摘要
基于酶联免疫原理,建立了定量测定细胞中磷酸化组蛋白H2AX(γH2AX)含量的新方法,并用于评价卷烟烟气的基因毒性。采用不同剂量的烟气气相物与烟气粒相物暴露于中国仓鼠卵巢细胞(CHO细胞),通过测定不同染毒时间下细胞中γH2AX含量水平变化,考察烟气染毒与细胞DNA损伤的剂量效应关系,以及不同烟气组分基因毒性的差异。此外,通过定量监测细胞中活性氧的变化,探讨卷烟烟气暴露诱发细胞DNA双链断裂损伤的作用机理。结果表明,随烟气暴露时间的增加,细胞中γH2AX的含量在几小时内达到峰值,然后逐渐下降;染毒浓度越高,细胞中γH2AX含量达到峰值的时间越长;随着烟气粒相物或气相物染毒浓度的增高,细胞中γH2AX含量水平呈上升趋势;烟气粒相物染毒相比烟气气相物对细胞中γH2AX含量的影响更为显著;此外,烟气成分直接染毒会引起细胞内活性氧的含量增加。细胞内活性氧的含量变化与细胞DNA双链断裂标志物γH2AX的含量变化有较强相关性,卷烟烟气中自由基成分可能是诱发细胞DNA双链断裂的原因之一。
Phospho-histone H2AX（TH2AX） has been widely used in vitro genotoxicity evaluation of ionizing radiation, carcinogenic substances and cigarette smoke as an important biomarker of DNA double stranded breaks （DSBs）. The study developed an enzyme-linked immunesorbent assay for detection of the content of γH2AX in cells to evaluate the genotoxicity of cigarette smoke. The study exposed CHO cells with cigarette smoke total particulate matter （TPM） and cigarette smoke condensate （CSC） of different dose, dose-effect relationship between smoke exposure and DNA damage components has been investigated by detecting the and the poisonous difference of cigarette smoke content change of γH2AX in cells under different exposure time and dose. In addition, reactive oxygen species （ROS） in cells were also detected to study the mechanism of cigarette smoke exposure induced DSBs. The experiment result showed that the required time for the level of γH2AX in cells grew to the peak prolonged with increased exposure dose. Once the level of γH2AX in cells grew to the peak value, then it decreased slowly; the level of γH2AX in cells rised with increasing dose of cigarette smoke; The effect of TPM on the content change of γH2AX was more than CSC; Moreover, smoke exposure could induce concentration increase of ROS in cells, and a good correlation of content change of ROS and γH2AX in cells were found, free radicals in cigarette smoke may be one major cause of DSBs.
出处
《分析化学》
SCIE
EI
CAS
CSCD
北大核心
2014年第3期320-325,共6页
Chinese Journal of Analytical Chemistry
基金
国家自然科学基金(Nos.21347002
21277174)资助~~
关键词
酶联免疫方法
基因毒性
Phospho-histone H2AX
Enzyme-linked immunesorbent assay
Genotoxicity
