Francisco Dave C, Peddi Prakash, Hair Jessica M, Flood Brittany A, Cecil Angela M, Kalogerinis Peter T, Sigounas George, Georgakilas Alexandros G
Department of Biology, Thomas Harriot College of Arts and Sciences, East Carolina University, Greenville, NC 27858, USA.
Free Radic Biol Med. 2008 Feb 15;44(4):558-69. doi: 10.1016/j.freeradbiomed.2007.10.045. Epub 2007 Nov 20.
Oxidatively induced stress and DNA damage have been associated with various human pathophysiological conditions, including cancer and aging. Complex DNA damage such as double-strand breaks (DSBs) and non-DSB bistranded oxidatively induced clustered DNA lesions (OCDL) (two or more DNA lesions within a short DNA fragment of 1-10 bp on opposing DNA strands) are hypothesized to be repair-resistant lesions challenging the repair mechanisms of the cell. To evaluate the induction and processing of complex DNA damage in breast cancer cells exposed to radiotherapy-relevant gamma-ray doses, we measured single-strand breaks (SSBs), DSBs, and OCDL in MCF-7 and HCC1937 malignant cells as well as MCF-10A nonmalignant human breast cells. For the detection and measurement of SSBs, DSBs, and OCDL, we used the alkaline single-cell gel electrophoresis, gamma-H2AX assay, and an adaptation of pulsed-field gel electrophoresis with E. coli repair enzymes as DNA damage probes. Increased levels for most types of DNA damage were detected in MCF-7 cells while the processing of DSBs and OCDL was deficient in these cells compared to MCF-10A cells. Furthermore, the total antioxidant capacity of MCF-7 cells was lower compared to their nonmalignant counterparts. These findings point to the important role of complex DNA damage in breast cancer and its potential association with breast cancer development especially in the case of deficient BRCA1 expression.
氧化诱导的应激和DNA损伤与多种人类病理生理状况相关，包括癌症和衰老。诸如双链断裂（DSB）和非DSB双链氧化诱导的簇状DNA损伤（OCDL）（在相对DNA链上1-10 bp的短DNA片段内的两个或更多个DNA损伤）等复杂DNA损伤被认为是具有挑战性的细胞修复机制的抗修复损伤。为了评估暴露于与放疗相关的伽马射线剂量的乳腺癌细胞中复杂DNA损伤的诱导和处理情况，我们测量了MCF-7和HCC1937恶性细胞以及MCF-10A非恶性人乳腺细胞中的单链断裂（SSB）、DSB和OCDL。为了检测和测量SSB、DSB和OCDL，我们使用了碱性单细胞凝胶电泳、γ-H2AX检测以及一种以大肠杆菌修复酶作为DNA损伤探针的脉冲场凝胶电泳改进方法。与MCF-10A细胞相比，在MCF-7细胞中检测到大多数类型的DNA损伤水平升高，而这些细胞中DSB和OCDL的处理存在缺陷。此外，与非恶性对应细胞相比，MCF-7细胞的总抗氧化能力较低。这些发现表明复杂DNA损伤在乳腺癌中的重要作用及其与乳腺癌发展的潜在关联，特别是在BRCA1表达缺陷的情况下。
