缺氧时肝癌HepG2细胞HIF-1α对GLUT-1表达的影响及机制探讨

Study on effect of HIF-1α on expression of GLUT-1 in human hepatocellular carcinoma HepG2 cells cultured under hypoxia

目的探讨缺氧状态下肝癌HepG2细胞中HIF-1α表达对GLUT-1的影响。方法 HepG2人肝癌细胞株,以DMEM培养基加胎牛血清培养,分为常氧对照组(21%O2),缺氧组(1%O2),常氧加吲哚美辛组,缺氧加吲哚美辛组。收获细胞后,分别行免疫荧光法检测细胞内HIF-1α的表达和核转位;Western blot检测各组的HIF-1α、GLUT-1蛋白表达;RT-PCR检测各组的GLUT-1mRNA含量。结果缺氧时HIF-1α(蛋白:68.25±11.72)和GLUT-1(mRNA:0.7424±0.1127;蛋白:70.33±12.83)在肝癌HepG2细胞中的表达增高(P<0.05);缺氧时以吲哚美辛抑制HIF-1α蛋白(36.34±9.50)稳定和积聚后,GLUT-1 mRNA(0.3710±0.1016)及蛋白(38.46±12.20)表达受到抑制(P<0.05)。结论缺氧可以上调HIF-1α和GLUT-1在肝癌HepG2细胞中的表达;HIF-1α可以调控其下游靶基因GLUT-1的转录和表达,影响糖酵解和细胞能量的产生。

Objective To investigate the effect of HIF-lα on the expression of GLUT-1 in human hepatocellular carcinoma HepG2 cells cultured under hypoxia. Methods Hepatoma carcinoma cell line HepG2, cultivated in DMEM medium with fetal bovine serum, was divided into control group（ normal oxygen, 21% O2 ） , hypoxia group（ 1% O2 ） , normal oxygen with indomethacin group and hypoxia with indomethacin group. After the cells were harvested, the expression and nucleus translocation of HIF-lα in cells of each group were investigated through immunofluorescence method. GLUT-1 protein and HIF-let protein in each group were tested by Western blot,respectively. The amount of GLUT-1 mRNA in each group was tested by RT-PCR. Results The expressions of HIF-lα （ protein ：68.25±11.72 ） and GLUT-1 （ mRNA ： 0. 7424 ± 0. 1127 ; protein ： 70.33 ±12.83 ） increased in hepatocellular carcinoma HepG2 cells cultured under hypoxia（P 〈 0.05 ） , and meanwhile the expressions of GLUT-1 mRNA（0. 3710 ± 0. 1016） and protein （38.46± 12. 20） were reduced since the stabilization and accumulation of HIF-let protein （36. 34± 9.50） were inhibited by indomethacin in anaerobic condition（P 〈 0.05）. Conclusions Hypoxia condition can upregulate the expressions of GLUT-1 and HIF- lα in hepatoma carcinoma HepG2 cells. HIF-lα can regulate the transcription and expression of GLUT-1, thereby influence the glycolysis and cellular energy production.