Analysis of trichloroethylene-induced global DNA hypomethylation in hepatic L-02 cells by liquid chromatography-electrospray ionization tandem mass spectrometry.
Analysis of trichloroethylene-induced global DNA hypomethylation in hepatic L-02 cells by liquid chromatography-electrospray ionization tandem mass spectrometry.
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DOI：
10.1016/j.bbrc.2014.03.015
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发表时间：
2014-04
影响因子：
3.1
通讯作者：
Hang Zhang;W. Hong;Jinbo Ye;Xifei Yang;X. Ren;Aibo Huang;Lin-qing Yang;Li Zhou;Hai-yan Huang-Hai-yan-Huan
中科院分区：
文献类型：
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作者：
Hang Zhang;W. Hong;Jinbo Ye;Xifei Yang;X. Ren;Aibo Huang;Lin-qing Yang;Li Zhou;Hai-yan Huang-Hai-yan-Huan
三氯乙烯（TCE）是一种主要的职业和环境污染物，近年来已被发现与实验动物和培养细胞的异常表观遗传变化有关。已知TCE会导致严重的肝毒性;然而，表观遗传学改变与TCE诱导的肝毒性之间的关联尚未得到充分研究。DNA甲基化是由DNA甲基转移酶（DNMT）催化的一种主要的表观遗传修饰，在调节许多细胞过程中起着关键作用。在这项研究中，我们分析了TCE诱导的人肝L-02细胞的DNA甲基化和DNMT酶活性的影响。采用液相色谱-电喷雾串联质谱（LC-ESI-MS/MS）联用技术，对三氯乙烯（TCE）诱导的L-02细胞DNA甲基化改变进行了检测。通过监测5-mdC的m/z242.1（分子离子）/126.3（碎片离子）和dG的m/z268.1/152.3的跃迁对，在多反应监测（MRM）模式下完成定量。5-mdC与dG的标准曲线相关系数大于0.9990。日内和日间相对标准偏差值（RSD）分别为0.53-7.09%和0.40- 2.83%。我们发现TCE暴露能够显著降低L-02细胞DNA甲基化水平，抑制DNMT活性。我们的研究结果不仅揭示了TCE暴露与表观遗传学改变之间的关联，而且还提供了一种基于质谱的替代方法，用于快速准确地评估哺乳动物细胞中化学诱导的DNA甲基化改变。
Trichloroethylene (TCE), a major occupational and environmental pollutant, has been recently associated with aberrant epigenetic changes in experimental animals and cultured cells. TCE is known to cause severe hepatotoxicity; however, the association between epigenetic alterations and TCE-induced hepatotoxicity are not yet well explored. DNA methylation, catalyzed by enzymes known as DNA methyltransferases (DNMT), is a major epigenetic modification that plays a critical role in regulating many cellular processes. In this study, we analyzed the TCE-induced effect on global DNA methylation and DNMT enzymatic activity in human hepatic L-02 cells. A sensitive and quantitative method combined with liquid chromatography and electrospray ionization tandem mass spectrometry (LC–ESI-MS/MS) was validated and utilized for assessing the altered DNA methylation in TCE-induced L-02 cells. Quantification was accomplished in multiple reaction monitoring (MRM) mode by monitoring a transition pair ofm/z242.1 (molecular ion)/126.3 (fragment ion) for 5-mdC andm/z268.1/152.3 for dG. The correlation coefficient of calibration curves between 5-mdC and dG was higher than 0.9990. The intra-day and inter-day relative standard derivation values (RSD) were on the range of 0.53–7.09% and 0.40–2.83%, respectively. We found that TCE exposure was able to significantly decrease the DNA methylation and inhibit DNMT activity in L-02 cells. Our results not only reveal the association between TCE exposure and epigenetic alterations, but also provide an alternative mass spectrometry-based method for rapid and accurate assessment of chemical-induced altered DNA methylation in mammal cells.