邻苯二甲酸二（2-乙基）己酯长期低剂量染毒对HepG2细胞DNA甲基化的影响

Effects of long term and low dose Di-（2-ethylhexyl） phthalate exposure on global genome DNA methylation in HePG2 cells

目的探索邻苯二甲酸二（2-乙基）己酯（DEHP）长期低剂量染毒对HepG2细胞全基因组DNA甲基化和细胞毒性的影响。方法用1．5、15．0、150．0μmol／L的DEHP对HepG2细胞进行24h持续染毒，连续处理20代后，对细胞的DNA甲基转移酶1（DNMT1）的基因和蛋白表达、基因组甲基化水平、凋亡水平和细胞周期进行检测，比较DEHP染毒细胞与正常对照细胞的差异。结果HepG2细胞经DEHP染毒处理后，DNMT1的mRNA与蛋白表达水平均明显下降，差异有统计学意义（P〈0．05或P〈0．01）；随着DEHP染毒剂量的增加，15．0、150．0μmol／L染毒组细胞的基因组甲基化水平明显下调，差异行统计学意义（P〈0．05和P〈0．01）；150．0μmol／L染毒组细胞晚期凋亡率明显高于对照组，差异有统计学意义（P〈0．05）；细胞周期无显著变化。结论HepG2细胞在接受长期低剂量的DEHP染毒后。细胞基因组DNA甲基化水平明显下调。最高剂量的DEHP长期染毒能诱导细胞出现一定程度的凋亡，DEHP具有细胞毒性，但其对细胞周期没有明显影响。

Objective To explore the potential effects of long-term and low-dose Di-（2-ethylhexyl） phihalate （DEHP） exposure on whole genome DNA methylation status and cytotoxicity of HePG2 cells. Methods HePG2 cells were exposed to 1.5,15.0 and 150.0 μmol/l, DEHP for 24 hours, after continuous exposure for 20 generalions, mRNA and protein expression level of DNA （cytosine-5）-methyltransferase 1 （DNMT 1）, whole genome DNA methylation, eell apoptosis levels and cell cycle were determined in both DEHP exposed cells and control cells. Results After DEHP exposure, the mRNA and protein expression levels of DNMT 1 were decreased significantly （P〈0.05 or P〈0.01 ）. The genome DNA methylation levels of HePG2 cells were down-regulated along with the increasing of DEHP exposure level （ 15.0, 150.0 μmol/L, P〈0.05 and P〈0.01 ）. In term of cell apoptosis rates, only the late-stage cell apoptosis rates of the highest DEHP dosage group （150.0 μmol/L DEHP） were observed to have a significant increase （P〈0.05）. There were no significant alterations in term of cell cycle. Conclusion After long term and low dose DEHP exposure, the whole genome DNA methylation levels of HePG2 cells were down regulated obviously, which might be one of the most important tnxie mechanism uf DEHP to induce pathophysiologie changes. Meanwhile, a certain content of cell apoptosis were observed in highest dosage group of DEHP exposure, which showed cytotnxicty of DEHP. However, there are no significanl effects of DEHP exposure on cell cycles.