Department of Plant, Food, and Environmental Sciences, Faculty of Agriculture, Dalhousie University, Truro, NS, Canada.
Department of Pathology, Faculty of Medicine, Dalhousie University, Halifax, NS, Canada.
Oxid Med Cell Longev. 2017;2017:1767198. doi: 10.1155/2017/1767198. Epub 2017 Jun 18.
Human neoplastic transformation due to DNA damage poses an increasing global healthcare concern. Maintaining genomic integrity is crucial for avoiding tumor initiation and progression. The present study aimed to investigate the efficacy of an apple flavonoid fraction (AF4) against various carcinogen-induced toxicity in normal human bronchial epithelial cells and its mechanism of DNA damage response and repair processes.
AF4-pretreated cells were exposed to nicotine-derived nitrosamine ketones (NNK), NNK acetate (NNK-Ae), methotrexate (MTX), and cisplatin to validate cytotoxicity, total reactive oxygen species, intracellular antioxidants, DNA fragmentation, and DNA tail damage. Furthermore, phosphorylated histone (-H2AX) and proteins involved in DNA damage (ATM/ATR, Chk1, Chk2, and p53) and repair (DNA-PKcs and Ku80) mechanisms were evaluated by immunofluorescence and western blotting, respectively. The results revealed that AF4-pretreated cells showed lower cytotoxicity, total ROS generation, and DNA fragmentation along with consequent inhibition of DNA tail moment. An increased level of -H2AX and DNA damage proteins was observed in carcinogen-treated cells and that was significantly ( ≤ 0.05) inhibited in AF4-pretreated cells, in an ATR-dependent manner. AF4 pretreatment also facilitated the phosphorylation of DNA-PKcs and thus initiation of repair mechanisms.
Apple flavonoids can protect in vitro oxidative DNA damage and facilitate repair mechanisms.
由于 DNA 损伤导致的人类肿瘤转化对全球医疗保健构成了日益严重的问题。维持基因组完整性对于避免肿瘤的发生和发展至关重要。本研究旨在探讨苹果类黄酮混合物（AF4）对正常人类支气管上皮细胞中各种致癌剂诱导的毒性的作用及其对 DNA 损伤反应和修复过程的作用机制。
用尼古丁衍生的亚硝胺酮（NNK）、NNK 醋酸盐（NNK-Ae）、甲氨蝶呤（MTX）和顺铂处理经 AF4 预处理的细胞，以验证细胞毒性、总活性氧、细胞内抗氧化剂、DNA 片段化和 DNA 尾部损伤。此外，通过免疫荧光和蛋白质印迹分别评估磷酸化组蛋白（-H2AX）和涉及 DNA 损伤（ATM/ATR、Chk1、Chk2 和 p53）和修复（DNA-PKcs 和 Ku80）机制的蛋白质。结果表明，经 AF4 预处理的细胞显示出较低的细胞毒性、总 ROS 生成和 DNA 片段化，以及 DNA 尾部瞬间的抑制。在致癌剂处理的细胞中观察到 -H2AX 和 DNA 损伤蛋白水平升高，并且在用 AF4 预处理的细胞中以 ATR 依赖性方式显著（≤0.05）抑制。AF4 预处理还促进了 DNA-PKcs 的磷酸化，从而启动了修复机制。
苹果类黄酮可以保护体外氧化 DNA 损伤并促进修复机制。
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