Chen Chen, Lai Yanhao, Wu Mei, Zhang Zunzhen
Department of Environmental Health, West China School of Public Health Sichuan University, Chengdu 610041, China.
Wei Sheng Yan Jiu. 2010 Nov;39(6):669-73.
To explore the effect of DNA polymerase beta (pol beta) expression level on genotoxicity in mouse fibroblast cell induced by hydroquinone (HQ).
pol beta wild-type cells (pol beta +/+), pol beta null cells (pol beta -/-) and pol beta overexpressed cells (pol beta oe) were applied as a model cell system. At various concentration of HQ, the cell cytotoxicities were detected by MTT assay. The DCFH-DA fluorescent probe was used to examine the cellular ROS levels. The effects of DNA damage/repair and chromosomal damage were observed by comet assay and micronuleus assay respectively.
MTT assay showed that the doses of HQ increased, the cells viabilities were decreased. The concentrations of HQ that inhibited cell growth by 50% (IC50) in the pol beta -/- cell were more lower than those of other cells (P < 0.05). The cellular ROS level in the three kinds of cells were increased in a concentration dependent way after treated with HQ and it was more stronger in pol beta -/- cell than those in other two kinds of cells (P < 0.05). Comet assay and micronucleus assay showed that HQ induced DNA damage and increased micronucleus formation. DNA and chromosomal damage of pol beta -/- cell were more serious than those of other cells. The DNA damage repair capacities of pol beta oe cell were more stronger than pol beta +/+ and pol beta -/- cells.
Cellular ROS generation could be effectively induced by HQ, leading to DNA and chromosomal damage, pol beta overexpression could help cells response to oxidative damage and protect cells from genotoxicity induced by HQ.
探讨DNA聚合酶β(polβ)表达水平对氢醌(HQ)诱导的小鼠成纤维细胞遗传毒性的影响。
以polβ野生型细胞(polβ +/+)、polβ缺失型细胞(polβ -/-)和polβ过表达细胞(polβ oe)作为模型细胞系统。在不同浓度的HQ作用下,采用MTT法检测细胞毒性。用DCFH-DA荧光探针检测细胞内活性氧水平。分别通过彗星试验和微核试验观察DNA损伤/修复及染色体损伤情况。
MTT试验显示,随着HQ剂量增加,细胞活力降低。polβ -/-细胞中抑制细胞生长50%(IC50)的HQ浓度低于其他细胞(P < 0.05)。HQ处理后,三种细胞内的活性氧水平均呈浓度依赖性升高,且polβ -/-细胞中的升高幅度大于其他两种细胞(P < 0.05)。彗星试验和微核试验表明,HQ可诱导DNA损伤并增加微核形成。polβ -/-细胞的DNA和染色体损伤比其他细胞更严重。polβ oe细胞的DNA损伤修复能力强于polβ +/+和polβ -/-细胞。
HQ可有效诱导细胞产生活性氧,导致DNA和染色体损伤,polβ过表达有助于细胞应对氧化损伤并保护细胞免受HQ诱导的遗传毒性。