吴家兵^1,2,
杨永坚^1,,,
叶丹³,
杨文静³,
王先良^2,3,,,
钱岩²,
吕占禄²,
郭辰²,
梁豹^1,2,
赵淑莉⁴
1. 安徽医科大学公共卫生学院劳动卫生与环境卫生系, 合肥 230032;
2. 中国环境科学研究院环境基准与风险评估国家重点实验室, 北京 100012;
3. 中国疾病预防控制中心环境与健康相关产品安全所, 北京 100050;
4. 中国环境监测总站, 北京 100029

作者简介: 吴家兵(1989-),男,硕士研究生,研究方向为职业环境毒理学,E-mail:wujiabing159@sina.cn.

通讯作者: 杨永坚,yyj580719@163.com ; 王先良,xlwang@craes.org.cn ;

基金项目: 国家环保公益性行业科研专项(201309045)
国家自然科学基金(20907047)
国家重点基础研究发展(973)计划(2012CB525005)


中图分类号: X171.5

Lead Toxicity to Human Liver Cancer Cells Bel-7402 and Their Effect on DNA Methylation Levels at Promoter Region of EGFP Gene

Wu Jiabing^1,2,
Yang Yongjian^1,,,
Ye Dan³,
Yang Wenjing³,
Wang Xianliang^2,3,,,
Qian Yan²,
Lv Zhanlu²,
Guo Chen²,
Liang Bao^1,2,
Zhao Shuli⁴
1. School of Public Health, Anhui Medical University, Hefei 230032, China;
2. State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012, China;
3. Institute of Environmental Health and Related Product Safety, Chinese Center for Disease Control and Prevention, Beijing 100050, China;
4. China National Environmental Monitoring Center, Beijing 100029, China

Corresponding authors: Yang Yongjian,yyj580719@163.com ; Wang Xianliang,xlwang@craes.org.cn ;

CLC number: X171.5

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摘要:以体外培养人Bel-7402肝癌细胞为模型，研究铅的3种常见化合物氯化铅(PbCl₂)、乙酸铅(Pb (CH₃COO)₂)、硝酸铅(Pb (NO₃)₂)的细胞毒性和去甲基化表观遗传毒性。应用MTS方法检测细胞的存活率，以前期研究建立的评价方法评价铅化合物的去甲基化表观遗传毒性。结果显示，PbCl₂、Pb (CH₃COO)₂、Pb (NO₃)₂均会抑制Bel-7402细胞的增值，计算求得PbCl₂、Pb (CH₃COO)₂、Pb (NO₃)₂相应的50%细胞存活浓度(IC₅₀)值分别为2 524 μmol·L^-1、1 977 μmol·L^-1、1 899 μmol·L^-1；80%细胞存活浓度(IC₈₀)值分别为264 μmol·L^-1、221 μmol·L^-1、281 μmol·L^-1，通过对3种染毒物不同染毒浓度的细胞存活率进行随机区组设计的方差分析显示3种化合物间的差异无统计学意义(F=0.11；P=0.897)。去甲基化表观遗传毒性检测结果显示，PbCl₂、Pb (CH₃COO)₂、Pb (NO₃)₂均可观察到明显的去甲基化表观遗传毒性，其相对于5-氮杂-2-脱氧胞苷(5-Aza-CdR)的去甲基化表观遗传毒性当量分别为2.82E-03、1.50E-03、5.09E-04，三者间也无显著性差异。结果表明，铅化合物会使Bel-7402细胞的细胞存活率和转染进细胞的质粒上增强型绿色荧光蛋白基因启动子的DNA甲基化水平下降。
关键词: 铅/
Bel-7402细胞/
去甲基化/
MTS

Abstract:Human liver cancer Bel-7402 cells were used to estimate the cell toxicity and demethylation toxicity of 3 lead compounds including lead chloride (PbCl₂), lead acetate (Pb(CH₃COO)₂) and lead nitrate (Pb(NO₃). Cell viability was estimated firstly by MTS assay, and the demethylation toxicity of lead compounds was quantified by the method established and reported in earlier study. The results showed PbCl₂, Pb(CH₃COO)₂ and Pb(NO₃ all significantly inhibited the Bel-7402 cell proliferation and the IC₅₀ values were 2 524 μmol·L^-1, 1 977 μmol·L^-1, 2 524 μmol·L^-1; the IC₈₀ values were 264 μmol·L^-1, 221 μmol·L^-1, 281 μmol·L^-1. There was no significant difference of cell viability among 3 cell groups treated with different lead compounds (F=0.11; P=0.897). However significant epigenetic demethylation toxicity was observed for 3 compounds and their demethylation toxicity equivalency was 2.82E-03, 1.50E-03 and 5.09E-04 folds of 5-AZA-CdR, respectively. No significant difference was observed among their demethylation toxicity equivalency. The study revealed that lead compounds can inhibit Bel-7402 cell proliferation and induce obvious demethylation effect on DNA methylation levels at promoter region of EGFP gene.
Key words:lead/
Bel-7402 cells/
demethylation/
MTS.