Nrf2信号通路在亚砷酸钠致L-02细胞氧化损伤过程中的作用

Objective To explore the role of nuclear factor-E2-related factor 2 (Nrf2) signaling pathway in the process of oxidative damage of human normal liver cells (L - 02 cells) caused by sodium arsenite (NaAsO₂), and to provide experimental evidence for the research on the mechanism of oxidative damage of liver injury caused by arsenic. Methods L - 02 cells were cultured in vitro and treated with 0 (control), 25, 50, 75, 100, 125, 150 µmol/L NaAsO₂ for 24 h respectively. The cell survival rate was detected by CCK8 method. The half inhibitory concentration (IC₅₀) was calculated according to the cell survival rate. L - 02 cells were grouped and experimented with 0, 1/8, 1/4, 1/2 times the dose of IC₅₀ of NaAsO₂ respectively. The protein expression of Nrf2 signaling pathway - related factors Nrf2, heme oxygenase - 1 (HO - 1), quinone oxidoreductase 1 (NQO1), glutathione peroxidase 1 (GPx1) in L - 02 cells and cell nuclei was detected by Western blot. Results The results of CCK8 experiment showed that the survival rates of L - 02 cells in the 25, 50, 75, 100, 125, 150 µmol/L NaAsO₂ groups [(69.53 ± 0.06)%, (41.33 ± 0.08)%, (23.65 ± 0.04)%, (26.51 ± 0.04)%, (31.63 ± 0.01)%, (26.24 ± 0.02)%] were significantly lower than that of the control group [(100.00 ± 0.00)%, P < 0.05 for all]; the IC₅₀ of cell survival rate was 40 µmol/L, and the doses of NaAsO₂ in the grouped experiment were 0 (control), 5, 10, 20 µmol/L respectively. The results of Western blot detection showed that compared with the control group, the protein levels of Nrf2, HO - 1 in L - 02 cells and HO - 1 in the nuclei of L - 02 cells in the 5, 10, 20 µmol/L NaAsO₂ groups were significantly increased (P < 0.05 for all); the protein level of GPx1 in L - 02 cells in the 10, 20 µmol/L NaAsO₂ groups was significantly decreased (P < 0.05 for all), and the protein level of Nrf2 in the nuclei of L - 02 cells was significantly increased (P < 0.05 for all); the protein level of NQO1 in the nuclei of L - 02 cells in the 5 µmol/L NaAsO₂ group was significantly increased (P < 0.05). Conclusion NaAsO₂ has an impact on the expression of factors related to the Nrf2 signaling pathway in L - 02 cells, and the mechanism of its oxidative damage to L - 02 cells may be related to the Nrf2 signaling pathway.

目的探讨核因子-E2相关因子2(Nrf2)信号通路在亚砷酸钠(NaAsO_2)致人正常肝细胞(L-02细胞)氧化损伤过程中的作用,为砷致肝损伤的氧化损伤作用机制研究提供实验依据。方法体外培养L-02细胞,分别以0(对照)、25、50、75、100、125、150 µmol/L NaAsO_2理细胞24 h,采用CCK8法检测细胞存活率;根据细胞存活率计算半抑制浓度(IC_(50)),分别以IC_(50)的0、1/8、1/4、1/2倍剂量NaAsO_2处理L-02细胞进行分组实验。采用蛋白免疫印迹法(Western blot)检测L-02细胞中和细胞核内Nrf2信号通路相关因子Nrf2、血红素氧化酶-1(HO-1)、醌氧化还原酶1(NQ01)、谷胱甘肽过氧化物酶1(GPx1)的蛋白表达情况。结果CCK8实验结果显示,25、50、75、100、125、150 µmol/L NaAsO_2组的L-02细胞存活率[(69.53 ± 0.06)%、(41.33 ± 0.08)%、(23.65 ± 0.04)%、(26.51 ± 0.04)%、(31.63 ± 0.01)%、(26.24 ± 0.02)%]明显低于对照组[(100.00 ± 0.00)%,P均 < 0.05];细胞存活率的IC_(50)为40 µmol/L,分组实验的NaAsO_2剂量分别采用0(对照)、5、10、20 µmol/L。Western blot检测结果显示,与对照组比较,5、10、20 µmol/LNaAsO_2组L-02细胞中Nrf2、HO-1及L-02细胞核内HO-1蛋白水平显著升高(P均< 0.05);10、20 µmol/L NaAsO_2组L-02细胞中GPxl蛋白水平显著降低(P均<0.05),L-02细胞核内Nrf2蛋白水平显著升高(P均<0.05);5 µmol/L NaAsO_2组L-02细胞核内NQ01蛋白水平显著升高(P< 0.05)。结论NaAsO_2对L-02细胞中Nrf2信号通路相关因子的表达有影响,其致L-02细胞氧化损伤的作用机制可能与Nrf2信号通路有关。