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. 2022 Jan 28;12:1559. doi: 10.1038/s41598-022-05339-z

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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GNP labelled Detroit-562 and HSC-3 HNCCs demonstrate increased fluorescence intensity (FI) of double stranded breaks (DSBs) with DTBR compared to NoNP cells with STBR. Cells were cultured in 6-well plates, irradiated with 8 Gy from the STB or DTB, then fixed 30 min after radiation. Cells were processed for immunohistochemistry (IHC) with γ-H2AX to assess DNA double strand breaks and DAPI nuclear stain. (a) The FI of γ-H2AX foci was determined and measured relative to the FI of DAPI nuclear stain within each nucleus. GNP-labeled Detroit-562 and HSC-3 cells that received DTBR demonstrated increased FI of γ-H2AX foci than No NP cells with STBR (p = 0.049 and p < 0.0001, respectively). With DTBR, Detroit-562, FaDu, and HSC-3 GNP-labeled cells demonstrated greater FI of γ-H2AX foci compared to No NPs (p < 0.0001 for all). GNP-labeled FaDu and HSC-3 cells exhibited increased FI with DTBR compared to STBR (p = 0.001 and p < 0.0001, respectively), but GNP-labeled Detroit-562 cells displayed decreased FI with DTBR compared to STBR (p < 0.0001). (b) The number of γ-H2AX foci was analysed and made relative to the nuclear area. With DTBR, GNP-labeled Detroit-562, FaDu, and HSC-3 cells exhibited significantly greater foci/nucleus than No NP cells (p < 0.0001, p < 0.0001, and p < 0.001, respectively). GNP-labeled Detroit-562 and HSC-3 cells that received DTBR display greater foci/nucleus than No NP cells with STBR (p = 0.0397 and p < 0.0001, respectively), but GNP-labeled FaDu cells with DTBR demonstrated a decrease in foci/nucleus compared to No NP cells with STBR (p < 0.0001). Lastly, GNP-labeled Detroit-562 and FaDu cells with DTBR displayed significantly less foci/nucleus than with STBR (p < 0.0001 for both), but HSC-3 cells exhibited more (p < 0.0001). (c) Representative confocal images of Detroit-562 cells post RT (Zeiss LSM 710 Laser Scanning Confocal microscope at 40 ×). Scale bars = 20 μm. Results are presented as means of [a] Relative fluorescence intensity and [b] Relative Foci count ± standard error of the mean, p* < 0.05, p** < 0.01, p*** < 0.001, p**** < 0.0001. Significance between groups was tested using a standard t-test with Tukey multiple comparison test (n = 3; 4 images/sample). GNP gold nanoparticles, NoNP no nanoparticles, HNCC head and neck cancer cells, STB(R) standard target beam (radiation), DTB(R) diamond target beam (radiation).