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,苏州215123 [2]贵州医科大学环境污染与疾病监控教育部重点实验室,贵州医科大学公共卫生学院卫生毒理学教研室,贵阳550025
出 处:《中华地方病学杂志》2017年第4期241-245,共5页Chinese Journal of Endemiology
基 金:国家自然科学基金(81573173、81473008、81673203);贵州医科大学环境污染与疾病监控省部共建教育部重点实验室开放课题(GMU-2016-HJZ-03);江苏省高校基金(16KJB330009)
摘 要:目的观察1.0μmol/L亚砷酸钠(NaAsO2)长期作用于永生化人皮肤角质形成细胞(HacaT)时,细胞凋亡及凋亡相关蛋白的动态变化。方法用1.0μmol/LNaAsO2长期作用HaCaT细胞成功建立恶性转化模型,收集细胞恶性转化模型建立过程中0、1、7、14、21、28、35代细胞。采用流式细胞仪检测细胞凋亡率,蛋白免疫印迹法检测细胞凋亡相关蛋白,包括活化的半胱氨酸蛋白酶3、8(cleaved—caspase-3、8)、转录因子C/EBP的同源蛋白(Chop)、淋巴细胞瘤-2基因(Bcl-2)、Bcl-2相关X蛋白(Bax)。结果随着染砷代数的增加,细胞凋亡率呈明显的下降趋势,各代数间比较差异有统计学意义(F=26.770,P〈0.05),其中染砷第7、14、21、28、35代细胞的凋亡率(0.307±0.049、0.213±0.055、0.163±0.057、0.147±0.035、0.053±0.012)低于对照组0代(0.393±0.021,P均〈0.05)。染砷组促凋亡的cleaved—caspase-3、Chop、Bax蛋白和抗凋亡的Bcl-2蛋白表达各代间比较,差异有统计学意义(cleaved—caspase-3:1.000±0.000、1.030±0.027、1.104±0.069、1.016±0.087、0.838±0.075、0.753±0.082、0.677±0.073;Chop:1.000±0.000、1.059±0.018、0.934±0.095、0.976±0.216、0.793±0.136、0.651±0.042、0.564±0.056;Bax:1.000±0.000、1.069±0.037、1.028±0.042、0.954±0.118、0.641±0.135、0.531±0.132、0.429±0.085;Bcl-2:1.000±0.000、1.072±0.023、1.249±0.134、1.334±0.143、1.633±0.221、1.507±0.152、1.461±0.145.F=7.730、7.355、27.802、12.438,P均〈0.05),且21代及以后与对照组0代(1.000±0.000)和同代对照组(1.000±0.000)比较,差异有统计学意义(P均〈0.05);而cleaved.caspase-8蛋白各代间比较,差异无统计学意义(F=0.832,P〉0.05)。结论低剂量NaAsO:可能通Objective To explore the mechanism of cell apoptosis of immortalized human keratinocytes (HaCaT cells) and protein expression related to this process after long term exposure to sodium arsenite (NaAsO2, 1.0 μmol/L). Methods Malignant transformation model was set up through long-term exposure of HaCaT cells to 1.0 μmol/L NaAsO2. Cell passage for 0, 1, 7, 14, 21, 28 and 35 generations in the process of malignant transformation were collected for measurement of cell apoptosis rate by flow cytometry, and apoptosis related proteins by Western blotting, including activation of cysteine protease 3, 8 (cleaved-caspase-3, 8), C/EBP homologous protein (CHOP), B-cell leukemia/lymphoma 2 (Bcl-2), and Bcl-2 associated X protein (Bax). Results Along with the arsenite treatment, the apoptosis levels were significantly decreased (F = 26.770, all P 〈 0.05), the apoptosis levels (0.307 ± 0.049, 0.213 ± 0.055, 0.163 ± 0.057, 0.147 ± 0.035, 0.053 ± 0.012) of the 7th, 14th, 21st, 28th and 35thgenerations ofcells after arsenite treatment were lower than that of control group of the 0th generation (0.393 ± 0.021, all P 〈 0.05). Compared between generations, there were statistical differences of the protein expression levels of cleaved-caspase-3, Chop, Bax and Bcl-2 in arsenite group (cleaved-caspase-3:1.000 ± 0.000, 1.030 ± 0.027, 1.104 ± 0.069, 1.016 ± 0.087, 0.838 ± 0.075, 0.753 ± 0.082, 0.677 ± 0.073; Chop: 1.000 ± 0.000, 1.059 ± 0.018, 0.934 ± 0.095, 0.976 ± 0.216, 0.793 ± 0.136, 0.651 ± 0.042, 0.564 ± 0.056; Bax: 1.000 ± 0.000, 1.069 ± 0.037, 1.028 ± 0.042, 0.954 ± 0.118, 0.641 ± 0.135, 0.531 ± 0.132, 0.429 ± 0.085; Bcl-2:1.000 ± 0.000, 1.072 ± 0.023, 1.249 ± 0.134, 1.334 ± 0.143, 1.633 ± 0.221, 1.507 ± 0.152, 1.461 ± 0.145, F = 7.730, 7.355, 27.802, 12.438, all P 〈 0.05), compared with control group of the 0th generation (1.000 ± 0.000) and the same generation control group (1.000 ± 0.000), after the 21st generation, the difference
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