Cell Cycle Disturbances and Mitotic Catastrophes in HeLa Hep2 Cells following 2.5 to 10 Gy of Ionizing Radiation

Abstract

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 G2-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 α-tubulin and γ-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 HeLa Hep2 cells into mitotic catastrophe and delayed apoptosis. These might be important cell death mechanisms involved in tumor growth retardation following radioimmunotherapy of solid tumors.

A novel Chk inhibitor, XL-844, increases human cancer cell radiosensitivity through promotion of mitotic catastrophe

“…Such expression of tubulin subunits has been described in cells undergoing mitotic catastrophe [35][36][37][38]. The induction of mitotic catastrophe with abnormal mitotic cells [39][40][41] may result in apoptosis or other forms of cell death [34,42,43].…”

Twilight effects of low doses of ionizing radiation on cellular systems: a bird’s eye view on current concepts and research

“…apoptosis and MC) are functionally linked since mitotic catastrophe may be considered as a subtype of caspasemediated death resulting from a persistent DNA damage and deficient cell cycle checkpoint control [148]. No wonder that a role of p53 in MC has been hypothesized by some authors [149] also in this instance. It has been demonstrated, in fact, that p53 regulates cellular levels of cyclin B1, through a potential transcriptional repression mechanism [35,150].…”

Radiation-induced cell death mechanisms

“…The mitotic catastrophe is generally considered a consequence of premature or improper entry of a cell into mitosis and can be induced by a multitude of DNA-damaging agents including radiation. The mitotic catastrophe is executed by most non-hematopoietic tumor cells in response to ionizing radiation [68] and is frequently observed in experimental tumors following radiotherapy and radioimmunotherapy [69][70][71][72][73]. Mitotic catastrophe is today considered to be the major cell death mechanism by which solid tumors respond to clinical radiotherapy.…”