胸腺素β4对膀胱癌上皮—间质转化调控作用的实验研究
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摘要
第一部分胸腺素β4与人膀胱移行细胞癌上皮间质转化的关系
目的：探讨人胸腺素β4与膀胱移行上皮癌(BTCC)上皮-间质转化的相关性,以及与临床病理特征之间的关系。方法：选用膀胱移行细胞癌手术切除标本60例,每例标本均选用癌组织中心、癌旁组织及其远端正常黏膜对照。采用定量RT-PCR、Western blot和免疫组织化学技术检测切除标本中的Tβ4、ILK、E-cadherin和β-catenin的表达,并分析与各临床病理因素之间的关系。结果：(1)Tβ4在BTCC组织中的表达明显高于正常黏膜(91.7% vs 15.0%,P<0.05),其在深层浸润组中的表达高于浅层浸润组(97.4%vs 81%,P<0.05)；ILK在BTCC组织中的表达明显高于正常黏膜(80.0%vs18.3%,P<0.05)；β-catenin在BTCC组织中的表达高于正常黏膜(76.7% vs 26.7%,P<0.05)；E-cadherin在BTCC组织中的表达明显低于正常黏膜(30.0% vs 90.0%,P<0.05)；(2)Tβ4与E-cadherin表达呈负相关,与ILK、β-catenin表达呈正相关。结论：胸腺素β4与膀胱移行上皮癌分化和转移有关,可能通过调控上皮-间质转化从而在膀胱移行上皮癌的侵袭转移过程中发挥作用。
第二部分基因沉默胸腺素β4对膀胱移行细胞癌上皮间质转化的逆转作用
目的：探讨胸腺素β4基因沉默对膀胱癌细胞EMT的逆转,进一步研究其在肿瘤侵袭转移中的作用机制。方法：使用针对Tβ4的慢病毒载体(Lenti-Tβ4)转染膀胱癌细胞株T24。采用定量RT-PCR、Western blot法检测Tβ4、ILK和上皮性标记基因E-cadherin、β-catenin的表达改变；Immunofluorescence法检测ILK、E-cadherin、β-catenin在转染后T24细胞中的表达改变；细胞划痕实验、Boyden小室体外侵袭实验、AO/EB荧光染色法反映细胞转移潜能和凋亡的变化。结果：转染后的T24细胞,Tβ4、ILK和β-catenin基因或蛋白表达下调,而上皮标记基因E-cadherin表达增强(p<0.05)；Immunofluorescence显示ILK和β-catenin在细胞质、细胞核中表达减弱,而E-cadherin在胞膜中表达增强,细胞形态向正常上皮细胞转化；转染后的T24细胞体外迁移能力与侵袭力下降,细胞凋亡增多。结论：肿瘤细胞中的Tβ4表达下调可以逆转肿瘤细胞的间质表型,而向正常上皮表型转化,降低肿瘤转移潜能。
第三部分Tβ4-siRNA慢病毒载体逆转膀胱癌细胞上皮间质转化的体内实验研究
目的：构建可荧光示踪的膀胱癌小鼠皮下移植瘤模型及肺转移模型,并探讨活体内抑制Tβ4基因表达对膀胱癌生长侵袭、转移及血管形成的影响,为后续研究奠定基础。方法：(1)选用膀胱移行细胞癌细胞株EJ,体外扩增培养制成细胞悬液后注射于BALB/C小鼠颈背部皮下,待有肉眼可见瘤块后,处死小鼠并取出肿瘤组织。对肿瘤组织进行病理组织学观察,免疫组化方法检测Tβ4、ILK、E-cadherin和β-catenin在移植瘤中的表达,TUNEL方法检测活体内抑制Tβ4基因表达对膀胱癌细胞的促凋亡作用,体外血管形成实验检测Tβ4表达下调对血管生成的影响。(2)将膀胱移行细胞癌细胞株EJ悬液接种于BALB/C小鼠尾静脉内,于术后第20天处死小鼠,观察胸腔及肺内转移灶大小、数量,观察生存天数,记录远处转移情况。结果：(1)EJ细胞以慢病毒Lenti-Tβ4转染后形成的膀胱癌移植瘤与EJ细胞慢病毒Lenti-GFP转染(阴性对照)后形成的移植瘤比较,移植瘤体积、湿重、小鼠平均生存天数均存在差异,差异有统计学意义(P<0.05)；移植瘤组织HE染色发现,干扰移植瘤中Tβ4基因表达,癌组织恶性程度相较于未干扰组低；Lenti-Tβ4组中Tβ4、ILK和β-catenin阳性表达率明显低于阴性对照组,而E-cadherin阳性表达率明显高于阴性对照组；Lenti-Tβ4组凋亡率明显高于阴性对照组；与空白对照组、阴性对照组相比,Lenti-Tβ4转染组促进血管形成能力下降66.8%,有统计学意义(P<0.01)。(2)BALB/C新生仔鼠尾静脉注入膀胱癌细胞EJ后,20d后解剖取出肺组织,阴性对照组肺组织表面不光滑,肉眼见肺表面弥漫性斑块,最大直径1.5 cm,而Lenti-Tβ4转染组肺组织中转移灶明显变小或未见明显转移灶,各组小鼠的肝脏、脾脏、心脏、骨均未发现明显转移。结论：携带干扰序列Tβ4-siRNA的慢病毒能有效地抑制Tβ4基因的表达,从而抑制膀胱癌细胞体内的侵袭能力,同时发现Tβ4基因表达与膀胱癌细胞的转移活性及血管生成相关,下调其表达水平能够明显抑制癌细胞的转移能力。
Part 1 Correlation between thymosinβ4 and epithelial-mesenchymal transition in human bladder transitional cell carcinoma
Objective:To investigate the relationship between human thymosinβ4 (T(34) and epithelial-mesenchymal transition (EMT) in bladder transitional cell carcinoma (BTCC), explore the association with clinicopathologic features. Methods:60 BTCC specimens from surgically resected were selected. For each specimen, the cancerous tissue、peri-cancer tissue and its remote normal mucosa were analyzed and compared. The expression of Tβ4、ILK、E-cadherin andβ-catenin proteins in resected cancer tissues were detected by using Real-time RT-PCR、Western blot and immunohistochemistry, and compared with clinicopathologic data. Results:(1) The expression rate of Tβ4 was significantly higher in BTCC tissues than in normal tissues (91.7% vs 15.0%, P＜0.05), and the expression rate of Tβ4 in deeply infiltrating group was significantly higher than that in superficially infiltrating group(97.4% vs 81%, P＜0.05); the level of ILK was significantly higher in BTCC than in normal tissues(80.0% vs 18.3%, P＜0.05); the expression rate ofβ-catenin was higher in BTCC tissues than in normal tissues(76.7% vs 26.7%, P＜0.05); the expression of E-cadherin in BTCC tissues were significantly lowered in comparison with those in normal bladder epithelium(30.0% vs 90.0%, P＜0.05); (2) Tβ4 expression was inversely correlated to E-cadherin, but positively to ILK andβ-catenin. Conclusion: Thymosinβ4 is related to the differentiation and metastasis of bladder transitional cell carcinoma, and may induce EMT to promote BTCC metastasis.
Part 2 The reverse effect of thymosinβ4 gene silencing on EMT in human bladder transitional cell carcinoma
Objective:To investigate the reverse effect of epithelial-mesenchymal transition (EMT) by silencing human gene thymosinβ4(Tβ4), and further reseach for its role on invasion and metastasis of cancer. Methods:Lentiviral shRNA vector encoding Tβ4 was transfected into T24 cell line. The expression of Tβ4、ILK and epithelial markers E-cadherin、β-catenin were detected by Real-time RT-PCR and Western blot assay; the expression of ILK、E-cadherin andβ-catenin in T24 cells were detected by immunofluorescence after transfection via lentiviral vector; the metastatic potential and apoptosis were examined by in vitro cell wound model、Boyden chamber invasion assay and acridine orange-ethidium bromide fluorescent staining. Results:The expression of Tβ4、ILK andβ-catenin were decreased in the T24 cells after transfected with Tβ4 shRNA, and the expression of E-cadherin was significantly up-regulated (p<0.05); the level of ILK in cytoplasm and level ofβ-catenin in nuclear were obviously decreased by immunofluorescence analysis, and the higher level of E-cadherin was observed, the morphology of T24 Cells was transformed into a normal epithelial phenotype; the motility、invation of T24 cells were inhibited and apoptosis was enhanced. Conclusion:The silencing of Tβ4 may reverse fibroblastoid morphology into a normal epithelial phenotype, and suppress tumor metastatic potential.
Part 3 The study on reversion of epithelial-mesenchymal transition (EMT) in bladder transitional cell carcinoma (BTCC) by using lentivirus-mediated Tp4-siRNA in vivo
Objective:To establish xenografted tumor and lung metastasis BALB/C mouse models of bladder transitional cell carcinoma (BTCC) expressing green fluorescent protein, and investigate effect of Tumor Growth, Metastasis and Apoptosis in vivo by inhibiting Tβ4 expression. so as to lay a foundation for future study. Methods:(1)EJ cell line was used in the present study. The cells were proliferated and the cell suspension was subcutaneously injected into the BALB/C mice. Histologic and immunohistochemical analysis were performed for Tβ4, ILK,β-catenin and E-cadherin. The formed tumor cells apoptosis was detected by TUNEL in the BALB/c nude mice and images were taken using light microscopy. Dark-brown apoptotic bodies were located in nucleus. (2)The in situ tumor information, lymphatic and pulmonary metastasis were all recorded in detail. EJ cells transfected with lenti-GFP (Cont) or lenti-Tβ4 shRNA (lenti-Tβ4) were injected into the tail veins of mice (20 mice per group), and the mice were killed 20 days after injection. The number of metastatic lung nodules was determined by direct counting of the nodules on the lung. Dark arrows indicate the nodules on the lung. Quantification of antiangiogenic activity was calculated by measuring the length of tube walls formed between discrete endothelial cells in each well. Results:(1)Note the E-cadherin stain in the adjacent epidermis of lenti-Tβ4 group xenografts. Note the cytoplasmic staining of the Tβ4 patterns, while ILK showed a corresponding pattern. This staining pattern stands in sharp contrast to the expression profile of E-cadherin, which was absent in the majority of cases in lenti-Tβ4 group, while positivity in staining was observed in control group. (2)The lung metastasis was observed in BALB/C mice, two weeks after mice were injected intravenously with lentivirus-infected cells, the mean number of metastatic lung nodules was 2 for mice injected with EJ cells infected with Tβ4 shRNA and 6 for mice injected with EJ cells infected with control lentivirus (P<0.05). Extensive tube formation of endothelial cells was observed in non-transfection and lenti-GFP (Cont) groups. However, when the endothelial cells were treated by the medium preconditioned with Tβ4 shRNA-transfected EJ cells, the tube formation was markedly suppressed. Conclusion:These data suggested that overexpression of Tβ4 was associated with increases in primary tumor growth and in the number of lung metastases, and transfection of Tβ4 shRNA remarkably inhibited cancer cell malign differentiation in vivo and decreased the angiogenesis of urothelial carcinoma cells in vitro.
目的：探讨人胸腺素β4与膀胱移行上皮癌(BTCC)上皮-间质转化的相关性,以及与临床病理特征之间的关系。方法：选用膀胱移行细胞癌手术切除标本60例,每例标本均选用癌组织中心、癌旁组织及其远端正常黏膜对照。采用定量RT-PCR、Western blot和免疫组织化学技术检测切除标本中的Tβ4、ILK、E-cadherin和β-catenin的表达,并分析与各临床病理因素之间的关系。结果：(1)Tβ4在BTCC组织中的表达明显高于正常黏膜(91.7% vs 15.0%,P<0.05),其在深层浸润组中的表达高于浅层浸润组(97.4%vs 81%,P<0.05)；ILK在BTCC组织中的表达明显高于正常黏膜(80.0%vs18.3%,P<0.05)；β-catenin在BTCC组织中的表达高于正常黏膜(76.7% vs 26.7%,P<0.05)；E-cadherin在BTCC组织中的表达明显低于正常黏膜(30.0% vs 90.0%,P<0.05)；(2)Tβ4与E-cadherin表达呈负相关,与ILK、β-catenin表达呈正相关。结论：胸腺素β4与膀胱移行上皮癌分化和转移有关,可能通过调控上皮-间质转化从而在膀胱移行上皮癌的侵袭转移过程中发挥作用。
第二部分基因沉默胸腺素β4对膀胱移行细胞癌上皮间质转化的逆转作用
目的：探讨胸腺素β4基因沉默对膀胱癌细胞EMT的逆转,进一步研究其在肿瘤侵袭转移中的作用机制。方法：使用针对Tβ4的慢病毒载体(Lenti-Tβ4)转染膀胱癌细胞株T24。采用定量RT-PCR、Western blot法检测Tβ4、ILK和上皮性标记基因E-cadherin、β-catenin的表达改变；Immunofluorescence法检测ILK、E-cadherin、β-catenin在转染后T24细胞中的表达改变；细胞划痕实验、Boyden小室体外侵袭实验、AO/EB荧光染色法反映细胞转移潜能和凋亡的变化。结果：转染后的T24细胞,Tβ4、ILK和β-catenin基因或蛋白表达下调,而上皮标记基因E-cadherin表达增强(p<0.05)；Immunofluorescence显示ILK和β-catenin在细胞质、细胞核中表达减弱,而E-cadherin在胞膜中表达增强,细胞形态向正常上皮细胞转化；转染后的T24细胞体外迁移能力与侵袭力下降,细胞凋亡增多。结论：肿瘤细胞中的Tβ4表达下调可以逆转肿瘤细胞的间质表型,而向正常上皮表型转化,降低肿瘤转移潜能。
第三部分Tβ4-siRNA慢病毒载体逆转膀胱癌细胞上皮间质转化的体内实验研究
目的：构建可荧光示踪的膀胱癌小鼠皮下移植瘤模型及肺转移模型,并探讨活体内抑制Tβ4基因表达对膀胱癌生长侵袭、转移及血管形成的影响,为后续研究奠定基础。方法：(1)选用膀胱移行细胞癌细胞株EJ,体外扩增培养制成细胞悬液后注射于BALB/C小鼠颈背部皮下,待有肉眼可见瘤块后,处死小鼠并取出肿瘤组织。对肿瘤组织进行病理组织学观察,免疫组化方法检测Tβ4、ILK、E-cadherin和β-catenin在移植瘤中的表达,TUNEL方法检测活体内抑制Tβ4基因表达对膀胱癌细胞的促凋亡作用,体外血管形成实验检测Tβ4表达下调对血管生成的影响。(2)将膀胱移行细胞癌细胞株EJ悬液接种于BALB/C小鼠尾静脉内,于术后第20天处死小鼠,观察胸腔及肺内转移灶大小、数量,观察生存天数,记录远处转移情况。结果：(1)EJ细胞以慢病毒Lenti-Tβ4转染后形成的膀胱癌移植瘤与EJ细胞慢病毒Lenti-GFP转染(阴性对照)后形成的移植瘤比较,移植瘤体积、湿重、小鼠平均生存天数均存在差异,差异有统计学意义(P<0.05)；移植瘤组织HE染色发现,干扰移植瘤中Tβ4基因表达,癌组织恶性程度相较于未干扰组低；Lenti-Tβ4组中Tβ4、ILK和β-catenin阳性表达率明显低于阴性对照组,而E-cadherin阳性表达率明显高于阴性对照组；Lenti-Tβ4组凋亡率明显高于阴性对照组；与空白对照组、阴性对照组相比,Lenti-Tβ4转染组促进血管形成能力下降66.8%,有统计学意义(P<0.01)。(2)BALB/C新生仔鼠尾静脉注入膀胱癌细胞EJ后,20d后解剖取出肺组织,阴性对照组肺组织表面不光滑,肉眼见肺表面弥漫性斑块,最大直径1.5 cm,而Lenti-Tβ4转染组肺组织中转移灶明显变小或未见明显转移灶,各组小鼠的肝脏、脾脏、心脏、骨均未发现明显转移。结论：携带干扰序列Tβ4-siRNA的慢病毒能有效地抑制Tβ4基因的表达,从而抑制膀胱癌细胞体内的侵袭能力,同时发现Tβ4基因表达与膀胱癌细胞的转移活性及血管生成相关,下调其表达水平能够明显抑制癌细胞的转移能力。
Part 1 Correlation between thymosinβ4 and epithelial-mesenchymal transition in human bladder transitional cell carcinoma
Objective:To investigate the relationship between human thymosinβ4 (T(34) and epithelial-mesenchymal transition (EMT) in bladder transitional cell carcinoma (BTCC), explore the association with clinicopathologic features. Methods:60 BTCC specimens from surgically resected were selected. For each specimen, the cancerous tissue、peri-cancer tissue and its remote normal mucosa were analyzed and compared. The expression of Tβ4、ILK、E-cadherin andβ-catenin proteins in resected cancer tissues were detected by using Real-time RT-PCR、Western blot and immunohistochemistry, and compared with clinicopathologic data. Results:(1) The expression rate of Tβ4 was significantly higher in BTCC tissues than in normal tissues (91.7% vs 15.0%, P＜0.05), and the expression rate of Tβ4 in deeply infiltrating group was significantly higher than that in superficially infiltrating group(97.4% vs 81%, P＜0.05); the level of ILK was significantly higher in BTCC than in normal tissues(80.0% vs 18.3%, P＜0.05); the expression rate ofβ-catenin was higher in BTCC tissues than in normal tissues(76.7% vs 26.7%, P＜0.05); the expression of E-cadherin in BTCC tissues were significantly lowered in comparison with those in normal bladder epithelium(30.0% vs 90.0%, P＜0.05); (2) Tβ4 expression was inversely correlated to E-cadherin, but positively to ILK andβ-catenin. Conclusion: Thymosinβ4 is related to the differentiation and metastasis of bladder transitional cell carcinoma, and may induce EMT to promote BTCC metastasis.
Part 2 The reverse effect of thymosinβ4 gene silencing on EMT in human bladder transitional cell carcinoma
Objective:To investigate the reverse effect of epithelial-mesenchymal transition (EMT) by silencing human gene thymosinβ4(Tβ4), and further reseach for its role on invasion and metastasis of cancer. Methods:Lentiviral shRNA vector encoding Tβ4 was transfected into T24 cell line. The expression of Tβ4、ILK and epithelial markers E-cadherin、β-catenin were detected by Real-time RT-PCR and Western blot assay; the expression of ILK、E-cadherin andβ-catenin in T24 cells were detected by immunofluorescence after transfection via lentiviral vector; the metastatic potential and apoptosis were examined by in vitro cell wound model、Boyden chamber invasion assay and acridine orange-ethidium bromide fluorescent staining. Results:The expression of Tβ4、ILK andβ-catenin were decreased in the T24 cells after transfected with Tβ4 shRNA, and the expression of E-cadherin was significantly up-regulated (p<0.05); the level of ILK in cytoplasm and level ofβ-catenin in nuclear were obviously decreased by immunofluorescence analysis, and the higher level of E-cadherin was observed, the morphology of T24 Cells was transformed into a normal epithelial phenotype; the motility、invation of T24 cells were inhibited and apoptosis was enhanced. Conclusion:The silencing of Tβ4 may reverse fibroblastoid morphology into a normal epithelial phenotype, and suppress tumor metastatic potential.
Part 3 The study on reversion of epithelial-mesenchymal transition (EMT) in bladder transitional cell carcinoma (BTCC) by using lentivirus-mediated Tp4-siRNA in vivo
Objective:To establish xenografted tumor and lung metastasis BALB/C mouse models of bladder transitional cell carcinoma (BTCC) expressing green fluorescent protein, and investigate effect of Tumor Growth, Metastasis and Apoptosis in vivo by inhibiting Tβ4 expression. so as to lay a foundation for future study. Methods:(1)EJ cell line was used in the present study. The cells were proliferated and the cell suspension was subcutaneously injected into the BALB/C mice. Histologic and immunohistochemical analysis were performed for Tβ4, ILK,β-catenin and E-cadherin. The formed tumor cells apoptosis was detected by TUNEL in the BALB/c nude mice and images were taken using light microscopy. Dark-brown apoptotic bodies were located in nucleus. (2)The in situ tumor information, lymphatic and pulmonary metastasis were all recorded in detail. EJ cells transfected with lenti-GFP (Cont) or lenti-Tβ4 shRNA (lenti-Tβ4) were injected into the tail veins of mice (20 mice per group), and the mice were killed 20 days after injection. The number of metastatic lung nodules was determined by direct counting of the nodules on the lung. Dark arrows indicate the nodules on the lung. Quantification of antiangiogenic activity was calculated by measuring the length of tube walls formed between discrete endothelial cells in each well. Results:(1)Note the E-cadherin stain in the adjacent epidermis of lenti-Tβ4 group xenografts. Note the cytoplasmic staining of the Tβ4 patterns, while ILK showed a corresponding pattern. This staining pattern stands in sharp contrast to the expression profile of E-cadherin, which was absent in the majority of cases in lenti-Tβ4 group, while positivity in staining was observed in control group. (2)The lung metastasis was observed in BALB/C mice, two weeks after mice were injected intravenously with lentivirus-infected cells, the mean number of metastatic lung nodules was 2 for mice injected with EJ cells infected with Tβ4 shRNA and 6 for mice injected with EJ cells infected with control lentivirus (P<0.05). Extensive tube formation of endothelial cells was observed in non-transfection and lenti-GFP (Cont) groups. However, when the endothelial cells were treated by the medium preconditioned with Tβ4 shRNA-transfected EJ cells, the tube formation was markedly suppressed. Conclusion:These data suggested that overexpression of Tβ4 was associated with increases in primary tumor growth and in the number of lung metastases, and transfection of Tβ4 shRNA remarkably inhibited cancer cell malign differentiation in vivo and decreased the angiogenesis of urothelial carcinoma cells in vitro.
引文
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9. Persad S, Attwell S, Gray V, et al. Regulation of protein kinaseB/Akt-serine 473 phosphorylation by integrin-linked kinase:critical roles for kinase activity and amino acids arginine 211 and serine 343[J]. J Biol Chem,2001,276 (29):27462-27469.
10. Tan C, Costello P, Sanghera J, et al. Inhibition of integrin linked kinase (ILK) suppresses beta-catenin Lef/Tcf-dependent transcription and expression of the E-cadherin repressor, snail, in APC-/-human colon carcinoma cells [J]. Oncogene,2001, 20(1):133-140.
1. Zhang Y, Feurino LW, Zhai Q, et al. Thymosin Beta 4 is Overexpressed in Human Pancreatic Cancer Cells and Stimulates Proinflammatory Cytokine Secretion and JNK Activation[J]. Cancer Biol Ther,2007 Dec 13;7(3).
2. Huang HC, Hu CH, Tang MC, et al. Thymosin beta4 triggers an epithelial-mesenchymal transition in colorectal carcinoma by upregulating integrin-linked kinase[J]. Oncogene,2006 2007; 26:2781-2790
3. Baumgart E, Cohen MS, Silva Neto B, et al. Identification and prognostic significance of an epithelial-mesenchymal transition expression profile in human bladder tumors[J]. Clin Cancer Res,2007 Mar 15;13(6):1685-94.
4. Nummela P, Yin M, Kielosto M, et al. Thymosin beta4 is a determinant of the transformed phenotype and invasiveness of S-adenosylmethionine decarboxylase-transfected fibroblasts[J]. Cancer Res,2006,66(2):701-712.
5. Yauch RL, Januario T, Eberhard DA, et al. Epithelial versus mesenchymal phenotype determines in vitro sensitivity and predicts clinical activity of erlotinib in lung cancer patient s[J]. Clin Cancer Res,2005,11(24):8686-8698.
6. Lee MY, Chou CY, Tang MJ, et al. Epithelial-mesenchymal transition in cervical cancer:correlation with tumor progression, epidermal growth factor receptor overexpression and snail up-regulation[J]. Clin Cancer Res,2008,14(15):4743-50.
7. Thuault S, Tan EJ, Peinado H, et al. HMGA2 and Smads co-regulate SNAIL1 expression during induction of epithelial-mesenchymal transition[J]. JBiol Chem,2008, 283(48):33437-46.
8. Liu X. Inflammatory cytokines augments TGF-b1-induced epithelial-mesenchymal transition in A549 cells by up-regulating TbR-I[J]. Cell Motil Cytoskeleton,2008, 65(12):935-44.
9. Clark EA, Golub TR, Lander ES, et al. Genomic analysis of metastasis reveals an essential role for Rhoc[J]. Nature,2000,406(6795):532-535.
10. Bachelder RE, Yoon S, Franci C, et al. Glycogen synthase kinase-3 is an endogenous inhibitor of Snail transcription:implications for the epithelial-mesenchymal transition[J]. J Cell Biol,2005,168 (1):29-33.
1. Lee MY, Chou CY, Tang MJ, et al. Epithelial-mesenchymal transition in cervical cancer:correlation with tumor progression, epidermal growth factor receptor overexpression and snail up-regulation[J]. Clin Cancer Res,2008,14(15):4743-4750.
2. Huang HC, Hu CH, Tang MC, et al. Thymosin beta4 triggers an epithelial-mesenchymal transition in colorectal carcinoma by upregulating integrin-linked kinase[J]. Oncogene,2007; 26:2781-2790.
3. Graham TR, Zhau HE, Odero-Marah VA, et al. Insulin-like growth factor-I-dependent up-regulation of ZEB1 drives epithelial-to-mesenchymal transition in human prostate cancer[J]. Cancer Res,2008,68(7):79-88.
4. Thiery J P. Epithelial-mesenchymal transitions in tumour progression[J]. CurrOp in Cell Biol,2003,15 (6):740-746.
5. Cha HJ, Jeong MJ, Kleinman HK. Role of thymosin beta 4 in tumor metastasis and angiogenesis[J]. J Natl Cancer Inst,2003,95:1674-1680.
6. Takanami I. Increased expression of integrin-linked kinase is associated with melanoma progression and poor patient shorter survival in non-small cell lung cancer[J]. BMC Cancer,2005,5(1):1.
7. Park SM, Gaur AB, Lengyel E, et al. The miR-200 family determines the epithelial phenotype of cancer cells by targeting the E-cadherin repressors ZEB1 and ZEB2[J]. Genes Dev,2008,22(7):894-907.
1. Xiao Z, McCallum TJ, Brown KM, et al. Characterization of a novel transplantable orthotopic rat bladder transitional cell tumor model[J]. Br J Cancer,1999,81 (4):638-646.
2. Kikuch iE, Menendez S, Ozu C, et al. Highly efficient gene delivery for bladder cancers by intravesically administered replication-competent retroviral vectors [J]. Clin Cancer Res,2007,13(15):4511-4518.
3. Develasco MA, Tanaka M, Anai S, et al. GFP image analysis in the mouse orthotopic bladder cancer model[J]. OncolRep,2008,20(3):543-547.
4. Kobayashi T, Okada F, Fujii N, et al. Thymosin-beta 4 regulates motility and metastasis of malignant mouse fibrosarcoma cells[J]. Am J Pathol,2002,160(3):869-882.
5. Grant DS, Kinsella JL, Kibbey MC, et al. Matrigel induces thymosin beta 4 gene in differrentiating endothelial cells[J]. J Cell Sci,1995,108(12):3685-3694.
6. Domanski M, Hertzog M, Coutant J, et al. Coupling of folding and binding of thymosin beta 4 upon interaction with monomeric actin monitored by nuclear magnetic resonance[J]. JBiol Chem,2004,279(22):23637-23645.
7. Clark EA, Golub TR, Lander ES, et al. Genomic analysis of metastasis reveals an essential role for Rhoc[J]. Nature,2000,406(6795):5322535.
1. Cha HJ, Jeong MJ, Kleinman HK. Role of thymosin beta 4 in tumor metastasis and angiogenesis[J]. J Natl Cancer Inst,2003,95(22):1674-1680.
2. Huang HC, Hu CH, Tang MC, et al. Thymosin beta4 triggers an epithelial-mesenchymal transition in colorectal carcinoma by upregulating integrin-linked kinase[J]. Oncogene,2007,26(19):2781.
3. Wang WS, Chen PM, Hsiao HL, et al. Overexpression of the thymosin beta-4 gene is associated with increased invasion of SW480 colon carcinoma cells and the distant metastasis of human colorectal carcinoma[J]. Oncogene,2004,23(39):6666.
4. Boyar B, Dufour S, Thiery JP. E-cadherin expression during the acidic FGF-induced dispersion of a rat bladder carcinoma cell line[J]. Exp Cell Res,1992,201(2):347-357.
5. Behrens J, Vakaet L, Friis R, et al. Loss of epitheliation and gain of invasiveness correlates with tyrosine phosphorylation of the E-cadherin/β-catenin complex in cells transformed with a temperature-sensitive v-SRC gene[J]. Cell Biol,1993,120 (3):757-766.
6. Martin Oft, Rosemary J, Balmain A, et al. Metastasis is driven by sequential elevation of H-ras and Smad2 levels[J]. Nature Cell Biology,2002,7(1):487-494.
7. Bates RC, Mercurio AM, DeLeo MJ, et al. The epithelial-mesenchymal transition of colon carcinoma involves expression of IL-8 and CXCR-1-mediated chemotaxis[J]. Exp Cell Res,2004,299(2):315-324.
8. Ikeguchi M, Makino M, Kaibara N. Clinical significance of E-caderin-catenin complex expression in metastatic foci of colorectal carcinoma[J]. J Surg Oncol,2001, 77:201-207.
9. Takahashi M, Fujita M, FurukawaY, et al. Isolation of a novel human gene, APCDD1, as a direct target of the beta-catenin/T cell factor 4 complex with probable involvement in colorectal carcinogenesis[J]. Cancer Res,2002,62:5651-5656.
10. Roura S, Miravet S, Piedra J, et al. Regulation of E-cadherin/Catenin association by tyrosine phosphorylation[J]. J Biol Chem,1999,17:36734-36740.
11. Kikuchi A. Regulation of beta-catenin signaling in the Wnt pathway [J]. Biochem Biophys Res Comun,2000,268:243-248.
12. Benetti R, Copetti T, Dell'orso S, et al. The calpain system is involved in the constitutive regulation of beta-catenin signaling functions[J]. J BiolChem,2005,280: 22070-22080.
13. Lim J, Kim JH, Paeng JY, et al. Prognostic value of acti-vated Akt expression in oral squamous cell carcinoma[J]. J Clin Pathol,2005,58(11):1199-1205.
14. Chang JY, Wright JM, Svoboda KK. Signal transduction pathways involved in epithelial-mesenchymal transition in oral cancer compared with other cancers[J]. Cells Tissues Organs,2007,185(1-3):40-47.
15. Sahai E, Marshall CJ. Rho-GTPases and Cancer[J]. Nat Rev Cancer,2002,133-142.
16. Bhowmick NA, Ghiassi M. Transforming growth factor beta mediates epithelial to mesenchymal transdifferentiation through a RhoA dependent mechanism [J]. Mol Biol Cell,2001,27-36.
17. Fukata M, Kaibuchi K. Rho-family GTPases in cadherin-mediated cell-cell adhesion[J]. Nat Rev Mol Cell Biol,2001,887-897.
18. Burridge K, Wennerberg K. Rho and Rac take center stage[J]. Cell,2004,116 (2):167-179.
19. Minard ME, Kim LS, Price JE, et al. The role of the guaninenucleotide exchange factor Tiaml in cellular migration, invasion, adhesion and tumor progression[J]. Breast Cancer Res Treat,2004,84(1):21-32.
20. Malliri A, van EsS, Huveneers S, et al. The Rac exchange factor Tiaml is required for the establishment and maintenance of cadherin-based adhesions[J]. J Biol Chem, 2004,279(29):30092-30098.
21. Minard ME, Ellis LM, Gallick GE. Tiaml regulates cell adhesion, migration and apoptosis in colon tumor cells[J]. Clin Exp Metastasis,2006,23(5-6):301-13.
22. Hung KF, Chang CS, Liu CJ, et al. Differential expression of E-cadherin in metastatic lesions comparing to primary oral squamous cell carcinoma[J].J Oral Pathol Med, 2006,35(10):589-594.
23. Yanjia H, Xinchun J. The role of epithelial-mesenchymal transition in oral squamous cell carcinoma and oral sub-mucous fibrosis[J]. Clin Chim Acta,2007,383(1-2):51-56.
24. Diamond ME, Sun L, Ottaviano AJ, et al. Differential growth factor regulation of N-cadherin expression and motility in normal and malignant oral epithelium[J]. J Cell Sci,2008,121(13):2197-2207.
25. Nakajima S, Doi R, Toyoda E, et al. N-cadherin expression and epithelial-mesenchymal transition in pancreatic carcinoma[J]. Clin Cancer Res, 2004,10(12):4125-4133.
26. Fukata M, Nakagawa M, Kaibuchi K, et al. Roles of Rho-family GTPases in cell polarisation and directional migration [J]. Curr Opin Cell Biol,2003,15(5):590-597.
27. Yang J, Mani S, Donaher J, et al. Twist, a master regulator of morphogenesis, play an essential role in tumor metastasis [J]. Cell,2004,117(7):927-939.
28. Gregory PA, Bert AG, Paterson EL, et al. The miR-200 family and miR-205 regulate epithelial to mesenchymal transition by targeting ZEB1and SIP1[J]. Nat Cell Biol, 2008,10(5):593-601
29. Kong WL, Yang H, He LL, et al. MicroRNA-155 is regulated by the transforming growth factor-β/Smad pathway and contributes to epithelial cell plasticity by targeting RhoA[J]. Mol Cell Biol,2008,28(22):6773-6784.
30. Putz E, Witter K, Offner S, et al. Phenotypic characteristics of cell lines derived from disseminated cancer cells in bone marrow of patients with solid epithelial tumors: establishment of working models for human micro-metastases[J]. Cancer Res, 1999,59(2):241-248.
31. Kobayashi T, Okada F, Fujii N, et al. Thymosin-beta 4 regulates motility and metastasis of malignant mouse fibrosarcoma cells[J]. Am JPathol,2002,160(3):869-882.
32. Philp D, Goldstein AL, Kleinman HK. Thymosin beta 4 promotes angiogenesis, wound healing, and hair follicle development[J]. Mech Age-ing Dev,2004,125(2):113-115.
33. Wang WS, Chen PM, Hsiao HL, et al. Overexpression of the thymosin beta 4 gene is associated with malignant progression of SW480colon cancer cells[J]. Oncogene, 2003,22(21):3297-3306.
34. Diamond DL, Zhang Y, Gaiger A, et al. Use of Protein Chip arraysurface enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOFMS) to identify thymosin beta 4, a differentially secreted protein from lymphoblastoid cell lines[J]. JAm Soc Mass Spectrom,2003,14(7):760-765.
35. Gu XM, Zhang H, Lai MD. Bioinformatics analysis to the differentially expressed genes of normal mucosa and carcinoma of colon[J]. Zhe-jiang Da Xue Xue Bao Yi Xue Ban,2004,33(2):95-101,124.
36. Li X, Zheng L, Peng F, et al. Recombinant thymosin beta4 can promote full-thickness cutaneous wound healing[J]. Protein Expr Purif,2007,56(2):229.
37. Larsson LI, Holck S. Occurrence of thymosin beta4 in human breast cancer cells and in other cell types of the tumor microenvironment[J]. Hum Pathol,2007,38(1):114.
38. Clark EA, Golub TR, Lander ES, et al. Genomic analysis of metastasis reveals an essential role for Rhoc[J]. Nature,2000,406(6795):5322535.
39. Grant DS, Kinsella JL, Kibbey MC, et al. Matrigel induces thymosin beta 4 gene in differrentiating endothelial cells[J]. J Cell Sci,1995,108(12):3685-3694.
40. Oh IS, So SS, Jahng KY, et al. Hepatocyte growth factor upregulates thymosin beta 4 in human umbilicalvein endothelial cell[J]. Biochem Biop hys Res Commun,2002, 296(2):4012405.
41. Ji P, Diederichs S, Wang W, et al. MALAT21, a novel no ncoding RNA, and thymosin beta 4 predict metastasis and survival in early stage non-small cell lung cancer [J]. Oncogene,2003,22:8031-8041.
42.周瑞,袁宏银,熊斌,等.大肠癌组织中胸腺素β4基因表达及其临床意义[J].肿瘤防治研究杂志,2005,32(9)：545-548.
43. Liu CR, Ma CS, Ning JY, et al. Thymosin beta 10 expression and actin filament organization in tumor cell lines with different metastatic potential[J]. Zhonghua BingLi Xue Za Zhi,2004,33 (1):67-71.
44. Domanski M, Hertzog M, Coutant J, et al. Coupling of folding and binding of thymosin beta 4 upon interaction with monomeric actin monitored by nuclear magnetic resonance[J]. J Biol Chem,2004,279(22):23637-23645.
45. Chaffer CL, Brennan JP, Slavin JL, et al. Mesenchymal-to-epithelial transition facilitates bladder cancer metastasis:role of fibroblast growth factor rectpor-2[J]. Cancer Res,2006,66(23):11271-11278.
46. Lee TK, Poon RT, Yuen AP, et al. Regulation of angiogenesis byld-1 through hypoxia-inducible factor-1 alpha-mediated vascular endothelial growth factor up-regulation in hepatocellular carcinoma[J]. Clin Cancer Res,2006, 12(18):5369-5376.
47. Alonso SR, Tracey L, Ortiz P, et al. A high-throughput study in melanoma identifies epithelial-mesenchymal transition as a major determinant of mestastasis[J]. Cancer Res, 2007,67(7):3450-3460.
48. Yang J, Maui SA, Weinberg RA. Exploring a new twist on tumor metastasis[J]. Cancer Res,2006,66(9):4549-4552.
2. Guarino M. Epithelial-mesenchymal transition and tumour invasion[J]. Int J Biochem Cell Biol,2007,39 (12):2153-2160.
3. KongW, Yang H, He L, et al. MicroRNA-155 is regulated by the transforming growth factor beta/Smad pathway and contributes to epithelial cell plasticity by targeting RhoA[J]. Mol Cell Biol,2008,28 (22):6773-6784.
4. Nummela P, Yin M, Kielosto M, et al.Thymosin beta4 is a determinant of the transformed phenotype and invasiveness of S-adenosylmethionine decarboxylase-transfected fibroblasts[J]. Cancer Res.2006,66(2):701-12.
5. Zhang Y, Feurino LW, Zhai Q, et al. Thymosin Beta 4 is Overexpressed in Human Pancreatic Cancer Cells and Stimulates Proinflammatory Cytokine Secretion and JNK Activation[J]. Cancer Biol Ther.2008;7(3):419-423.
6. Huang HC, Hu CH, Tang MC, et al. Thymosin beta4 triggers an epithelial-mesenchymal transition in colorectal carcinoma by upregulating integrin-linked kinase[J]. Oncogene.2006 2007; 26:2781-2790.
7. Graham TR, Zhau HE, Odero-Marah VA, et al. Insulin-like growth factor-I-dependent up-regulation of ZEB1 drives epithelial to mesenchymal transition in human prostate cancer cells [J]. Cancer Res,2008,68 (7):2479-2488.
8. Zhau HE, Odero-Marah V, Lue HW, et al. Epithelial to mesenchymal transition (EMT) in human prostate cancer:lessons learned from ARCaP model[J]. Clin Exp Metastasis, 2008,25(6):601-610.
9. Persad S, Attwell S, Gray V, et al. Regulation of protein kinaseB/Akt-serine 473 phosphorylation by integrin-linked kinase:critical roles for kinase activity and amino acids arginine 211 and serine 343[J]. J Biol Chem,2001,276 (29):27462-27469.
10. Tan C, Costello P, Sanghera J, et al. Inhibition of integrin linked kinase (ILK) suppresses beta-catenin Lef/Tcf-dependent transcription and expression of the E-cadherin repressor, snail, in APC-/-human colon carcinoma cells [J]. Oncogene,2001, 20(1):133-140.
1. Zhang Y, Feurino LW, Zhai Q, et al. Thymosin Beta 4 is Overexpressed in Human Pancreatic Cancer Cells and Stimulates Proinflammatory Cytokine Secretion and JNK Activation[J]. Cancer Biol Ther,2007 Dec 13;7(3).
2. Huang HC, Hu CH, Tang MC, et al. Thymosin beta4 triggers an epithelial-mesenchymal transition in colorectal carcinoma by upregulating integrin-linked kinase[J]. Oncogene,2006 2007; 26:2781-2790
3. Baumgart E, Cohen MS, Silva Neto B, et al. Identification and prognostic significance of an epithelial-mesenchymal transition expression profile in human bladder tumors[J]. Clin Cancer Res,2007 Mar 15;13(6):1685-94.
4. Nummela P, Yin M, Kielosto M, et al. Thymosin beta4 is a determinant of the transformed phenotype and invasiveness of S-adenosylmethionine decarboxylase-transfected fibroblasts[J]. Cancer Res,2006,66(2):701-712.
5. Yauch RL, Januario T, Eberhard DA, et al. Epithelial versus mesenchymal phenotype determines in vitro sensitivity and predicts clinical activity of erlotinib in lung cancer patient s[J]. Clin Cancer Res,2005,11(24):8686-8698.
6. Lee MY, Chou CY, Tang MJ, et al. Epithelial-mesenchymal transition in cervical cancer:correlation with tumor progression, epidermal growth factor receptor overexpression and snail up-regulation[J]. Clin Cancer Res,2008,14(15):4743-50.
7. Thuault S, Tan EJ, Peinado H, et al. HMGA2 and Smads co-regulate SNAIL1 expression during induction of epithelial-mesenchymal transition[J]. JBiol Chem,2008, 283(48):33437-46.
8. Liu X. Inflammatory cytokines augments TGF-b1-induced epithelial-mesenchymal transition in A549 cells by up-regulating TbR-I[J]. Cell Motil Cytoskeleton,2008, 65(12):935-44.
9. Clark EA, Golub TR, Lander ES, et al. Genomic analysis of metastasis reveals an essential role for Rhoc[J]. Nature,2000,406(6795):532-535.
10. Bachelder RE, Yoon S, Franci C, et al. Glycogen synthase kinase-3 is an endogenous inhibitor of Snail transcription:implications for the epithelial-mesenchymal transition[J]. J Cell Biol,2005,168 (1):29-33.
1. Lee MY, Chou CY, Tang MJ, et al. Epithelial-mesenchymal transition in cervical cancer:correlation with tumor progression, epidermal growth factor receptor overexpression and snail up-regulation[J]. Clin Cancer Res,2008,14(15):4743-4750.
2. Huang HC, Hu CH, Tang MC, et al. Thymosin beta4 triggers an epithelial-mesenchymal transition in colorectal carcinoma by upregulating integrin-linked kinase[J]. Oncogene,2007; 26:2781-2790.
3. Graham TR, Zhau HE, Odero-Marah VA, et al. Insulin-like growth factor-I-dependent up-regulation of ZEB1 drives epithelial-to-mesenchymal transition in human prostate cancer[J]. Cancer Res,2008,68(7):79-88.
4. Thiery J P. Epithelial-mesenchymal transitions in tumour progression[J]. CurrOp in Cell Biol,2003,15 (6):740-746.
5. Cha HJ, Jeong MJ, Kleinman HK. Role of thymosin beta 4 in tumor metastasis and angiogenesis[J]. J Natl Cancer Inst,2003,95:1674-1680.
6. Takanami I. Increased expression of integrin-linked kinase is associated with melanoma progression and poor patient shorter survival in non-small cell lung cancer[J]. BMC Cancer,2005,5(1):1.
7. Park SM, Gaur AB, Lengyel E, et al. The miR-200 family determines the epithelial phenotype of cancer cells by targeting the E-cadherin repressors ZEB1 and ZEB2[J]. Genes Dev,2008,22(7):894-907.
1. Xiao Z, McCallum TJ, Brown KM, et al. Characterization of a novel transplantable orthotopic rat bladder transitional cell tumor model[J]. Br J Cancer,1999,81 (4):638-646.
2. Kikuch iE, Menendez S, Ozu C, et al. Highly efficient gene delivery for bladder cancers by intravesically administered replication-competent retroviral vectors [J]. Clin Cancer Res,2007,13(15):4511-4518.
3. Develasco MA, Tanaka M, Anai S, et al. GFP image analysis in the mouse orthotopic bladder cancer model[J]. OncolRep,2008,20(3):543-547.
4. Kobayashi T, Okada F, Fujii N, et al. Thymosin-beta 4 regulates motility and metastasis of malignant mouse fibrosarcoma cells[J]. Am J Pathol,2002,160(3):869-882.
5. Grant DS, Kinsella JL, Kibbey MC, et al. Matrigel induces thymosin beta 4 gene in differrentiating endothelial cells[J]. J Cell Sci,1995,108(12):3685-3694.
6. Domanski M, Hertzog M, Coutant J, et al. Coupling of folding and binding of thymosin beta 4 upon interaction with monomeric actin monitored by nuclear magnetic resonance[J]. JBiol Chem,2004,279(22):23637-23645.
7. Clark EA, Golub TR, Lander ES, et al. Genomic analysis of metastasis reveals an essential role for Rhoc[J]. Nature,2000,406(6795):5322535.
1. Cha HJ, Jeong MJ, Kleinman HK. Role of thymosin beta 4 in tumor metastasis and angiogenesis[J]. J Natl Cancer Inst,2003,95(22):1674-1680.
2. Huang HC, Hu CH, Tang MC, et al. Thymosin beta4 triggers an epithelial-mesenchymal transition in colorectal carcinoma by upregulating integrin-linked kinase[J]. Oncogene,2007,26(19):2781.
3. Wang WS, Chen PM, Hsiao HL, et al. Overexpression of the thymosin beta-4 gene is associated with increased invasion of SW480 colon carcinoma cells and the distant metastasis of human colorectal carcinoma[J]. Oncogene,2004,23(39):6666.
4. Boyar B, Dufour S, Thiery JP. E-cadherin expression during the acidic FGF-induced dispersion of a rat bladder carcinoma cell line[J]. Exp Cell Res,1992,201(2):347-357.
5. Behrens J, Vakaet L, Friis R, et al. Loss of epitheliation and gain of invasiveness correlates with tyrosine phosphorylation of the E-cadherin/β-catenin complex in cells transformed with a temperature-sensitive v-SRC gene[J]. Cell Biol,1993,120 (3):757-766.
6. Martin Oft, Rosemary J, Balmain A, et al. Metastasis is driven by sequential elevation of H-ras and Smad2 levels[J]. Nature Cell Biology,2002,7(1):487-494.
7. Bates RC, Mercurio AM, DeLeo MJ, et al. The epithelial-mesenchymal transition of colon carcinoma involves expression of IL-8 and CXCR-1-mediated chemotaxis[J]. Exp Cell Res,2004,299(2):315-324.
8. Ikeguchi M, Makino M, Kaibara N. Clinical significance of E-caderin-catenin complex expression in metastatic foci of colorectal carcinoma[J]. J Surg Oncol,2001, 77:201-207.
9. Takahashi M, Fujita M, FurukawaY, et al. Isolation of a novel human gene, APCDD1, as a direct target of the beta-catenin/T cell factor 4 complex with probable involvement in colorectal carcinogenesis[J]. Cancer Res,2002,62:5651-5656.
10. Roura S, Miravet S, Piedra J, et al. Regulation of E-cadherin/Catenin association by tyrosine phosphorylation[J]. J Biol Chem,1999,17:36734-36740.
11. Kikuchi A. Regulation of beta-catenin signaling in the Wnt pathway [J]. Biochem Biophys Res Comun,2000,268:243-248.
12. Benetti R, Copetti T, Dell'orso S, et al. The calpain system is involved in the constitutive regulation of beta-catenin signaling functions[J]. J BiolChem,2005,280: 22070-22080.
13. Lim J, Kim JH, Paeng JY, et al. Prognostic value of acti-vated Akt expression in oral squamous cell carcinoma[J]. J Clin Pathol,2005,58(11):1199-1205.
14. Chang JY, Wright JM, Svoboda KK. Signal transduction pathways involved in epithelial-mesenchymal transition in oral cancer compared with other cancers[J]. Cells Tissues Organs,2007,185(1-3):40-47.
15. Sahai E, Marshall CJ. Rho-GTPases and Cancer[J]. Nat Rev Cancer,2002,133-142.
16. Bhowmick NA, Ghiassi M. Transforming growth factor beta mediates epithelial to mesenchymal transdifferentiation through a RhoA dependent mechanism [J]. Mol Biol Cell,2001,27-36.
17. Fukata M, Kaibuchi K. Rho-family GTPases in cadherin-mediated cell-cell adhesion[J]. Nat Rev Mol Cell Biol,2001,887-897.
18. Burridge K, Wennerberg K. Rho and Rac take center stage[J]. Cell,2004,116 (2):167-179.
19. Minard ME, Kim LS, Price JE, et al. The role of the guaninenucleotide exchange factor Tiaml in cellular migration, invasion, adhesion and tumor progression[J]. Breast Cancer Res Treat,2004,84(1):21-32.
20. Malliri A, van EsS, Huveneers S, et al. The Rac exchange factor Tiaml is required for the establishment and maintenance of cadherin-based adhesions[J]. J Biol Chem, 2004,279(29):30092-30098.
21. Minard ME, Ellis LM, Gallick GE. Tiaml regulates cell adhesion, migration and apoptosis in colon tumor cells[J]. Clin Exp Metastasis,2006,23(5-6):301-13.
22. Hung KF, Chang CS, Liu CJ, et al. Differential expression of E-cadherin in metastatic lesions comparing to primary oral squamous cell carcinoma[J].J Oral Pathol Med, 2006,35(10):589-594.
23. Yanjia H, Xinchun J. The role of epithelial-mesenchymal transition in oral squamous cell carcinoma and oral sub-mucous fibrosis[J]. Clin Chim Acta,2007,383(1-2):51-56.
24. Diamond ME, Sun L, Ottaviano AJ, et al. Differential growth factor regulation of N-cadherin expression and motility in normal and malignant oral epithelium[J]. J Cell Sci,2008,121(13):2197-2207.
25. Nakajima S, Doi R, Toyoda E, et al. N-cadherin expression and epithelial-mesenchymal transition in pancreatic carcinoma[J]. Clin Cancer Res, 2004,10(12):4125-4133.
26. Fukata M, Nakagawa M, Kaibuchi K, et al. Roles of Rho-family GTPases in cell polarisation and directional migration [J]. Curr Opin Cell Biol,2003,15(5):590-597.
27. Yang J, Mani S, Donaher J, et al. Twist, a master regulator of morphogenesis, play an essential role in tumor metastasis [J]. Cell,2004,117(7):927-939.
28. Gregory PA, Bert AG, Paterson EL, et al. The miR-200 family and miR-205 regulate epithelial to mesenchymal transition by targeting ZEB1and SIP1[J]. Nat Cell Biol, 2008,10(5):593-601
29. Kong WL, Yang H, He LL, et al. MicroRNA-155 is regulated by the transforming growth factor-β/Smad pathway and contributes to epithelial cell plasticity by targeting RhoA[J]. Mol Cell Biol,2008,28(22):6773-6784.
30. Putz E, Witter K, Offner S, et al. Phenotypic characteristics of cell lines derived from disseminated cancer cells in bone marrow of patients with solid epithelial tumors: establishment of working models for human micro-metastases[J]. Cancer Res, 1999,59(2):241-248.
31. Kobayashi T, Okada F, Fujii N, et al. Thymosin-beta 4 regulates motility and metastasis of malignant mouse fibrosarcoma cells[J]. Am JPathol,2002,160(3):869-882.
32. Philp D, Goldstein AL, Kleinman HK. Thymosin beta 4 promotes angiogenesis, wound healing, and hair follicle development[J]. Mech Age-ing Dev,2004,125(2):113-115.
33. Wang WS, Chen PM, Hsiao HL, et al. Overexpression of the thymosin beta 4 gene is associated with malignant progression of SW480colon cancer cells[J]. Oncogene, 2003,22(21):3297-3306.
34. Diamond DL, Zhang Y, Gaiger A, et al. Use of Protein Chip arraysurface enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOFMS) to identify thymosin beta 4, a differentially secreted protein from lymphoblastoid cell lines[J]. JAm Soc Mass Spectrom,2003,14(7):760-765.
35. Gu XM, Zhang H, Lai MD. Bioinformatics analysis to the differentially expressed genes of normal mucosa and carcinoma of colon[J]. Zhe-jiang Da Xue Xue Bao Yi Xue Ban,2004,33(2):95-101,124.
36. Li X, Zheng L, Peng F, et al. Recombinant thymosin beta4 can promote full-thickness cutaneous wound healing[J]. Protein Expr Purif,2007,56(2):229.
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