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28 September 2010 Analysis of Flow Cytometry DNA Damage Response Protein Activation Kinetics after Exposure to X Rays and High-Energy Iron Nuclei
Lori J. Chappell, Mary K. Whalen, Sheena Gurai, Artem Ponomarev, Francis A. Cucinotta, Janice M. Pluth
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Abstract

We developed a mathematical method to analyze flow cytometry data to describe the kinetics of γ-H2AX and pATF2 phosphorylation in normal human fibroblast cells after exposure to various qualities of low-dose radiation. Previously reported flow cytometry kinetics for these DSB repair phospho-proteins revealed that distributions of intensity were highly skewed, severely limiting the detection of differences in the very low-dose range. Distributional analysis revealed significant differences between control and low-dose samples when distributions were compared using the Kolmogorov-Smirnov test. Differences in radiation quality were found in the distribution shapes and when a nonlinear model was used to relate dose and time to the decay of the mean ratio of phospho-protein intensities of irradiated samples to controls. We analyzed cell cycle phase- and radiation quality-dependent characteristic repair times and residual phospho-protein levels with these methods. Characteristic repair times for γ-H2AX were higher after exposure to iron nuclei compared to X rays in G1 cells and in S/G2 cells. The RBE in G1 cells for iron nuclei relative to X rays for γ-H2AX was 2.1 ± 0.6 and 5.0 ± 3.5 at 2 and 24 h after irradiation, respectively. For pATF2, a saturation effect was observed with reduced expression at high doses, especially for iron nuclei, with much slower characteristic repair times (>7 h) compared to X rays. RBEs for pATF2 were 0.7 ± 0.1 and 1.7 ± 0.5 at 2 and 24 h, respectively. Significant differences in γ-H2AX and pATF2 levels when irradiated samples were compared to controls were noted even at the lowest dose analyzed (0.05 Gy). These results show that mathematical models can be applied to flow cytometry data to identify important and subtle differences after exposure to various qualities of low-dose radiation.

Lori J. Chappell, Mary K. Whalen, Sheena Gurai, Artem Ponomarev, Francis A. Cucinotta, and Janice M. Pluth "Analysis of Flow Cytometry DNA Damage Response Protein Activation Kinetics after Exposure to X Rays and High-Energy Iron Nuclei," Radiation Research 174(6a), 691-702, (28 September 2010). https://doi.org/10.1667/RR2204.1
Received: 10 March 2010; Accepted: 1 June 2010; Published: 28 September 2010
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