Abstract
BackgroundTumour radiosensitivity would be particularly useful in optimizing the radiation dose during radiotherapy. The aim of the current study was to evaluate the potential value of phosphorylated H2AX (γH2AX) and ATM (pATM) in assessing 12C6+ radiosensitivity of tumour cells.MethodsHuman cervical carcinoma HeLa cells, hepatoma HepG2 cells, and mucoepidermoid carcinoma MEC-1 cells were irradiated with different doses of 12C6+. The survival fraction was assayed with clonogenic survival method and the foci of γH2AX and pATM was visualized using immunocytochemical methods. Flow cytometry was used to assay γH2AX, pATM and the cell cycle.ResultsThe survival fraction decreased immediately in dose-dependent manner, but in turn, significantly increased during 24 h after 12C6+ irradiation. Both γH2AX and pATM foci accumulated linearly with doses and with a maximum induction at 0.5 h for γH2AX and 0.5 or 4 h for pATM, respectively, and a fraction foci kept for 24 h. The expression of γH2AX and pATM was in relation to cell cycle. The G0/G1 phase cells had the highest expression of γH2AX after 0.5 h irradiation and then decreased to a lower level at 24 h after irradiation. An obvious increase of pATM in G2/M phase was shown after 24 h of 2 and 4 Gy irradiation. The significant G2/M phase arrest was shown. There was a close relationship between the clonogenic survival and γH2AX and pATM expression both in timing and dose in response to 12C6+.ConclusionsThe rate of γH2AX and pATM formation and loss may be an important factor in the response of cells to 12C6+. pATM and γH2AX are effective radiation biomarkers in assessing the radiosensitivity of 12C6+ in human tumor cells.
Highlights
Tumour radiosensitivity would be useful in optimizing the radiation dose during radiotherapy
Our studies emphasize the rate of H2AX phosphorylation (γH2AX) and phosphorylated H2AX (γH2AX) and ATM (pATM) formation and loss may be an important factor in the response of cells to 12C6+. pATM and γH2AX are effective radiation biomarkers in assessing the radiosensitivity of 12C6+ in human tumor cells
Immunofluorescence staining of phosphorylated H2AX and Ataxia Telangiectasia Mutation (ATM) foci Phosphorylated H2AX and ATM foci were observed with anti-γH2AX antibodies, anti- ATMpSer1981 antibodies and the nuclei were stained with DAPI
Summary
Tumour radiosensitivity would be useful in optimizing the radiation dose during radiotherapy. Activation of the Ataxia Telangiectasia Mutation (ATM) through its phosphorylation on Ser1981 (ATM–S1981P, pATM), and phosphorylation of one of the variants of histone H2AX, histone H2AX on Ser139 (γH2AX), are the main participants, and the early markers of a cell’s response to DNA damage, if the damage involves formation of DSB [5, 6]. These modifications of ATM and H2AX trigger pathways are involved in DNA repair and in activating checkpoints that halt progression through the cell cycle [7, 8]. The pause in cell cycle progression is needed to allow for DNA repair to succeed prior to resumption of DNA replication or cell division
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