SET基因缺陷对三氯乙烯诱导人正常肝细胞DNA甲基化水平改变的影响  

Effect of SET deficiency on the trichloraethylene-induced alteration of DNA methylation in human hepatic L-02 cells

作　　者：洪文旭[1] 黄爱博 许华[2] 张航[1] 王宏菊[3] 赵琼晖[3] 叶金波[1] 刘建军[1] 

机构地区：[1]深圳市疾病预防控制中心现代毒理学重点实验室,518055 [2]暨南大学药学院微生物与生化药学系 [3]深圳出入境检验检疫局食品检验检疫技术中心

出　　处：《中华预防医学杂志》2015年第3期206-211,共6页Chinese Journal of Preventive Medicine

基　　金：基金项目：国家自然科学基金（81273126、21102094）;广东省自然科学基金（$2012020010903）;广东省科技计划（20138031800001）;深圳市科技计划（201201027）

摘　　要：目的比较三氯乙烯（trichloroethylene）诱导人正常肝细胞（L-02细胞）和SET基因缺陷L-02细胞（SET缺陷细胞）中DNA甲基化相关指标的变化，探讨SET与三氯乙烯诱导的表观遗传调控作用之间的关系。方法选用前期建立的SET缺陷细胞为研究对象，使用三氯乙烯分别对L一02细胞和SET缺陷细胞进行处理，检测细胞增殖水平、细胞DNA甲基化水平和DNA甲基转移酶（DNAmethvltrans伯rases，DNMTs）活性的变化，通过Westernblot法从蛋白水平分析DNMTs（DNMT1、DNMT3a、DNMT3b）的表达变化。结果三氯乙烯处理L-02细胞和SET缺陷细胞24h后，两种细胞的增殖水平均呈现下降趋势。使用0、1．0、2．0、4．0和8．0mmol／L的三氯乙烯处理细胞后，L-02细胞的相对增殖水平分别为100．00±2．70、83．34±2．38、75．56±4．51、71．67±2．77、66．67±1．63（F=58．29，P〈0．001）；SET缺陷细胞的相对增殖水平分别为101．12±1．67、85．01±2．33、79．44±1．67、78．337±3．89、76．11±3．33（F=42．41，P〈0．001）。0、1．0、2．0、4．0和8．0mmol／L的三氯乙烯处理L-02细胞和SET缺陷细胞24h后，L-02细胞的DNA甲基化水平分别为3．77±0．08、3．48±0．08、3．38±0．10、3．14±0．15、2．91±0．07，呈下降趋势（F=212．87，P〈0．001）；SET缺陷细胞DNA甲基化水平分别为3．77±0．15、3．57±0．15、3．30±0．11、3．35±0．13呈下降趋势（F=79．32，P〈0．001）。L-02细胞经0、1．0、2、0、4．0、8．0mmol／L的三氯乙烯处理24h后，DNMTI蛋白的相对表达量分别为1．00±0．03、1．28±0．04、1．20±0．04、1．62±0．05、1．43±0．04，表达升高（F=103．00，P〈0．001）；而在SET缺陷细胞中DNMT1的相对表达量分别为1．00±0．04、0．96±0．02、1．19±0．05、0．85±0．03、0．83±0．03，出现下降（F=44．18，P〈0．001）。结论SET缺陷可以明显抑制三氯乙烯暴露引起的L-02�Objective To compare the DNA methylation-related alteration induced by trichloroethylene （TCE） in human hepatic L-02 cells （L-02 cells） and SET deficient cells, and reveal the role of SET on the mechanisms in TCE-induced epigenetic pathway. Methods The L-02 cells and pre- established SET deficient cells were treated with different TCE concentrations, and the changes of total cell viability, DNA methylation level and DNA methyltransferases （DNMTs） activity were measured, respectively. In addition, the TCE-induced alteration in the protein expression of DNMT1, DNMT3a and DNMT3b were analyzed by Western blotting. Results After treatment with TCE for 24 h, the cell proliferation level was significantly decreased in both cell lines. When concentrations of TCE were 0, 1.0, 2.0, 4. 0 and 8.0 retool/L, the proliferation levels of L-02 cells were 100. 00± 2.70, 83.34 ± 2.38,75.56±4.51, 71.67 ± 2. 77 and 66.67 ± 1.63, respectively （F = 58.29, P 〈 0.001）; the cell proliferation levels ot SET deficient cells were 101.12 ± 1.67, 85.01 ±2. 33,79. 44 ± 1.67, 78. 337 ±3.89 and 76. 11 ± 3.33, respectively （F = 42.41, P 〈 0. 001 ）. When concentration of TCE reached 4. 0 mmol/L, the difference of cell proliferation level between two groups was statistically significant （t = - 3.51; P =0. 013）. After treated by TCE for 24 h, the global DNA methylafion significantly decreased in both cell lines （F value was 212. 87 and 79.32, respectively, P 〈 0. 001 ）. The difference between two groups was not statistically significant. After treated by TCE for 24 h, the methyhransferases activities were significantly decreased in both cell cells （ F values were 77.92 and 113.80, respectively, P 〈 0. 001 ） . The SET deficiency could inhibit the decrease of methyltransferases activity under TCE treatment. When the concentration of TCE reached 8.0 mmol/L, the enzymatic activity of L-02 cells and SET deficient cells decreased to 67.61% ± 2.85% and 72. 97% ± 1.94% , respectively. The difference between

关 键 词：三氯乙烯 DNA甲基化 人正常肝细胞 SET缺陷 DNA甲基转移酶 

分 类 号：R114[医药卫生—卫生毒理学]