绿茶对苯并芘致人胚肺成纤维细胞DNA损伤的保护作用

Protective Effect of Chinese Green Tea on B （a） P-induced DNA Damage in Human Embryonic Lung Fibroblasts

目的探讨绿茶对苯并芘（benzo[a]pyrene，B（a）P）诱导的DNA损伤的保护作用。方法①采用稳定转染抑癌基因p53-GFP质粒的p53-/-MEF细胞（p53-GFP细胞）分为DMSO对照组，B（a）P处理组，B（a）P和绿茶共处理组检测各组细胞中GFP荧光的激活情况；②将WI-38细胞分为DMSO对照组，DOX阳性对照组，B（a）P处理组，B（a）P和绿茶共处理组，用Western印迹检测不同处理组WI-38细胞中p-p53，γ-H2Ax，PARP和Caspase-3蛋白表达的改变。结果荧光结果显示，10μmol／L的B（a）P能够明显激活p53-GFP荧光，而绿茶和B（a）P共处理后荧光激活被抑制；用Western印迹检测发现，与DMSO对照组相比，B（a）P处理的wI-38细胞中P-p53和γ-H2AX蛋白表达水平升高，PARP和Caspase-3蛋白的切割增强，而绿茶和B（a）P共处理的WI-38细胞中p-p53和γ-H2AX蛋白表达水平较B（a）P处理组明显降低，PARP和Caspase-3蛋白的切割也明显减弱。结论本研究采用稳定转染p53-GFP质粒的p53-／-MEF细胞初步观察到绿茶可能对苯并芘诱导的DNA损伤具有保护作用。在WI-38细胞中的进一步研究证明，绿茶与B（a）P共处理能降低WI-38中p-p53和1-H2AX的表达水平，并且减弱PARP和Caspase-3蛋白的切割。提示，绿茶对苯并芘致WI-38细胞DNA损伤具有保护作用，其机制可能与抑制DNA损伤和凋亡通路有关。

Objective To explore the protective effect of Chinese green tea on DNA damage in- duced by benzo [ a] pyrene （B （a） P） . Methods ① The p53 -/-MEF cells were in vitro cul- tured and transfected with p53-GFP to obtain p53-GFP cells. Then the transfected cells were divided into DMSO control group, B （a） P treatment group and B （a） P ＋ Chinese green tea treatment group. GFP fluorescence was detected by fluorescence microscopy in each group. ② The WI-38 cells were divided into DMSO control group, DOX positive control group, B （a） P treatment group and B （a） P ＋ Chinese green tea treatment group. The expression levels of p-p53, γ-H2AX, PARP and caspase-3 were detected by Western blotting in each group. Results Fluorescence microscopy showed that10 μmol/L B （a） P could significantly activate the fluorescence in p53-GFP cells, which was suppressed after treatment with Chinese green tea. Western blotting revealed that the ex- pression levels of p-p53 and γ-H2AX were significantly increased in B （a） P-treated WI-38 cells when compared with the DMSO control group, and B （a） P could increase the cleaved levels of PARP and caspase-3. Chinese green tea could reduce the expression levels of p-p53 and γ-H2AXand attenuate the increased cleaved levels of PARP and caspase-3. Conclusion Chinese green tea can protect against B （a） P-induced DNA damage in WI-38 cells by inhibiting the pathways associ- ated with DNA damage and apoptosis.