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. 2019 Mar 20;9:4913. doi: 10.1038/s41598-019-41211-3

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© The Author(s) 2019

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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ASC peptides effectively block IL-1β and IL-18 release and ASC oligomerization in human blood monocytes. (A) IL-1β and IL-18 were measured in cell-free supernatants from THP-1 cells incubated with FAM-ASC^PYD #5 or FAM-ASC^PYD #6 (12.5, 25,50 and 100 μM) for 24 h and primed with LPS (0.1 μg/ml, 4 h) followed by stimulation with nigericin (10 μM, 1 h), as assessed by ELISA. (B) Production of IL-1β and IL-18 from human blood monocytes stimulated or not with LPS (0.1 μg/ml, 4 h) alone or in a combination of ATP (5 mM, 30 min) in the presence of FAM-ASC^PYD #5 or FAM-ASC^PYD #6 (25, 50 and 100 μM) peptides. (C,D) Western blot analysis of processed IL-1β p17 (C) and Casp-1 p20 (D) in cell-free supernatants of THP-1 cells primed with LPS (0.1 μg/ml, 4 h), followed by stimulation with nigericin (10 μM, 1 h) in the presence of FAM-ASC^PYD #5 (25, 50 and 100 μM), FAM-ASC^PYD #6 (50 and 100 μM) peptides, or DMSO vehicle. Expression of pro-IL-1β p35 and pro-Casp-1 p50 was assessed in cell lysates. GAPDH was used as loading control. (E) Imaging of ASC speck oligomerization in THP-1 cells treated with LPS (0.1 μg/ml, 4 h) and nigericin (10 μM, 1 h) in the absence or presence of FAM-ASC^PYD #5 or FAM-ASC^PYD #6 (100 μM) peptides. The number of ASC specks were counted manually in at least 100 cells, and the proportion of cells containing a speck was shown. Objective 100× , Magnification 1.0× . Data represent the mean ± standard error of triplicate wells and are representative of three independent experiments. *p < 0.05; **p < 0.01; ***p < 0.001.