Dose threshold for residual γH2AX, 53BP1, pATM and p-p53 (Ser-15) foci in X-ray irradiated human fibroblasts.

Abstract

Background: Enumeration of residual DNA repair foci 24 hours or more after exposure to ionizing radiation (IR) is often used to assess the efficiency of DNA double-strand break repair. However, the relationship between the number of residual foci in irradiated cells and the radiation dose is still poorly understood. The aim of this work was to investigate the dose responses for residual DNA repair foci in normal human fibroblasts after X-ray exposure in the absorbed dose range from 0.1 to 5 Gy.

Materials and methods: Fibroblasts were irradiated using a X-ray unit at an absorbed dose rate of 0.2 Gy/min. Irradiated cells were incubated for 0.5, 24, 48 and 72 h. Immunofluorescence visualized γH2AX, 53BP1, pATM and p-p53 (Ser-15) foci were enumerated using DARFI software and by manual scoring. Additionally, clonogenic survival analysis was performed.

Results: The data analysis performed with the hockey stick model showed the presence of a dose threshold for the residual foci of all proteins studied. The estimated threshold doses are close to the quasi-threshold dose (Dq = 0.99 ± 0.09 Gy) calculated from the cell survival curve.

Conclusion: The excellent agreement between the calculated values of the threshold dose and Dq in irradiated fibroblasts proves that residual foci are sites, where cells are still attempting to repair potentially lethal DNA damage.