Development and field-test validation of an assay for DNA repair in circulating human lymphocytes.
Development and field-test validation of an assay for DNA repair in circulating human lymphocytes.
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发表时间：
1991-11
期刊：
影响因子：
11.2
通讯作者：
W. F. Athas;M. Hedayati;Genevieve M. Matanoski;Evan R. Farmer;Lawrence Grossman
中科院分区：
文献类型：
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作者：
W. F. Athas;M. Hedayati;Genevieve M. Matanoski;Evan R. Farmer;Lawrence Grossman
一种测量核苷酸切除修复对紫外线照射和化学诱导的DNA损伤的反应的方法已经开发，验证，并在培养的人类淋巴细胞中进行了现场测试。该方法适用于基于人群的筛查，并应促进未来寻求调查DNA修复熟练程度与癌症易感性之间关系的流行病学研究。这种努力的动力来自于这样一种建议，即遗传性疾病色素性干皮病中皮肤癌的高发病率表现为患者细胞修复被模拟紫外线剂损伤的DNA的能力降低。在该实验中，利用deae -葡聚糖/DNA复合短期转染条件，将含有氯霉素乙酰转移酶（cat）报告基因的受损的、不复制的重组质粒DNA引入淋巴细胞。在40小时的修复/表达潜伏期后，对受损细菌cat基因的切除修复按比例进行监测，作为cat酶活性再激活的功能。该方法的有效性表明，该试验能够区分严重（互补组A和D）和中度（互补组C）的着色性干皮病病毒转化淋巴细胞系的切除修复缺陷和修复熟练细胞系。将染性干皮病a患者受丝裂原刺激的外周血淋巴细胞培养物与健康人受丝裂原刺激的外周血淋巴细胞同时测定，也观察到类似的结果。将这种DNA修复试验作为一种现场试验，在一组经过中试的基底细胞癌患者和无癌对照中进行了调整，初步确定了一个特定的亚群，由于光化学（UV）损伤的质粒DNA的修复显著减少，因此有患这种疾病的风险。
A method for measuring nucleotide excision repair in response to UV irradiation and chemical-induced DNA damage has been developed, validated, and field tested in cultured human lymphocytes. The methodology is amenable to population-based screening and should facilitate future epidemiological studies seeking to investigate associations between DNA repair proficiency and cancer susceptibility. The impetus for such endeavors derives from the suggestion that the high incidence of skin cancer in the genetic disorder xeroderma pigmentosum is manifested as a result of the reduced capacity of patients' cells to repair DNA damaged by UV-mimetic agents. For the assay, damaged, nonreplicating, recombinant plasmid DNA harboring a chloramphenicol acetyltransferase (cat) reporter gene is introduced into lymphocytes by using a DEAE-dextran/DNA complex short-term transfection conditions. Excision repair of the damaged bacterial cat gene is monitored proportionately as a function of reactivated CAT enzyme activity following a 40-h repair/expression incubation period. The validity of the approach was indicated by the ability of the assay to discriminate xeroderma pigmentosum virus-transformed lymphocyte cell lines of both severe (complementation groups A and D) and moderate (complementation group C) excision repair deficiencies from repair-proficient cell lines. Similar results were observed when a mitogen-stimulated peripheral blood lymphocyte culture from an xeroderma pigmentosum A patient was assayed concurrently with mitogen-stimulated peripheral blood lymphocytes obtained from healthy individuals. Adaptation of this DNA repair assay as a field test in a pilot-tested select group of basal cell carcinoma patients and cancer-free controls led to the preliminary identification of a specific subset at risk for this disease as a consequence of significant reduction to the repair of photochemically (UV)-damaged plasmid DNA.