摘要
目的基于抑制Kelch样环氧氯丙烷相关蛋白1(Kelch-like ECH-assoicated protein 1,Keap1)基因,研究砷对人永生化角质形成细胞凋亡及抗氧化水平的影响。方法培养细胞72 h,分为5组:空白对照组、阴性对照(Keap1基因抑制未染砷)、Keap1基因抑制+低砷(2.9μmol/L)、Keap1基因抑制+中砷(5.8μmol/L)和Keap1基因抑制+高砷(29.0μmol/L)组;用流式细胞仪检测细胞凋亡率;用实时荧光定量聚合酶链式反应检测核因子E2相关因子2(nuclear factor erythroid 2-related factor 2,Nrf2)、Keap1 mRNA水平;采用酶联免疫吸附试验检测谷胱甘肽、血红素氧化酶、环氧化酶、S-腺苷甲硫氨酸与同型半胱氨酸蛋白水平。结果 Nrf2和Keap1 mRNA表达不全相等(均有P<0.05);与阴性对照组相比,低砷组S-腺苷甲硫氨酸和同型半胱氨酸蛋白表达上调(均有P<0.05),血红素氧化酶蛋白在低、中、高砷组均上调(均有P<0.05),环氧化酶蛋白在中、高砷组上调(均有P<0.05),谷胱甘肽蛋白在高砷组上调(F=7.24,P=0.001)。结论基于Keap1基因抑制,砷暴露上调抗氧化酶活性,提高抗氧化水平,细胞凋亡下降;随着砷剂量增加,Nrf2表达下调,抗氧化水平失衡,细胞凋亡增加。
Objective Based on the inhibition of Keapl （ Kelch-like ECH-associated protein 1 ） gene, we studied the effects of arsenic exposure on the apoptosis and amioxidant capacity of human immortalized keratinocyte （HaCaT） cells. Methods The cells were cultured for 72 h and was divided into 5 groups including blank control, negative control （ Keapl gene inhibition of untouched arsenic） and Keapl gene inhibition ＋ low-dose arsenic group ：（ 2. 9 μmol/L） , Keapl gene inhibition ＋ medium-dose arsenic group （5.8 μmol/L） and Keapl gene inhibition ＋ high-dose arsenic group （29. 0 μmol/L） ; The apoptosis rate was detected by flow cytometry. The expression of Nrf2 and Keapl mRNA was detected by Real-Time Quantitative PCR. The expressions of GSH, HO-1, COX-2, SAM and HCY were detected by ELISA. Results The expression of Nrf2 and Keapl mRNA was not equal in each groups （ all P 〈 0. 05 ）. Compared with the negative control group, the expression of SAM and HCY protein in low-dose arsenic group was significantly increased （ all P 〈 0.05） ; The protein expression of COX-2 was increased in medium-dose and high-dose arsenic groups （ all P 〈 0. 05 ） ; GSH protein expression was increased in high arsenic group （ F = 7.24, P = 0. 001 ）. Conclusions In the case of Keapl gene inhibition, the arsenic exposure increased the antioxidant enzyme activity, increased the level of antioxidation and decreased the apoptosis rate. With the increasing of arsenic dose, the expression of Nrf2 was down-regulated and the antioxidant level was irabalanced and death increased.
作者
白雪
马瑶
李煜
陈柔锦
吴军
郑玉建
BAI Xue;MA Yao;LI Yu;CHEN Rou-jin;WU Jun;ZHENG Yu-jian(Department of Environmental Health, School of Public Health, Xinjiang Medical University, Urumqi 830011, China;Department of Planned Immunization, Centers for Disease Control and Prevention, New Urban Area, Urumqi 830000, China;Department of Scientific Research, the Sixth Affiliated Hospital of Xinjiang Medical University, Urumqi 830002, China)
出处
《中华疾病控制杂志》
CAS
CSCD
北大核心
2018年第4期386-389,410,共5页
Chinese Journal of Disease Control & Prevention
基金
国家自然科学基金(81560513)
新疆医科大学研究生科研创新基金(CXCY2017013)
