Effect of gold nanoparticle on radiation induced DNA damage in MCF7 breast cancer cells.
Janic B, Liu F, Bobbitt K, Brown S, Mao G, Chetty IJ, Movsas B, and Wen NW. Effect of gold nanoparticle on radiation induced DNA damage in MCF7 breast cancer cells. Cancer Res 2018; 78(13)1376.
Nanoparticles (NPs) are 1-100 nm constructs explored for their application in cancer diagnosis and treatment. Exposure of cells to ionizing irradiation leads to DNA damage, with DNA double strand breaks (DSBs) being the most toxic that can lead to apoptosis. The presence of gold in NPs enhances radiation damage and DSBs, the latter measured by the levels of phosphorylated DNA histone protein H2AX (γH2AX). The goal of this study was to decipher biological mechanisms of NP radio-sensitization. We analyzed the effects of two different sizes of gold nanoparticles (AuNP) on DSBs in MCF-7 breast cancer cells by assessing H2AX phosphorylation at three photon energies. Cells were incubated with either 4 nm or 14 nm AuNP and irradiated with 2, 4 or 8 Gy using 2.5 FFF MV (60 MU/min), 6 MV (600 MU/min) or 10 FFF MV (2400 MU/min) X-rays. Percent of cells positive for γH2AX was determined by flow cytometry. Live cells (100000) were gated using forward (FSC; cell size) versus side scatter (SSC; complexity) characteristics (gate P1). Cells positive for γH2AX were further gated to identify subpopulations exhibiting brighter (gate P2) or dimmer (gate P3) fluorescence intensity. Treatment with 4 nm AuNP resulted in significantly more γH2AX positive cells after irradiation at all three energies, compared to their respective controls. When treated at 6 MV energy, increases in the percentage of cells positive for H2AX phosphorylation was detected at all three doses (2, 4 and 8 Gy). Increased phosphorylation was accompanied by an increase of cells in the P2 gate accompanied by an increase in their FCS and SSC characteristics, consistent with activation. In cells treated with 10 MV the effect was most pronounced at 4Gy dose, while 2 and 8 Gy resulted in a slight increase in the percentage of vH2AX positive cells. Nevertheless, these cells still exhibited prominent increases within the P2 gate and FCS vs SSC characteristics, compared to their irradiated controls. In cells treated with 2.5 MV energy similar results were observed using 4 and 8 Gy, while no effect was detected with 2 Gy. Data from the experiments using 14 nm size AuNP were less clear and require further investigation. The difference may be due to the localization of the smaller NPs within the nucleus and therefore cause more DNA damage thereby greater H2AX phosphorylation. We hypothesize that the NP radio-sensitization mechanism involves directing cells towards apoptosis by enhancing DNA damage and interfering with DNA repair. Hence, vH2AX positive cells exhibiting bright fluorescence (P2) and an increase in FCS and SSC identified even in the conditions without significant increase in the percent of total vH2AX positive cells, may reflect cells primed for apoptosis. Future studies are planned to elucidate the exact intracellular mechanisms of NP biological radio-sensitization effect.