摘要目的:观察亚砷酸钠（NaAsO 2）长期作用于永生化人支气管上皮细胞（HBE细胞）致其恶性转化过程中，核因子-E2相关因子2（Nrf2）对细胞凋亡的调控作用。 方法:用含0.0和1.0 μmol/L NaAsO 2的培养基培养HBE细胞，分别为对照组和染砷组。用含1.0 μmol/L NaAsO 2处理HBE细胞至43代，建立恶性转化模型。检测细胞染砷后不同代数（0、1、8、15、22、29、36、43代）各指标的动态变化，包括用流式细胞仪检测细胞凋亡率，用蛋白免疫印迹法（Western blot）检测凋亡相关蛋白和Nrf2蛋白的表达情况。用Nrf2小干扰RNA（siRNA）转染恶性转化的HBE细胞（T-HBE细胞）来沉默Nrf2，验证Nrf2蛋白的沉默效果，并检测凋亡率和凋亡相关蛋白的表达。 结果:随着染砷代数增加，HBE细胞凋亡率呈下降趋势（0、1、8、15、22、29、36、43代分别为0.370 ± 0.029、0.443 ± 0.069、0.357 ± 0.046、0.330 ± 0.016、0.273 ± 0.050、0.160 ± 0.024、0.110 ± 0.022、0.097 ± 0.012， F趋势 = 22.981， P < 0.05），与0代细胞比较，第22、29、36、43代细胞的凋亡率均较低，差异均有统计学意义（ P均< 0.05）。随着染砷代数增加，染砷组细胞促凋亡蛋白半胱氨酸蛋白酶（caspase）-3、活化的caspase-3（cleaved-caspase-3）、转录因子C/EBP的同源蛋白（CHOP）、B淋巴细胞瘤-2基因（Bcl-2）相关X蛋白（Bax）的表达均呈下降的趋势（ F趋势 = 22.356、3.738、6.130、8.061， P均< 0.05），抗凋亡蛋白髓样细胞白血病-1蛋白（Mcl-1）、Bcl-2蛋白表达呈上升趋势（ F趋势 = 58.201、7.691， P均< 0.05）。分别与0代和同代对照组细胞比较，22代及以后染砷组细胞caspase-3、cleaved-caspase-3蛋白表达，15代及以后染砷组细胞CHOP、Mcl-1、Bcl-2蛋白表达，29代及以后染砷组细胞Bax蛋白表达，差异均有统计学意义（ P均< 0.05）；而各代染砷组细胞caspase-8、cleaved-caspase-8、caspase-12和cleaved-caspase-12蛋白表达比较，差异均无统计学意义（ P均> 0.05）。分别与0代和同代对照组细胞比较，8代及以后染砷组细胞Nrf2蛋白表达差异均有统计学意义（ P均<0.05）。与Con siRNA（对照）转染组T-HBE细胞比较，Nrf2 siRNA转染组T-HBE细胞的凋亡率较高（ P < 0.05）。与Con siRNA转染组T-HBE细胞比较，Nrf2 siRNA转染组T-HBE细胞的Nrf2、Bcl-2、Mcl-1蛋白表达均较低（ P均< 0.05），cleaved-caspase-3/caspase-3、caspase-3、cleaved-caspase-3、CHOP、Bax蛋白表达均较高（ P均< 0.05）。 结论:Nrf2可能通过Bcl-2、Mcl-1和Bax调控线粒体凋亡途径、通过CHOP调节内质网凋亡途径，抑制HBE细胞的凋亡，参与NaAsO 2致HBE细胞恶性转化的过程。

abstractsObjective:To observe the role of nuclear factor erythroid 2-related factor 2 (Nrf2) in regulating apoptosis during malignant transformation of human bronchial epithelial cells (HBE cells) induced by sodium arsenite (NaAsO 2). Methods:HBE cells were treated with 0.0 and 1.0 μmol/L NaAsO 2, which were control group and arsenic exposed group respectively. HBE cells were treated with 1.0 μmol/L NaAsO 2 for 43 passages to establish a malignant transformation model. The dynamic changes of indexes in different passages (0, 1st, 8th, 15th, 22nd, 29th, 36th, and 43rd) after exposure to NaAsO 2 were monitored, including the apoptosis rate detected by flow cytometry and apoptosis-related proteins and Nrf2 protein detected by Western blotting. Nrf2 siRNA was transfected into malignant transformed HBE cells (T-HBE cells) to silence Nrf2. The silencing effect of Nrf2 protein was verified. And, the apoptosis rate and apoptosis-related proteins were detected. Results:With the increase of arsenic exposure, the apoptosis rates of HBE cells decreased (0, 1, 8, 15, 22, 29, 36 and 43 passages were 0.370 ± 0.029, 0.443 ± 0.069, 0.357 ± 0.046, 0.330 ± 0.016, 0.273 ± 0.050, 0.160 ± 0.024, 0.110 ± 0.022, 0.097 ± 0.012, respectively, Ftrend = 22.981, P < 0.05). Compared with the 0 passage cells, the apoptosis rates of the 22nd, 29th, 36th and 43rd passages in the arsenite group were lower. The differences between them were statistically significant ( P < 0.05). With the increase of arsenic exposure, the expressions of pro-apoptotic proteins caspase-3, cleaved-caspase-3, C/EBP-homologous protein (CHOP) and B-cell lymphoma-2 (Bcl-2) associated X protein (Bax) showed downward trends ( Ftrend = 22.356, 3.738, 6.130, 8.061, P < 0.05), while the anti-apoptotic proteins myeloid cell leukemia 1 protein (Mcl-1) and Bcl-2 showed upward trends ( Ftrend = 58.201, 7.691, P < 0.05). Compared with the 0 passage and the control group of the same passage, from the 22nd passage of caspase-3, cleaved-caspase-3, from the 15th passage of CHOP, Mcl-1, and Bcl-2, from the 29th passage of Bax in the arsenite group, the differences of protein were statistically significant ( P < 0.05). However, there were no significant differences in caspase-8, cleaved-caspase-8, caspase-12 and cleaved-caspase-12 protein expressions in the arsenic group ( P > 0.05). Compared with the 0 passage and the control group of the same passage, from the 8th passage of Nrf2 proteins in the arsenite group, the differences of expressions were statistically significant ( P < 0.05). Compared with T-HBE cells transfected with Con siRNA (control), the apoptosis rate of T-HBE cells transfected with Nrf2 siRNA was higher ( P < 0.05). Compared with T-HBE cells transfected with Con siRNA, the expression levels of Nrf2, Bcl-2 and Mcl-1 in T-HBE cells transfected with Nrf2 siRNA were lower ( P < 0.05), while the expression levels of cleaved-caspase-3/caspase-3, caspase-3, cleaved-caspase-3, CHOP, and Bax were higher ( P < 0.05). Conclusion:Nrf2 may regulate mitochondrial apoptotic pathway through Bcl-2, Mcl-1 and Bax, and endoplasmic reticulum apoptotic pathway through CHOP, so as to inhibit the apoptosis of HBE cells and participate in the process of malignant transformation of HBE cells induced by NaAsO 2.