Liang Fei, Zhu Xiao-jie, Wang Xiu-hong
Department of Biochemistry and Molecular biology, Harbin Medical University, Harbin 150086, China.
Zhonghua Zhong Liu Za Zhi. 2009 Aug;31(8):587-91.
To investigate the protective effect of overexpressed heme oxygenase-1 (HO-1) in hypoxia and the correlation between HO-1 overexpressoin and hypoxia inducible factor-1alpha (HIF-1alpha) in human hepatoma HepG2 cells.
The expressions of HO-1 and HIF-1alpha mRNA as well as the protein were detected by RT-PCR and Western blotting, respectively. MTT assay was used to examine the relative cell survival rate. The total superoxide dismutase (SOD) activity was examined with a SOD kit.
Hypoxia induced overexpression of HO-1 gene in HepG2 cells at transcriptional and translational levels. The relative survival rate of HepG2 cells under hypoxia was significantly decreased after the HO-1 protein overexpression was inhibited by ZnPPIX (P < 0.01). The total SOD activity of cells was significantly increased after cells were treated by hypoxia for 16 hours (P < 0.05), while decreased significantly by HO-1 inhibitor ZnPPIX treatment (P < 0.01). HIF-1alpha was upregulated under hypoxia. In addition, the HO-1 overexpression under hypoxia was decreased by HIF-1alpha inhibitor, while the HIF-1alpha expression level under hypoxia was not significantly changed after HO-1 expression was inhibited by ZnPPIX.
The overexpression of HO-1 in hypoxic HepG2 cells is HIF-1alpha-dependent or at least partly HIF-1alpha-dependent. The relative survival rate of hypoxic hepatoma cells was significantly decreased by HO-1 inhibitor treatment. The results of this study may offer new thought and drug target for the therapy of human hepatoma in the future.
研究血红素加氧酶-1（HO-1）过表达在缺氧环境下对人肝癌HepG2细胞的保护作用以及HO-1过表达与缺氧诱导因子-1α（HIF-1α）之间的相关性。
分别采用逆转录聚合酶链反应（RT-PCR）和蛋白质免疫印迹法检测HO-1和HIF-1α mRNA及蛋白的表达。采用MTT法检测细胞相对存活率。用超氧化物歧化酶（SOD）试剂盒检测总SOD活性。
缺氧可在转录和翻译水平诱导HepG2细胞中HO-1基因的过表达。当锌原卟啉（ZnPPIX）抑制HO-1蛋白过表达后，缺氧条件下HepG2细胞的相对存活率显著降低（P<0.01）。细胞经缺氧处理16小时后，总SOD活性显著升高（P<0.05），而经HO-1抑制剂ZnPPIX处理后显著降低（P<0.01）。缺氧条件下HIF-1α上调。此外，HIF-1α抑制剂可降低缺氧条件下HO-1的过表达，而ZnPPIX抑制HO-1表达后，缺氧条件下HIF-1α的表达水平无显著变化。
缺氧的HepG2细胞中HO-1的过表达依赖于HIF-1α或至少部分依赖于HIF-1α。HO-1抑制剂处理可显著降低缺氧肝癌细胞的相对存活率。本研究结果可能为未来人类肝癌的治疗提供新的思路和药物靶点。
