γH2AX responses in human buccal cells exposed to ionizing radiation

Abstract

DNA double strand breaks are induced by ionizing radiation (IR), leading to the phosphorylation of the core histone protein H2AX (termed γH2AX). The understanding of the γH2AX responses in irradiated human buccal cells is still very limited. We used visual scoring and laser scanning cytometry (LSC) methods to investigate γH2AX signaling following exposure of human buccal cells (from six individuals) to ionizing radiation at 0-4 Gy. The frequency of nuclei containing 15-30 γH2AX foci was significantly elevated 30 min post-IR exposure (by visual scoring). Concomitantly, there was a significant decrease in the frequency of cells without foci following exposure to IR. IR-induced γH2AX signal as determined by laser scanning cytometry (which included γH2AX integral and MaxPixel value) increased significantly in all individual's 2N nuclei 30 min post-IR and was similar for all three nuclear shapes identified. Individuals with the lowest baseline γH2AX integral (i.e., in nonirradiated cells) showed the greatest fold stimulation of γH2AX and significant dose-responses to IR doses of 1, 2, and 4 Gy. In 5 out of 6 individuals, the frequency of visually scored γH2AX in nuclei showed a strong correlation (up to r = 0.999) with LSC scored γH2AX integrals. The γH2AX response and subsequent decline varied between individuals but remained elevated above baseline levels 24 h post IR exposure. γH2AX response in irradiated human buccal cells has potential to be used as an index of baseline DNA damage in population studies. The variable response to IR exposure between individuals should be taken into consideration when using the γH2AX assay for radiation biodosimetry.