Cell cycle disturbances and mitotic catastrophes in HeLa hep2 cells following 2.5 to 10 Gy of ionizing radiation
Cell cycle disturbances and mitotic catastrophes in HeLa hep2 cells following 2.5 to 10 Gy of ionizing radiation
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DOI：
10.1158/1078-0432.ccr-07-0980
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发表时间：
2007-09-15
影响因子：
11.5
通讯作者：
Stigbrand, Torgny
中科院分区：
文献类型：
--
作者：
Eriksson, David;Loefroth, Per-Olov;Stigbrand, Torgny
目的：实验性放射免疫治疗提供的吸收剂量为2.5至10戈伊已被证明会导致肿瘤生长迟缓。本研究的目的是阐明暴露于这些剂量的HeLa Hep 2细胞中发生的连续分子和细胞事件。在2.5- 10-戈伊照射后，对HeLa Hep 2细胞的剂量-反应曲线、细胞周期检查点的激活和有丝分裂行为进行了研究。（4，5-二甲基噻唑-2-基）-2，5-二苯基四唑溴化物测定、蛋白质印迹、荧光激活细胞分选分析和免疫荧光染色。结果：荧光激活的细胞分选分析显示G2-M阻滞。p53和p21被发现上调，但并不直接相关的逮捕。G2-M期阻滞是短暂的，细胞重新进入细胞周期，仍然含有未修复的细胞损伤。这种过早进入引起后期桥，落后的染色体材料，多极有丝分裂纺锤体的增加，如碘化丙啶染色和α-微管蛋白和γ-微管蛋白抗体的免疫荧光染色所示。此外，在最高吸收剂量为10戈伊时，中心体数量从12.6 +/- 6.6%显著增加至67 +/- 5.3%，多倍体细胞从2.8 +/- 1.3%显著增加至17.6 +/- 2.1%。结论：2.5 ~ 10戈伊的吸收剂量可使He La Hep 2细胞发生有丝分裂灾难，并延迟凋亡。这些可能是实体瘤放射免疫治疗后肿瘤生长迟缓的重要细胞死亡机制。
Purpose: Experimental radioimmunotherapy delivering absorbed doses of 2.5 to 10 Gy has been shown to cause growth retardation of tumors. The purpose of this study was to elucidate the sequential molecular and cellular events occurring in HeLa Hep2 cells exposed to such doses.Methods: Dose-response curves, activation of cell cycle checkpoints, and mitotic behavior were investigated in HeLa Hep2 cells following 2.5- to 10-Gy irradiation by carrying out 3-(4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide assays, Western blots, fluorescence-activated cell sorting analysis, and immunofluorescence stainings. Terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling staining was used to detect apoptosis.Results: A G(2)-M arrest was shown by fluorescence-activated cell sorting analysis. p53 and p21 were found to be up-regulated but were not immediately related to the arrest. The G2-M arrest was transient and the cells reentered the cell cycle still containing unrepaired cellular damage. This premature entry caused an increase of anaphase bridges, lagging chromosomal material, and multipolar mitotic spindles as visualized by propidium iodide staining and immunofluorescence staining with alpha-tubulin and gamma-tubulin antibodies. Furthermore, a dose-dependent significant increase in centrosome numbers from 12.6 +/- 6.6% to 67 +/- 5.3% was identified as well as a dose-dependent increase of polyploid cells from 2.8 +/- 1.3% to 17.6 +/- 2.1% with the highest absorbed dose of 10 Gy. These disturbances caused the cells to progress into mitotic catastrophe and a fraction of these dying cells showed apoptotic features as displayed by terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling staining 5 to 7 days after irradiation.Conclusion: An absorbed dose of 2.5 to 10 Gy was shown to force He La Hep2 cells into mitotic catastrophe and delayed apoptosis. These might be important cell death mechanisms involved in tumor growth retardation following radio immunotherapy of solid tumors.