Prediction of clonogenic cell survival curves based on the number of residual DNA double strand breaks measured by γH2AX staining

Purpose:

To assess the potential of using the residual phosphorylation of histone H2AX (γH2AX) after irradiation as a marker of radiosensitivity invitro.

Material and methods:

Confluent cell cultures of FaDu and SKX human squamous cell carcinoma lines were irradiated with graded single doses. Twenty-four hours after irradiation cells were seeded for standard colony forming assay (CFA). In parallel, staining for γH2AX was performed to visualise the residual foci.

Results:

In the CFA, FaDu showed a higher radioresistance than SKX. After analysis of the residual foci data, we constructed ‘predicted’ survival curves using two different methods. First, the proportion of nuclei with <3 foci was found to correlate closely with the observed surviving fraction (SF) in FaDu, with a slight overestimation of the true SF in SKX. Second, there was a strong linear correlation of the mean number of residual foci and observed −lnSF. Based on regression analysis, we calculated the SF for both cell lines based on the mean number of residual γH2AX foci. This second approach again led to a good correlation of predicted and observed SF values in FaDu and a (slight) overestimation in SKX.

Conclusion:In the two cell lines investigated the mean number of residual foci of γH2AX can be used to predict differences in the radiation dose response relationship invitro.