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. 2021 May 5;11:9529. doi: 10.1038/s41598-021-88897-y

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© The Author(s) 2021

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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Streptococcus pneumoniae (Spn) induces epithelial microvesicle (MV) production via pneumolysin. (A) A549 cells were treated with Spn WT, Spn Δply (MOI 50), or PBS (Ctr) for 4 h and MVs were isolated from conditioned media. Annexin V + MVs were quantified by FACS. N = 4 *p < 0.01 vs Ctr, #p < 0.01 vs Spn Δply (One-way Anova). (B,C) Human lung specimens (pieces from the same lung) were treated with Spn WT, Spn Δply (1 × 10⁶ CFU/ml), or PBS. (B) Graph depicts the relative amount of annexin V/EPCAM double positive MVs (normalized to Ctr). N = 6, *p < 0.05 vs Ctr, #p < 0.05 vs Spn ΔPly (One-way Anova). (C) MV lysates from human lung tissue supernatants probed for pan-cytokeratin (epithelial marker). Depicted is a representative blot (cropped image). (D) MVs of epithelial origin (annexin V/EPCAM double positive) were quantified in BAL of mice treated intranasally with Spn or PBS (Ctr). Each mouse is represented by a dot. N = 4–6, *p < 0.05 (unpaired t-test).