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. 2021 Jun 17;18:21. doi: 10.1186/s12989-021-00415-0

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© The Author(s) 2021

Open AccessThis 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/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 3 — Silica-NPs accumulated in alveolar macrophages of silica-NPs instilled murine lungs. A-B. Distribution of silica particles in murine lungs was studied by confocal fluorescence microscopy. A. Representative images showing the localization of silica-NPs and macrophages labeled with anti-CD68 antibody. In the images, blue indicates cell nucleus, green indicates FITC-labeled silica particles, and red indicates Alexa 594 labeled CD68. Scale bars in low magnification (top panels) = 50 μm, in high magnification (bottom panels) = 10 μm. B. Representative images showing the localization of silica-NPs and neutrophils labeled with anti-Ly-6G antibody. In the images, blue indicates cell nucleus, green indicates FITC- labeled silica particles, and red indicates Alexa 594 labeled Ly6G. The white arrows indicate Ly-6G-postive cells with multi-segmented nucleus, which are absent from FITC-labeled silica-NPs. Scale bars in low magnification (top panels) = 50 μm, in high magnification (bottom panels) = 10 μm. C-D. Distribution of silica particles in cells in BALF were studied by confocal fluorescence microscopy. C. Representative images showing the localization of silica-NPs and macrophages labeled with anti-CD68 antibody. In the images, blue indicates cell nucleus, green indicates FITC-labeled silica particles, and red indicates Alexa 594-labeled CD68. Note that the CD68-negative, smaller cells with multi-segmented nucleus indicated by white arrows, are absent from FITC-labelled silica-NPs. D. Plots of the fluorescence intensity of CD68 (red), FITC-labelled silica-NPs (green), and nucleus (blue) over a cross-section along the indicated yellow arrows. Data are representative of at least three independent experiments. E-F. Flow cytometric detection of intracellular silica-NP accumulation in cells retrieved from bronchoalveolar lavage. BAL cells from three animals were pooled in each experiment. E. Representative histograms. F. The ratio of FITC-positive cells in F4/80-positive macrophages. Data are shown as mean ± SD of three independent experiments. **p < 0.01, and ***p < 0.001 by ANOVA with Tukey’s multiple comparison tests