Interindividual differences in initial DNA repair capacity when evaluating H2O2-induced DNA damage in extended-term cultures of human lymphocytes using the comet assay
Interindividual differences in initial DNA repair capacity when evaluating H2O2-induced DNA damage in extended-term cultures of human lymphocytes using the comet assay
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DOI：
10.1007/s10565-007-9002-5
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发表时间：
2007-11-01
影响因子：
6.1
通讯作者：
Hellman, B.
中科院分区：
文献类型：
--
作者：
Andersson, M.;Stenqvist, P.;Hellman, B.
有人建议，在测试化学品的遗传毒性时，可以使用人类淋巴细胞的长期培养物作为基于转化细胞的细胞系的补充。为了研究基于不同献血者的培养物之间诱导的 DNA 损伤及其随后修复的模式是否存在显着差异，使用彗星测定在来自四个不同受试者的培养物中测量了过氧化氢 (H2O2) 诱导的 DNA 损伤。暴露于 0.25 mmol/L H2O2 10 分钟后，所有培养物中的 DNA 损伤均显着增加，而在 DNA 修复 30 分钟后，所有培养物中 H2O2 诱导的 DNA 损伤均显着减少。不同捐献者的损伤程度各不相同，尤其是修复后。使用 PCR 和 DNA 测序，对残留损伤最低和最高的供体淋巴细胞中 p53 基因的外显子 5 进行了测序。没有发现这样的突变。携带外显子 5 中 p53 突变的小鼠淋巴瘤 L5178Y 细胞被纳入作为参考。这些细胞对 H2O2 引起的 DNA 损伤不太敏感，而且 DNA 修复能力也相当低。从风险评估的角度来看，不同受试者的培养物之间 H2O2 诱导的 DNA 损伤和 DNA 修复能力的差异可能很重要，但在开发遗传毒性常规检测时显然不具有决定性的重要性。
It has been suggested that extended-term cultures of human lymphocytes could be used as a complement to cell lines based on transformed cells when testing the genotoxicity of chemicals. To investigate whether the pattern of induced DNA damage and its subsequent repair differs significantly between cultures based on different blood donors, hydrogen peroxide (H2O2)-induced DNA damage was measured in cultures from four different subjects using the comet assay. The DNA damage was significantly increased in all cultures after 10 min exposure to 0.25 mmol/L H2O2, and there was a significant decrease in the H2O2-induced DNA damage in all cultures after 30 min of DNA repair. The level of damage varied between the different donors, especially after the repair. Using PCR and DNA sequencing, exon 5 of the p53 gene was sequenced in the lymphocytes from the donors with the lowest and highest residual damage. No such mutation was found. Mouse lymphoma L5178Y cells carrying the p53 mutation in exon 5 were included as a reference. These cells were found to be less sensitive toward the H2O2-induced DNA damage, and they were also found to have a rather low DNA repair capacity. The demonstrated variation in H2O2-induced DNA damage and DNA repair capacity between the cultures from the different subjects may be important from a risk assessment perspective, but is obviously not of decisive importance when it comes to the development of a routine assay for genotoxicity.