目的

探讨缺氧时肝癌细胞SMMC-7721中乏氧诱导因子-1α（HIF-1α）、己糖激酶II（HKII）表达及对细胞周期调控机制的影响。

方法

人肝癌细胞SMMC-7721 分成3组：缺氧12 h组、缺氧24 h 组及对照组。缺氧组分别在缺氧条件下(37℃,5%CO₂、2%O₂饱和度)培养12 h 和24 h，对照组置于正常氧浓度条件下(37℃,5%CO₂、21%O₂饱和度)培养24 h。流式细胞仪检测细胞周期分布，免疫组织化学SP法检测HIF-1α表达，荧光定量PCR法检测HKII mRNA表达量。

结果

缺氧状态下细胞周期分析处于G0/G1期细胞百分比明显增多，处于G₂/M期的细胞减少，并且可见到较明显的凋亡峰的形成，缺氧12 h 组、缺氧24 h组的G0/G1期比例显著高于对照组（P<0.05）；HIF-1α和HKII 在两组缺氧组与对照组比较，其表达量明显增加（P<0.05），缺氧24h组与缺氧12h组比较差异有统计学意义（P <0.05）。

结论

缺氧导致细胞阻滞在G0/G1期，HIF-1α可刺激人肝癌细胞HKII表达的增加，以HIF-1α为药物靶点可望成为治疗肝癌的重要新方法。

Objective

To investigate the impact of cell cycles arrest,expression of hypoxia-inducible factor-1α(HIF-1α)and hexokinaseII(HKII) in hepatocellular carcinoma cells under the hypoxic environment.

Methods

Human hepatocellular carcinoma cell line SMMC-7721 were divided into 3 groups: group of hypoxia for 12h and for 24h, and the control group. The two hypoxic groups were exposed to hypoxic condition(37℃,5%CO₂,2%O₂)for 12h and 24h respectively.The control group was exposed to normal oxygen condition(37℃, 5%CO₂, 21%O₂) for 24h. Cell cycle was examined by flow cytometry analysis; HIF-1α was measured by immunohistochemistry and HKII was detected by real-time quantitative PCR.

Results

Cells accumulated in the gap 0 and gap 1(G0/G1)phases(groups of hypoxia for 24h and 12h vs the control group,P<0.05);while reduced in the gap2 and mitosis(G2/M)phases of the cell cycle,and the apoptosis peak significantly occurred after hypoxia treatment.HIF-1α and HKII expression was significantly higher in hypoxic groups than that in control group(groups of hypoxia for 24h and 12h vs the control group,P<0.05;group of hypoxia for 24h vs for 12h,P<0.05).

Conclusion

A marked G0/G1 arrest of cell cycle and a high level of HKII expression are induced under the hypoxic environment, demonstrating the potential role of HIF-1α-targeted therapy in hepatocellular carcinoma.