INFIDELITY OF DNA REPAIR AND HEALTH RISK

DNA 修复的不忠和健康风险

• 批准号: 6246346
• 负责人: William W Au
• 金额: $ 2.71万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-01-06 至 1997-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6246346
• 关键词: DNA repair; cytogenetics; genetic techniques; human genetic material tag; method development; molecular genetics; tissue /cell culture; ultraviolet radiation

We intend to develop cytogenetic (broad based) and molecular (specific) assays for documentation of pollutant-induced DNA repair problem in cigarette smokers and in benzene-exposed workers. In the cytogenetic assay, blood lymphocytes from exposed individuals, matched controls and positive controls (cell line from zeroderma pigmentosum patients) will be irradiated with UV-light in vitro. These cells are challenged to repair the UV-light induced DNA damage. Abnormal/insufficient repair of the damage will cause the pollutant exposed cells and positive control cells to have more chromosome aberrations than those from normal controls. The repair defect will be confirmed biochemically as reduced uptake of labeled nucleotides. Three molecular assays will be developed for use in population studies to confirm and detect specific DNA repair problems. In one assay, cell homogenates from the same individuals will be used to repair UV-irradiated non-replicating plasmids, pSV2catSVgpt. Repair activities will be quantitated by measuring the uptake of labeled dAMP in recovered plasmids. In addition, alteration in the level of expression of two DNA repair enzymes, b-polymerase and O6-methylguanine-DNA methyltransferase, will be documented by using monoclonal antibodies in Western blot analyses. We expect that chronic exposure to toxic pollutants will affect most DNA repair mechanisms and the defect should be detected by our combination of assays. In addition, our protocols using unique and state of the art techniques should provide us with the sensitivities to detect the problem. This study will allow us to evaluate and recommend the most appropriate biomarker(s) for identification of potential human health risk from exposure to toxic pollutants.

我们打算发展细胞遗传学(基础广泛)和分子(特异性) 污染物引起的DNA修复问题的文献记录方法 吸烟者和苯接触者。在细胞遗传学中 检测，接触者的血淋巴细胞，匹配的对照组和 阳性对照(来自色素性零皮病患者的细胞系)将 体外用紫外光照射。这些细胞被挑战为 修复紫外光诱导的DNA损伤。维修不正常/不充分 这种损害会使污染暴露的细胞和阳性对照 细胞比正常细胞有更多的染色体异常 控制。修复缺陷将被生化确认为 减少对标记核苷酸的摄取。将有三种分子分析方法 开发用于人口研究，以确认和检测特定的 DNA修复问题。在一次化验中，细胞匀浆来自相同的 个人将被用来修复紫外线照射的非复制 表达载体pSV2catSVgpt。维修活动将通过以下方式进行量化 测定回收的质粒中标记的DAMP的摄取。在……里面 另外，两种DNA修复表达水平的改变 酶，b-聚合酶和O6-甲基鸟嘌呤-DNA甲基转移酶，将 通过在Western印迹分析中使用单抗来记录。 我们预计，长期接触有毒污染物将影响最大 DNA修复机制和缺陷应由我们的 多种检测方法的结合。此外，我们的协议使用唯一和 最先进的技术应该为我们提供对 检测问题。这项研究将使我们能够评估和推荐 识别潜在人类最合适的生物标志物(S) 暴露在有毒污染物中的健康风险。