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. 2017 Sep 29;7:12409. doi: 10.1038/s41598-017-12635-6

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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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Effect of MB on the priming step of inflammasome activation and expression of other cytokine genes. (A and B) BMDMs were treated with the indicated concentration of MB with/without LPS (10 ng/mL) as indicated in the schematic graph. (A) Pro-IL-1β and NLRP3 expression levels were analyzed by immunoblotting and further presented with band density. (B) Expression levels of mouse IL-1β, IL-1α, IL-6, IL-10, IL-12b, and TNFα mRNAs were quantitated by real-time PCR. C, BMDMs were treated with MB and/or LPS as the 1^st signal, after which cells were replaced by media containing nigericin (NG, 2^nd signal) with/without MB as the 2^nd signal. IL-1β and IL-18 secretion levels were measured by ELISA, and Casp1 secretion and pro-IL1β expression were analyzed by immunoblotting. All immunoblot data shown are representative of at least three independent experiments. Bar graph presents the mean ± SD.