Apel Anja, Herr Ingrid, Schwarz Heinz, Rodemann H Peter, Mayer Andreas
Division of Radiobiology and Molecular Environmental Research, University of Tuebingen, Germany.
Cancer Res. 2008 Mar 1;68(5):1485-94. doi: 10.1158/0008-5472.CAN-07-0562.
Autophagy or "self eating" is frequently activated in tumor cells treated with chemotherapy or irradiation. Whether autophagy represents a survival mechanism or rather contributes to cell death remains controversial. To address this issue, the role of autophagy in radiosensitive and radioresistant human cancer cell lines in response to gamma-irradiation was examined. We found irradiation-induced accumulation of autophagosomes accompanied by strong mRNA induction of the autophagy-related genes beclin 1, atg3, atg4b, atg4c, atg5, and atg12 in each cell line. Transduction of specific target-siRNAs led to down-regulation of these genes for up to 8 days as shown by reverse transcription-PCR and Western blot analysis. Blockade of each autophagy-related gene was associated with strongly diminished accumulation of autophagosomes after irradiation. As shown by clonogenic survival, the majority of inhibited autophagy-related genes, each alone or combined, resulted in sensitization of resistant carcinoma cells to radiation, whereas untreated resistant cells but not sensitive cells survived better when autophagy was inhibited. Similarly, radiosensitization or the opposite was observed in different sensitive carcinoma cells and upon inhibition of different autophagy genes. Mutant p53 had no effect on accumulation of autophagosomes but slightly increased clonogenic survival, as expected, because mutated p53 protects cells by conferring resistance to apoptosis. In our system, short-time inhibition of autophagy along with radiotherapy lead to enhanced cytotoxicity of radiotherapy in resistant cancer cells.
自噬，即“自我吞噬”，在接受化疗或放疗的肿瘤细胞中常常被激活。自噬是一种生存机制还是会导致细胞死亡，这一问题仍存在争议。为了解决这个问题，我们研究了自噬在人源放射敏感和放射抗性癌细胞系对γ射线照射的反应中的作用。我们发现，照射会诱导自噬体的积累，同时每个细胞系中自噬相关基因beclin 1、atg3、atg4b、atg4c、atg5和atg12的mRNA也会强烈诱导表达。通过逆转录PCR和蛋白质免疫印迹分析表明，转导特异性靶标小干扰RNA可导致这些基因下调长达8天。照射后，阻断每个自噬相关基因均与自噬体积累的显著减少相关。克隆形成存活实验表明，大多数被抑制的自噬相关基因，无论是单独作用还是联合作用，都会使抗性癌细胞对辐射敏感，而当自噬被抑制时，未处理的抗性细胞比敏感细胞存活得更好。同样，在不同的敏感癌细胞系以及抑制不同的自噬基因时，也观察到了放射增敏或相反的情况。正如预期的那样，突变型p53对自噬体的积累没有影响，但略微增加了克隆形成存活率，因为突变型p53通过赋予细胞抗凋亡能力来保护细胞。在我们的系统中，短时间抑制自噬并结合放疗会增强放疗对抗性癌细胞的细胞毒性。
