Nrf2调控细胞凋亡在砷致HBE细胞恶性转化过程中的作用  

Role of Nrf2 regulating apoptosis in arsenite induced malignant transformation of HBE cells

作　　者：徐梦柔 李春春[1] 丁思 梅成镐 杨乾磊 毛佳渊 武婧 安艳 Xu Mengrou;Li Chunchun;Ding Si;Mei Chenghao;Yang Qianlei;Mao Jiayuan;Wu Jing;An Yan(Department of Toxicology,School of Public Health,Medical College of Soochow University,Suzhou 215123,China)

机构地区：[1]苏州大学医学部公共卫生学院卫生毒理学教研室,215123

出　　处：《中华地方病学杂志》2021年第8期627-634,共8页Chinese Journal of Endemiology

基　　金：国家自然科学基金(81872646、81811540034、81573173)。

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1.0μmol/L NaAsO_(2)的培养基培养HBE细胞,分别为对照组和染砷组。用含1.0μmol/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Objective: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 1.0μmol/L NaAsO_(2),which were control group and arsenic exposed group respectively.HBE cells were treated with 1.0μmol/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 statistic

关 键 词：亚砷酸盐类 永生化人支气管上皮细胞 恶性转化 凋亡 核因子-E2相关因子2 

分 类 号：R114[医药卫生—卫生毒理学]