Ivanov Andrei, Cragg Mark S, Erenpreisa Jekaterina, Emzinsh Dzintars, Lukman Henny, Illidge Timothy M
Cancer Research UK, Wessex Oncology Unit, Cancer Sciences Division, School of Medicine, Southampton University Hospital, Southampton SO16 6YD, UK.
J Cell Sci. 2003 Oct 15;116(Pt 20):4095-106. doi: 10.1242/jcs.00740. Epub 2003 Sep 2.
p53 mutant tumour cells respond to genotoxic insults by bypassing G1 arrest and halting in G2. Following release from G2 arrest they undergo mitotic catastrophe, whereby mitotic cycling is suppressed, delayed apoptosis begins and endopolyploid cells are produced. The ability of these endopolyploid cells to participate in the restitution process is controversial. To facilitate recovery, these endopolyploid cells must repair the extensive DNA damage induced. DNA damage and its resolution were studied by observing the kinetics of gamma-H2AX foci formation and by comet assay analysis. Subsequently, the kinetics and distribution of Rad51 foci were studied as a measure of homologous recombination. Here we present evidence of the resolution of DNA damage in endopolyploid cells through a decrease of tail moment by comet assay and in the number of cells expressing gamma-H2AX foci. Rad51 foci expression reached a maximum in endopolyploid cells on days 5-6 after irradiation, when delayed apoptosis was maximal, indicating that cells were being selected for survival at this time. Furthermore, the proportion of Annexin-V-positive polyploid cells decreased as they continued ongoing rounds of DNA replication, suggesting endoreduplication is involved in selecting cells resistant to apoptosis. Our findings suggest that after severe genotoxic insult endopolyploid cells have a transient survival advantage that may contribute to radioresistance of tumours that undergo mitotic catastrophe.
p53 突变肿瘤细胞通过绕过 G1 期阻滞并在 G2 期停滞来应对基因毒性损伤。从 G2 期阻滞释放后，它们会经历有丝分裂灾难，在此过程中，有丝分裂循环受到抑制，延迟凋亡开始，并且会产生内多倍体细胞。这些内多倍体细胞参与修复过程的能力存在争议。为了促进恢复，这些内多倍体细胞必须修复所诱导的广泛 DNA 损伤。通过观察 γ-H2AX 焦点形成的动力学以及彗星试验分析来研究 DNA 损伤及其修复。随后，研究 Rad51 焦点的动力学和分布，作为同源重组的一种衡量指标。在这里，我们通过彗星试验中尾矩的降低以及表达 γ-H2AX 焦点的细胞数量的减少，提供了内多倍体细胞中 DNA 损伤修复的证据。Rad51 焦点表达在照射后第 5 - 6 天在内多倍体细胞中达到最大值，此时延迟凋亡最为明显，表明此时细胞正在被选择存活。此外，随着 Annexin-V 阳性多倍体细胞持续进行多轮 DNA 复制，其比例下降，这表明核内复制参与了选择抗凋亡细胞。我们的研究结果表明，在严重的基因毒性损伤后，内多倍体细胞具有短暂的生存优势，这可能导致经历有丝分裂灾难的肿瘤产生放射抗性。
