Effect of Hypoxia and HIF-1α on Epithelial-mesenchymal Transition，Invasion and Proliferation in Colorectal Cancer
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摘要: 目的 研究不同缺氧条件下缺氧诱导因子-1α（HIF-1α）、锌指转录因子Snail及上皮钙黏蛋白（E-cadherin）在结直肠癌SW480细胞中的表达水平，以及它们对细胞的增殖凋亡和侵袭迁移力的影响，探讨三者在结直肠癌的缺氧及上皮间质转化中的作用及可能机制。方法利用氯化钴（CoCl2）化学诱导细胞缺氧模型，实验分组:常氧组、低氧组和无氧组。采用MTT法检测结直肠癌SW480细胞的生长增殖活性，体外侵袭实验（Transwell小室法）检测细胞的侵袭迁移力，流式细胞术（FCM）检测细胞的凋亡及周期变化，RT -PCR及Western blot检测细胞中HIF-1α、Snail及E-cadherin mRNA与蛋白的表达。结果MTT结果显示随CoCl2浓度的增加结直肠癌SW480细胞的增殖活性逐渐增加（P<0.05）;Transwell结果显示随缺氧程度的增加穿过基质膜和聚碳酸酯膜的细胞数逐渐增多，侵袭迁移率增加（P<0.05）;FCM检测结果显示与常氧组比较，无氧组和低氧组G0/G1期SW480细胞明显减少，G2/S期细胞明显增多，细胞凋亡率降低（P<0.05）;RT-PCR与Western blot检测结果显示低氧组、无氧组与常氧组比较，SW480细胞内HIF-1α和Snail mRNA与蛋白的表达增加，E-cadherin mRNA和蛋白的表达降低（P<0.05），且HIF-1α和Snail的表达呈显著正相关，HIF-1α、Snail与E-cadherin的表达均呈显著负相关（P<0.05）。结论缺氧及HIF-1α可促进结直肠癌SW480细胞株的体外生长增殖活性，抑制其凋亡，增强其体外侵袭迁移能力，其机制可能是缺氧诱导HIF-1α表达上调直接作用于SW480细胞产生，亦可能是缺氧或HIF-1α表达上调通过促进Snail表达，抑制E-cadherin表达，间接导致EMT的发生发展而产生。
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关键词:
- 缺氧诱导因子-1α缺氧 /
- 上皮-间质转化 /
- Snail /
- 上皮钙黏蛋白
Abstract: Objective To study the expression level of the hypoxia inducible factor-1α（HIF-1α）、zinc finger transcription factor Snail and E-cadherin in different hypoxia condition in the colorectal cancer SW480 cells,and to explore their effect on the proliferation,apoptosis,invasion and transference,approach the roles and mechanisms of them three in the hypoxia and epithelial-mesenchymal transition of colorectal cancer. Methods Chemical was used to induce the cell hypoxia model by cobalt chloride(CoCl2),and three groups:normoxia group,hypoxia group and non-oxygen group were involved in this study.Methyl thiazolyl tetrazolium(MTT) method was used to detect the proliferative of colorectal cancer SW480 cells,extraorgan invasion experiment(Transwell method) was applied to evaluate the invasion and transference capability of cells,the cell cycle and apoptosis was examined by flow cytometry (FCM),the mRNA and protein expression levels of HIF-1α,Snail and E-cadherin were detected by real time PCR(RT-PCR) and Western blot. Results MTT disclosed that the proliferative activity of the colorectal cancer SW480 cells was increased gradually following the concentration increase of the cobalt chloride(P<0.05);transwell showed that the cells permeated the matrix membrane and polycarbonate membrane was increased gradually accompanied with the degree of hypoxia(P<0.05);FCM revealed that in hypoxia group and non-oxygen group,the cells in G0/G1 period was decreased obviously,the cells in G2/S period was increased obviously,the cells apoptosis was degrade(P<0.05) compared with normoxia group;RT-PCR and Western blot displayed that hypoxia group and non-oxygen group compared with normoxia group,the mRNA and protein level of HIF-1α and Snail in SW480 cells was increased,the mRNA and protein of E-cadherin was decreased(P<0.05),furthermore,HIF-1α and Snail had positive correlation,and they had negative correlation with E-cadherin conspicuously(P<0.05). Conclusion The hypoxia and HIF-1α can promote the extraorgan proliferation activity of the colorectal cancer SW480 cells,inhibit its apoptosis and enhance its extraorgan invasion and migration.The possible mechanism is that hypoxia induced HIF-1α affect SW480 directly,or the hypoxia and HIF-1α expression promote Snail expression and inhibit E-cadherin expression,which lead to EMT and take it place.-
Key words:
- HIF-1αHypoxia /
- Epithelial-mesenchymal transition(EMT) /
- Snail /
- E-cadherin
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[1] Thiery JP,Acloque H,Huang RY,et al.Epithelial-mesenchymal transitions in development and disease[J].Cell,2009,139(5) :871-90. [2] Pea C,García JM,Larriba MJ,et al.SNAIL expression in colon cancer related with CDH1 and VDR downregulationin normal adjacent tissue [J].Oncogene,2009,28 (49):4375-85. [3] Savagner P.The epithelial-mesenchymal transition (EMT) phenomenon[J].Ann Oncol,2010,21(Suppl 7):vii89-92. [4] Liu Y.New insights into epithelial-mesenchymal transition in kidney fibrosis[J].J Am Soc Nephrol,2010,21(2):212-22. [5] Hpfl G,Wenger RH,Ziegler U,et al.Rescue of hypoxia inducible factor 1alpha deficient tumor growth by wildtype cells is independent of vascular endothelial growth factor[J].Cancer Res,2002,62(10):2962-70. [6] Trisciuoglio D,Gabellini C,Desideri M,et al.Bcl-2 regulates HIF-1alpha protein stabilization in hypoxicmelanoma cells via the molecular chaperone HSP90[J].PLoS One,2010,5(7):e11772. [7] Zhao JH,Luo Y,Jiang YG,et al.Knockdown of β-Catenin through shRNA cause a reversal of EMT and metastaticphenotypes induced by HIF-1α[J].Cancer Invest，2011,29(6):377-82. [8] Wells A,Yates C,Shepard CR.E-cadherin as an indicator of mesenchymal to epithelial reverting transitionsduring the metastatic seeding of disseminated carcinomas[J].Clin Exp Metastasis,2008,25(6):621-8. [9] Natalwala A,Spychal R,Tselepis C.Epithelial-mesenchymal transition mediated tumourigenesis in thegastrointestinal tract [J].World J Gastroenterol,2008,14 (24):3792-7. [10] Browne G,Sayan AE,Tulchinsky E.ZEB proteins link cell motility with cell cycle control and cell survival incancer[J].Cell Cycle,2010,9(5):886-91. [11] Larriba MJ,Bonilla F,Muoz A.The transcription factors Snail1 and Snail2 repress vitamin D receptor duringcolon cancer progression [J].J Steroid Biochem Mol Biol,2010,121(1-2):106-9. [12] Galván JA,González MV,Crespo G,et al.Snail nuclear expression parallels higher malignancy potential inneuroendocrine[J].Lung Cancer,2010,69 (3):289-95.
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