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ATR-dependent radiation-induced γH2AX foci in bystander primary human astrocytes and glioma cells

Oncogene volume 26, pages 993–1002 (2007)Cite this article

Abstract

Radiotherapy is an important treatment for patients suffering from high-grade malignant gliomas. Non-targeted (bystander) effects may influence these cells' response to radiation and the investigation of these effects may therefore provide new insights into mechanisms of radiosensitivity and responses to radiotherapy as well as define new targets for therapeutic approaches. Normal primary human astrocytes (NHA) and T98G glioma cells were irradiated with helium ions using the Gray Cancer Institute microbeam facility targeting individual cells. Irradiated NHA and T98G glioma cells generated signals that induced γH2AX foci in neighbouring non-targeted bystander cells up to 48 h after irradiation. γH2AX bystander foci were also observed in co-cultures targeting either NHA or T98G cells and in medium transfer experiments. Dimethyl sulphoxide, Filipin and anti-transforming growth factor (TGF)-beta 1 could suppress γH2AX foci in bystander cells, confirming that reactive oxygen species (ROS) and membrane-mediated signals are involved in the bystander signalling pathways. Also, TGF-beta 1 induced γH2AX in an ROS-dependent manner similar to bystander foci. ROS and membrane signalling-dependent differences in bystander foci induction between T98G glioma cells and normal human astrocytes have been observed. Inhibition of ataxia telangiectasia mutated (ATM) protein and DNA-PK could not suppress the induction of bystander γH2AX foci whereas the mutation of ATM- and rad3-related (ATR) abrogated bystander foci induction. Furthermore, ATR-dependent bystander foci induction was restricted to S-phase cells. These observations may provide additional therapeutic targets for the exploitation of the bystander effect.

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Acknowledgements

We are grateful to Stuart Gilchrist and Bob Sunderland for providing excellent technical support for the microbeam irradiation. Graeme Smith, KuDos Pharmaceuticals, Cambridge, UK, kindly provided the ATM inhibitor KU-55933 and the DNA-PK inhibitor NU 7026. Penny Jeggo, University of Sussex, UK, kindly provided ATR Seckel fibroblasts (F02/98 htert) and suitable control cells (48BR htert). This study was funded by a grant from the James S McDonnell Foundation to SC Short, M Folkard and KM Prise with additional support from the Gray Cancer Institute.

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  1. Gray Cancer Institute, Mount Vernon Hospital, Northwood, UK

    S Burdak-Rothkamm, S C Short, M Folkard & K M Prise

  2. Gray Cancer Institute & Radiation Oncology & Biology, University of Oxford, Oxford, UK

    K Rothkamm

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Correspondence to S Burdak-Rothkamm.

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Burdak-Rothkamm, S., Short, S., Folkard, M. et al. ATR-dependent radiation-induced γH2AX foci in bystander primary human astrocytes and glioma cells. Oncogene 26, 993–1002 (2007). https://doi.org/10.1038/sj.onc.1209863

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