Department of Oncology, Institute of Clinical Sciences, University of Gothenburg, Sahlgrenska University Hospital, Göteborg, Sweden.
Radiat Res. 2010 Mar;173(3):272-82. doi: 10.1667/RR1891.1.
Clustered DNA damages are induced by ionizing radiation and are defined as two or more lesions within one or two helical turns. The aim of this study was to investigate the induction and repair of clustered DNA damage in cells with emphasis on the influence of structural differences in the chromatin organization. Human fibroblasts were irradiated with X rays and induced DSBs and clustered damages were quantified using pulsed-field gel electrophoresis combined with postirradiation incubation with the base excision repair endonuclease Fpg, which recognizes oxidized purines and cleaves the strand at sites inducing strand breaks. Hence clustered damages appear in enzyme-treated samples as additional DSBs. The chromatin was modified by different pretreatments that resulted in structures with varying compactness and levels of free radical scavenging capacity. We found that the induction of DSBs and clustered damages increased linearly with dose in all structures and that both types of lesions were allocated randomly within the nucleus. The induction yields increased with decreasing compactness of chromatin, and the chromatin effect was larger for clustered lesions than for DSBs. Clustered damages were processed efficiently with a fast and a slow repair component similar to that for induced DSBs.
簇状 DNA 损伤是由电离辐射诱导的，定义为一个或两个螺旋圈中的两个或多个损伤。本研究的目的是研究细胞中簇状 DNA 损伤的诱导和修复，重点研究染色质组织结构差异的影响。用 X 射线照射人成纤维细胞，诱导双链断裂，并使用脉冲场凝胶电泳结合碱基切除修复内切酶 Fpg 进行后续孵育来定量检测簇状损伤，Fpg 可以识别氧化嘌呤，并在诱导链断裂的位点切割链。因此，在酶处理的样品中，簇状损伤表现为额外的双链断裂。通过不同的预处理来修饰染色质，得到不同紧密程度和自由基清除能力的结构。我们发现，在所有结构中，双链断裂和簇状损伤的诱导都随剂量呈线性增加，并且两种损伤类型都在核内随机分布。随着染色质紧密程度的降低，诱导产率增加，簇状损伤的染色质效应大于双链断裂。簇状损伤的修复与诱导的双链断裂相似，具有快速和缓慢的修复成分。
