]VanTrappen PO, Ryan A, CarrollM, et al. A model for co-expression pattern analysis of genes implicated in angiogenesis and tumour cell invasion in cervical cancer. Br J Cancer.2002; 87:537-544.