DNA-PKcs基因沉默抑制人乳腺上皮细胞对低剂量辐射损伤的修复

DNA-PKcs silencing inhibit the DNA repair induced by low dose radiation on human breast epithelial cells

摘要:

DNA-PKcs作为DNA依赖性蛋白激酶(DNA-PK)的催化亚基在DNA双链断裂(DSBs)的非同源末端重组(NHEJ)通路中起重要的作用。本实验以人乳腺上皮细胞株MCF10F为研究对象, 通过siRNA技术抑制细胞内DNA-PKcs的表达,用50 cGy 137CS照射细胞, 测定细胞生长曲线以确定细胞对低剂量辐射(LDR)的敏感性, 同时检测DNA 修复相关蛋白表达的变化, 旨在探讨DNA依赖蛋白激酶(DNA-PKcs)基因沉默对人乳腺上皮细胞株MCF10F低剂量辐射敏感性的影响及机制。结果显示: 转染特异性siRNA可使人乳腺上皮细胞(MCF10F) DNA-PKcs基因沉默, 增殖受到明显的抑制; 50 cGy g射线辐射可使乳腺细胞内DNA-PKcs、Ku80、ATM、P53等DNA修复相关蛋白表达增多, 但DNA-PKcs基因沉默细胞(MCF10Fpk)中, 这些蛋白表达显著低于对照组(MCF10Fmock)。以上结果提示, DNA-PKcs基因沉默可引起乳腺细胞对低剂量辐射敏感性增加, 其原因可能与相关DNA修复蛋白表达减少有关。

Abstract:

DNA-PKcs, the catalytic subunit of DNA-dependent protein kinase (DNA-PK), plays an important role in the nonhomologous end-joining (NHEJ) pathway of DNA double-strand breaks (DSBs) repair. To investigate the effects of DNA-PKcs down-regulation on cell growth and sensitization to low dose radiation (LDR), we transfected DNA-PKcs siRNA into human mammary epithelia cells MCF10F, then, detected the proliferation curve of the cells and the expression of protiens in DNA repair pathways. The results showed that DNA-PKcs gene silencing, induced by the transfection of DNA-PKcs siRNA could suppress significantly the cell proliferation and inhibit the expression of the DNA repair proteins, such as Ku80, ATM and P53 after 50 cGy 137Cs g-irradiation.The results suggested that DNA-PKcs gene silencing could increase the sensitivity of human breast epithelial cells to the LDR, which might be relative with the decrease of the proteins.