LGK-974对人肝癌细胞系HepG2的辐射增敏作用

Effect of LGK-974 in enhancing the radiosensitivity of HepG2 hepatoma cells

观察LGK-974(CAS:1243244-14-5)对人肝癌细胞系HepG2的辐射增敏作用,并探讨可能的作用机制。应用四甲基偶氮唑盐(MTT)比色法检测LGK-974对HepG2细胞的生长抑制作用;细胞克隆形成法确定1.0mmol/L LGK-974对HepG2细胞放疗敏感性的影响;H2DCFH-DA探针流式细胞法检测细胞活性氧(Reactive oxygen species,ROS)水平;RT-PCR及Western Blot法检测Nrf2及其下游NQO-1、HO-1基因的转录水平和蛋白表达水平。结果显示,LGK-974抑制HepG2细胞的生长分裂和增殖,24、48、72 h的半抑制浓度(IC50)为4.3、2.14、0.536mmol/L(p<0.05);1.0mmol/L LGK-974对HepG2细胞的放射增敏比(SERD0)为1.37;LGK-974能增大辐射产生的ROS水平(p<0.05);照射后Nrf2及其下游基因HO-1、NQO-1的转录水平和表达水平均增加;LGK-974能降低辐射对Nrf2、HO-1和NQO-1的转录和表达水平的增加作用(p<0.05)。由此得出,LGK-974对人肝癌细胞系HepG2具有明显的辐射增敏作用,其机制可能是抑制Nrf2信号通路,阻止细胞缓解辐射产生的氧化压力。

To observe the effects of LGK-974 on radiosensitivity of HepG2 cells and explore its possible mechanism, MTT assay was applied to examine the growth inhibition of LGK-974 to HepG2; the radiosensitivity of HepG2 cells treated by 1.0 μmol/L LGK-974 was detected by clone formation assay; the reactive oxygen species level was examined by H2DCFH-DA probe flow eytometry method; the mRNA and protein levels of Nrf2, and the downstream gene 1-10-1, NQO-1 were accessed by RT-PCR and Western Blot, respectively. The results showedLGK-974 could inhibit HepG2 cells growth, and the IC5o of 24 h, 48 h and 72 h were 4.3 ~tmol/L, 2.14 ~tmol/L, and 0.536 ~tmol/L, respectively （p〈0.05）; the SERoo of 1.0 μmol/L LGK-974 was 1.37; LGK-974 could increase the ROS level produced by irradiation （p〈0.05）; irradiation could increase the transcription and expression of Nrf2, 1t0-1, and NQO-1, and LGK-974 could weaken these effects. The study reveals that LGK-974 can enhance the radiosensitivity of HepG2 cells. The mechanism might inhibit the Nrf2 signaling pathway and prevent cells reducing the oxidative stress produced by irradiation.