Computational modelling of γ-H2AX foci formation in human cells induced by alpha particle exposure

Abstract

AbstractIn cellular experiments, radiation-induced DNA damage can be quantified by counting the number of γ-H2AX foci in cell nucleus by using an immunofluorescence microscope. Quantification of DNA damage carries uncertainty, not only due to lack of full understanding the biological processes but also limitations in measurement techniques. The causes of limited certainty include the possibility of expressing foci in varying sizes responding individual DSBs and the overlapping of foci on the two-dimensional (2D) immunofluorescence microscopy image of γ-H2AX foci, especially when produced due to high-LET radiation exposure. There have been discussions on those limitations, but no successful studies to overcome them. In this paper, a practical modelling has been developed to simulate the occurrences of double-strand breaks (DSBs) and the formations of γ-H2AX foci in response to individual DSB formations, in cell nucleus due to exposure to alpha particles. Cell irradiation and DSB production were simulated using a user-written code that utilizes Geant4-DNA physics models. A C +  + code was used to simulate the formation γ-H2AX foci, which were spatially correlated to the loci of DBSs, and to calculate the number of individual foci from the observed 2D image of the cell nucleus containing the overlapping γ-H2AX foci. The average size of focal images was larger from alpha particle exposure than that from X-ray exposure, whereas the number of separate focal images were comparable except at doses up to 0.5 Gy. About 40% of separate focal images consisted of overlapping γ-H2AX foci at 1 Gy of alpha particle exposure. The foci overlapping ratios were obtained by simulation for individual size groups of focal images at varying doses. The size distributions of foci at varying doses were determined with experimentally obtained separate focal images. The correction factor for foci number was calculated using the foci overlapping ratio and foci size distribution, which are specific to dose from alpha particle exposure. The number of individual foci formations induced by applying the correction factor to the experimentally observed number of focal images better reflected the quality of alpha particles in causing DNA damage. Consequently, the conventional γ-H2AX assay can be better implemented by employing this computational modelling of γ-H2AX foci formation.