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. 2020 May 11;11:2331. doi: 10.1038/s41467-020-16241-5

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© The Author(s) 2020

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 — a Tracking of EVs by dynamic light-scattering microscopy (DLSM). EVs were isolated from 5 × 10⁵ uninfected or opsonized C. albicans-infected monocytes (MEVs or MEVs_Ca, respectively) by polymer precipitation. Representative data of three independent experiments (three donors) are shown. b MEVs_Ca are significantly increased compared with MEVs (data are presented as mean values + /− SD, p = 0.0027, unpaired two-tailed t test, n = 3 different donors). EVs isolated from same number of infected or uninfected monocytes were counted by DLSM. c Size distribution of MEVs and MEVs_Ca as determined by DLSM using NanoSight NTA 3.2 software. Graphs were generated by overlaying the size distribution of MEVs and MEVs_Ca from n = 3 donors. d The double-membrane structure and round shape of the MEVs_Ca are visible by cryogenic electron microscopy (Cryo-EM). Bars: 100 nm. e MEVs_Ca structure and shape is confirmed by freeze-fracture electron microscopy (FFEM). Bars: 100 nm. f MEVs_Ca show significantly higher protein content compared with MEVs (p < 0.05, unpaired two-tailed t test, n = 3 different donors). Proteins from MEVs and MEVs_Ca (each from 1 × 10⁷ monocytes) were detected by label-free LC-MS/MS-based proteomics. g Most of the proteins enriched in MEVs_Ca compared with MEVs are extracellular exosome-related proteins (Gene Ontology (GO) enrichment analysis). h Complement pathway proteins were significantly increased in MEVs_Ca (Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis). i Heatmap of complement and coagulation proteins upregulated in MEV_Cas. Comparisons in g–i rely on protein content of MEVs_Ca from three different donors. j Diagram of an MEVs_Ca showing the presence of receptors, marker proteins, and cytokines detected in the current study. k TGF-β1 (data are presented as mean values + /− SD, p = 0.0013, unpaired two-tailed t test, n = 3 donors), l but not IL-6, is significantly increased in MEVs_Ca compared with MEVs (data are presented as mean values + /− SD, p = 0.2432, unpaired two-tailed t test, n = 3 different donors). Cytokines from MEVs and MEVs_Ca (each isolated from monocytes (5 × 10⁵)) were determined by ELISA.