环境污染物遗传毒性高通量、高内涵体外生物筛查方法的研究与应用
文献类型：学位论文
| 作者 | 言野 |
| 学位类别 | 博士 |
| 答辩日期 | 2013-11 |
| 授予单位 | 中国科学院研究生院 |
| 授予地点 | 北京 |
| 导师 | 马梅，王子健 |
| 关键词 | DNA损伤效应 染色体损伤效应 苯并三氮唑 苯并噻唑 水源水体 DNA-damaging effect Chromosom-damaging effect Benzotriazoles Benzothiazoles Source waters |
| 其他题名 | Research and application of high-throughput and high-content in vitro bioassays for screening environmental genotoxic pollutants |
| 学位专业 | 环境科学 |
| 中文摘要 | 遗传毒物是一类能引起有机体DNA或（和）染色体结构或（和）数量改变的物质，它们与许多疾病如新生婴儿畸形、各种癌症、帕金森症、阿兹海默症等相关。由于环境污染物可以通过各种途径与人相互作用，因此，它们的遗传毒性筛查与评价就显得尤为重要。本研究改进和建立了两种高通量体外遗传毒性实验组合，从DNA和染色体两个层次、多指标快速且全面的评价了两类环境污染物和全国多个水源水样遗传毒性的可能作用水平。 针对DNA损伤的检测，在已有微孔板实验基础上改进了SOS/umu实验方案，保证甚至提高检测灵敏度的前提下，将整个实验周期缩短至8小时内，为原来实验方法的1/2以下；针对染色体损伤的检测，建立了针对人胃癌细胞MGC-803和人肺癌细胞A549，基于图像自动获取、数据自动分析的微孔板高内涵筛选体外细胞微核实验。高内涵筛选可以在保证细胞完整性的情况下得到亚细胞结构相关信息，从而保证体外细胞微核实验可以获得客观的染色体损伤效应评价结果。 4种BTris（苯并三氮唑类）类物质遗传毒性测试的结果表明，5-甲基苯并三氮唑（5-TTri）和5,6-二甲基苯并三氮唑（XTri）既可以引起DNA水平也可以染色体水平上的遗传损伤，但苯并三氮唑（BTri）和5-氯代苯并三氮唑（Cl-BTri）仅可以引起染色体水平上的遗传损伤。9种BTs（苯并噻唑）类物质遗传毒性测试的结果表明，BT、FBT和MBT没有表现出遗传毒性；仅有2-羟基苯并噻唑（OHBT）和2-氨基苯并噻唑（ABT）可以引起DNA水平上的遗传损伤；而ABT、OHBT、2-溴苯并噻唑（BrBT）、2-甲硫基苯并噻唑（MTBT）、2-甲基苯并噻唑（MeBT）和2-氯苯并噻唑（CBT）可以引起染色体水平上的遗传损伤。 16个水源水体样品的遗传毒性筛查和评价结果表明，它们均具有一定的遗传毒性，或是在DNA水平上，或是在染色体水平水，或是两种都有。在长江6个水源水样中，通过非目标化合物筛查、数据收集和Read-across方法数据填补（data filling）我们发现了6种具有DNA损伤效应的遗传毒物和23种具有染色体损伤效应的遗传毒物。 因此，联合应用SOS/umu实验和基于高内涵筛选的体外细胞微核实验，可以有效的筛查和评价环境污染物和复杂环境样品的遗传毒性。 |
| 英文摘要 | Genotoxins are a class of chemicals which can induce damage to DNA or (and) chromosome, due to their relationships with many adverse health effects such as teratogenic effects, cancers, Parkinson's disease and Alzheimer's disease, the genotoxicity of environmental pollutants is really a need to be screened and evaluated. In this study, we modified and established two high-throughput in vitro genetic toxicity test, and applied them to screen and evaluate the genotoxicity of two types of environmental pollutants and source waters from China at both DNA-damaging and chromosome-damaging level in a high-content way. For detection of DNA damage, the microplate-based SOS/umu test was shorten to be within 8 hours which is less than 1/2 of the time used in published methods, and similar or higher sensitivity of typical genotoxins compared to published methods were obtained. For detection of chromosomal damage, microplate-based high-content screening in vitro MN tests employing human gastric carcinoma cell line (MGC-803) and human lung carcinoma cell line (A549) were developed. The automatic imaging, automatic data analysis and information involving subcellular structures were accomplished by the high-content screening and therefore objective results were guranteed in the newly developed MNt. The results of genotoxicity of BTris domenstrated that 5-TTri (5-Methybenzotriazole) and XTri (5,6-Dimethylbenzotriazole) showed genotoxicity at both DNA and chromosomal level while BTri and Cl-BTri (5-Chlorobenzotriazole) showed genotoxicity only at chromosomal level. No observed genotoxicity was obtained for BT, FBT and MBT, DNA-damaging effect was only induced by OHBT and ABT, and chromosome-damaging effect was induced by ABT, OHBT, BrBT and MTBT Cl-BTri to MGC -803, by MeBT to A549 and by CBT to both. The genotoxicity were showed either at DNA level or at chromosomal level or at both level by the 16 source waters. Using automatic screening of non-target organic pollutants, data collection and data filling by read-across method, 6 DNA-damaging agents and 23 chromosome-damage agents were found in the 6 source waters collected from Yangtze river. Therefore, the SOS/umu test combined with the high-content screening in vitro MNt can be used to quick but comprehensive screen and evaluate the genotoxicity of environmental pollutants and mixtures. |
| 公开日期 | 2015-06-16 |
| 源URL | [http://ir.rcees.ac.cn/handle/311016/13477]  |
| 专题 | 生态环境研究中心_环境水质学国家重点实验室 |
| 推荐引用方式 GB/T 7714 | 言野. 环境污染物遗传毒性高通量、高内涵体外生物筛查方法的研究与应用[D]. 北京. 中国科学院研究生院. 2013. |
入库方式： OAI收割
来源：生态环境研究中心
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