Department of Chemistry, Indian Institute of Technology Patna, Patna 800 013, India.
J Biosci. 2011 Mar;36(1):105-16. doi: 10.1007/s12038-011-9008-2.
Clustered damage in DNA includes two or more closely spaced oxidized bases, strand breaks or abasic sites that are induced by high- or low-linear-energy-transfer (LET) radiation, and these have been found to be repair-resistant and potentially mutagenic. In the present study we found that abasic clustered damages are also induced in primary human fibroblast cells by low-LET X-rays even at very low doses. In response to the induction of the abasic sites, primary fibroblasts irradiated by low doses of X-rays in the range 10-100 cGy showed dose-dependent up-regulation of the DNA repair enzyme, ApeI. We found that the abasic clusters in primary fibroblasts were more lethal to cells when hApeI enzyme expression was down-regulated by transfecting primary fibroblasts with hApeI siRNA as determined by clonogenic survival assay. Endonuclease activity of hApeI was found to be directly proportional to hApeI gene-silencing efficiency. The DNA repair profile showed that processing of abasic clusters was delayed in hApeI-siRNA-silenced fibroblasts, which challenges the survival of the cells even at very low doses of X-rays. Thus, the present study is the first to attempt to understand the induction of cluster DNA damage at very low doses of low- LET radiation in primary human fibroblasts and their processing by DNA repair enzyme ApeI and their relation with the survival of the cells.
DNA 中的聚集性损伤包括两个或更多紧密间隔的氧化碱基、链断裂或无碱基位点，这些损伤是由高或低线性能量转移 (LET) 辐射诱导的，并且已经被发现具有修复抗性和潜在的诱变作用。在本研究中，我们发现低 LET X 射线甚至在非常低的剂量下也会在原代人成纤维细胞中诱导碱基缺失的聚集性损伤。为了应对碱基缺失的诱导，用低剂量 X 射线（范围为 10-100cGy）辐照的原代成纤维细胞显示出 DNA 修复酶 ApeI 的剂量依赖性上调。我们发现，用 hApeI siRNA 转染原代成纤维细胞下调 hApeI 酶表达后，原代成纤维细胞中的碱基缺失簇对细胞的杀伤作用更强，通过集落形成存活试验测定。发现 hApeI 的内切核酸酶活性与 hApeI 基因沉默效率成正比。DNA 修复谱显示，在 hApeI-siRNA 沉默的成纤维细胞中，碱基缺失簇的处理被延迟，这使得细胞即使在非常低剂量的 X 射线下也难以存活。因此，本研究首次尝试了解低 LET 辐射极低剂量下原代人成纤维细胞中簇状 DNA 损伤的诱导及其被 DNA 修复酶 ApeI 处理的情况，并探讨其与细胞存活的关系。
