长程缺氧对结肠癌细胞增殖的影响  

Effect of long-term exposure to hypoxia on the proliferation of colon cancer cells in vitro

作　　者：杨小丁[1] 程勇[1] 贾健锋[1] 曹先圣[1] 杨勇[1] 

机构地区：[1]重庆医科大学附属第一医院胃肠外科,重庆400016

出　　处：《肿瘤》2013年第1期42-47,共6页Tumor

摘　　要：目的:体外观察长程缺氧环境对结肠癌细胞SW480和HCT116增殖能力的影响并探讨其可能的作用机制。方法:缺氧培养结肠癌SW480和HCT116细胞14～28d,采用MTT法和平板克隆法检测缺氧条件下2种结肠癌细胞增殖能力的改变;激光共聚焦显微镜下观察缺氧诱导因子-1α(hypoxia-inducible factor-1α,HIF-1α)和Wnt/β-catenin信号通路关键因子β-catenin的表达;蛋白质印迹法检测HIF-1α和β-catenin、糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)、Ser9被磷酸化的GSK-3β[phospho-glycogen synthase kinase-3β(Ser9),pGSK-3β(Ser9)]以及增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)的表达。以常氧下培养的这2种细胞作为对照。结果:细胞生长曲线和平板克隆检测结果均显示,长程缺氧适应后SW480和HCT116细胞的增殖能力较常氧下明显增强(P<0.01);激光共聚焦显微镜下观察发现,HCT116细胞在缺氧条件下HIF-1α的表达部位发生改变;蛋白质印迹法检测发现,与常氧组比较长程缺氧组细胞中HIF-1α(P<0.05)、β-catenin(P<0.05)和pGSK-3β(Ser9)(P<0.01)蛋白的表达均上调,而总GSK-3β的表达无明显变化,PCNA在缺氧条件下表达上调(与常氧组比较,在SW480细胞中P<0.05,在HCT116细胞中P<0.01)。结论:缺氧适应可促进结肠癌细胞SW480和HCT116的增殖,这可能是HIF-1α与Wnt/β-catenin信号转导通路共同作用的结果。Objective： To investigate the effect of long-term exposure to hypoxia on the proliferation of colon cancer SW480 and HCT116 cells, and to explore its possible mechanism. Methods： SW480 and HCT116 cells were cultured under hypoxic conditions for 14-28 days in vitro. The changes of proliferative ablilities of SW480 and HCT116 cells were detected by MTT method and colony formation assay. The expressions of HIF-1α （hypoxia-inducible factor 1α） and β-catenin-Wnt/β-catenin signalling pathway- associated key factor were observed under a LSCM （laser scanning confocal microscope）. The expression levels of HIF-1α, β-catenin, GSK-3β （glycogen synthase kinase-3β）, pGSK-3β （Ser9） （phosphorylation of GSK-3β on the Ser9 residue） and PCNA （proliferating cell nuclear antigen） proteins were detected by Western blotting. The results from these detection methods were compared with those obtained from W480 and HCT116 cells exposed to normoxic conditions. Results： The results of MTT method and colony formation assay revealed that the SW480 and HCT116 cells exposed to hypoxic conditions for 2 weeks had higher proliferative ablilities as compared with the cells exposed to normoxic conditions （P 〈 0.01）. The result of LSCM showed that the subcellular localization of HIF-1α expression was changed. Western blotting result revealed that the expressions of HIF-1α （P 〈 0.05）, β-catenin （P 〈 0.05） and pGSK-3β （Ser9） （P 〈 0.01） proteins of SW480 and HCT116 cells exposed to hypoxic conditions were increased as compared with the cells exposed to normoxic conditions, but the overall expression of GSK-3β did not change. The expression level of PCNA was increased after exposure to normoxic conditions （exposure to hypoxic conditions vs exposure to normoxic conditions; SW480 cells, P 〈 0.05; HCT116 cells, P 〈0.01）. Conclusion： Adaptation to hypoxia may promote the proliferative ability of colon cancer SW480 and HCT116 cell lines and it maybe the result of HIF-1α＇s in

关 键 词：结肠肿瘤 细胞低氧 缺氧诱导因子 Α亚基 WNT Β-CATENIN信号通路 

分 类 号：R735.35[医药卫生—肿瘤]