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. 2017 Nov 6;8:1319. doi: 10.1038/s41467-017-01433-3

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© The Author(s) 2017

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — Exo^NM increase PMo population. C57BL/6 mice were treated with Exo^M or Exo^NM, prior to BM isolation. a Flow cytometry shows the uptake of fluorophore-tagged exosomes by the myeloid cells in the BM. Left: Flow plots of the total CD11b⁺Gr1⁺ BM populations. Right: Plots show a higher percentage of CD11b⁺Gr1⁺ cells among the BM cells positive for exosome uptake. A representative of three independent experiments is shown. b Treatment with Exo^NM leads to an increase in Ly6C^lowCD11b⁺ cells compared to Exo^M in the total myeloid population. c qRT-PCR of mRNA encoding Nr4a1. Mouse BM monocytes were treated 24 h with exosomes (10 μg ml⁻¹). The experiment was performed in biological triplicates and technical replicates of two. *P < 0.05 by t-test in pairwise comparisons. Averages and s.d. values shown. d Western blot shows an increase in Nr4a1 in mouse BM monocytes treated with Exo^NM. Quantification was performed using ImageJ (National Institutes of Health), utilizing plot profile (areas under the peaks) and integrated density measures. A representative of three experiments is shown. e–h Exosomes increase the presence of PMo in the lung. C57BL/6 mice were treated with 10 μg Exo^NM or Exo^M 72 and 24 h prior to sacrifice. CX3CR1⁺ cells present in the lung were detected by FACS or in situ IF. e Mouse lungs were lavaged, smashed through a strainer and single-cell suspension stained for CD11b, CD45, and CX3CR1. FACS plots were gated for CD11b⁺CD45⁺ cells and CX3CR1⁺ cells quantified. f Quantification of the data shown in a (percent CX3CR1⁺of total CD45⁺ cells). *P < 0.05 calculated as above. Mean and s.d. values are shown. g IF analysis shows the increased abundance of Nr4a1⁺ cells in the lungs of mice treated with Exo^NM. Scale bar, 100 μm. h Quantification of the data in g, expressed as number of Nr4a1⁺ cells field⁻¹. Scale bar: 50 μm. n = 3 mice per group, five images per section. *P < 0.05, **P < 0.001, and ***P < 0.0005 by pairwise two-tailed t-test; n = 5 mice per group, >5 random images per lung. Mean and s.d. values are shown