缺氧对肺腺癌细胞HIF-1α、GLUT-1、MMP-2表达影响的研究

Effect of hypoxia on HIF-1α,GLUT-1,and MMP-2 in human lung adenocarcinoma cells

目的:探讨人肺腺癌细胞株A549在缺氧条件下HIF-1α、GLUT-1、MMP-2的表达变化情况。方法:对肺腺癌细胞A549分别进行常规和氯化钴模拟缺氧培养,采用MTY法检测缺氧对A549细胞增殖的影响;Transwell侵袭小室测定法评价缺氧下A549细胞的侵袭力;分别采用RT-PCR法和western blot、免疫组化法测定缺氧不同时间HIF-1α和GLUT-1、MMP-2 mRNA和蛋白表达水平的变化。结果:A549细胞在缺氧条件下增殖能力和侵袭力较正常时增强。正常氧条件下的A549细胞即有一定水平的HIF-1α、GLUT-1、MMP-2的表达,随着缺氧时间的延长,HIF-1α和、GLUT-1、MMP-2 mRNA和蛋白水平呈升高趋势,与正常对照组比较,各缺氧组差异有统计学意义(P<0.01)。结论:氯化钴诱导缺氧可以上调HIF-1α、GLUT-1、MMP-2蛋白在肺腺癌细胞中的表达水平,加速肺癌细胞的生长和转移,使之进一步耐受缺氧。

Objective：To explore the changes of hypoxia-indacible factor let（HIF-1α） , glucose transporter protein-1 （ GLUT-1 ）, and matrix metalloproteinase-2 （MMP-2） in hypoxic human lung adenocarcinoma A549 cells. Methods.The human lung adenocarcinoma A549 cells were cultured in normoxic and CoCl2-induced hypoxic conditions respectively. MTF assay was used to test the effect of hypoxia on the proliferation of lung cancer ceils, and Transwell assay to determine the invasive capacity of hypoxic ceils. Reverse transcriptionpolymerase chain reaction analyses （RT-PCR） was used to detect the mRNA expression of HIF-1α, GLUT-1 and MMP-2 under hypoxia. The protein levels of HIF-1 α, GLUT-1, and MMP-2 were measured with immunohistochemical staining and western blot at different time points after hypoxia. Results：The proliferative and invasive capabilities of lung cancer cells increased under hypoxia condition. HIF-1 α, GLUT-1, and MMP-2 were expressed at certain levels in A549 cells under normoxic condition. After hypoxia exposure, the mRNA and protein expression of the three proteins increased in a time-dependent manner. The difference was significant between hypoxia group and normal control group （ P 〈 0.05 ）. Conclusion： Hypoxia induced by CoCl2 up-regulates the expression of HIF-1 α, GLUT-1 and MMP-2 in lung adenocarcinoma A549 cells and stimulates the proliferation and metastasis of lung cancer cells, thus further promotes their tolerance to hypo-xia.