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. 2017 Sep 19;7:11889. doi: 10.1038/s41598-017-12349-9

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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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Effects of MI-2 on inflammatory TNFα production in macrophages. (a) TNFα concentrations in the culture supernatants from untreated or LPS-treated macrophages, with or without MI-2 treatment, as measured by ELISA. Pooled results are shown from 3 independent experiments. ** and *** indicate significant differences based on 2-tailed unpaired Student’s t-tests at P < 0.01 and P < 0.001, respectively. The error bars show the s.e.m. values. (b) Human monocytes were incubated in differentiation medium (α-MEM containing 10% FBS, and 20 ng/mL MCSF), with or without 10 ng/mL LPS, at the indicated time points. Protein expression in the cell lysates was analysed by immunoblotting with antibodies against phosphor-p65 (p-p65), p65, phosphor-IkBα (p-IkBα), and IkBα. β-actin expression was analysed as a loading control. Full-length blots are presented in Supplementary Figure 2. The results shown are representative of 3 independent experiments. (c) Human monocytes were incubated in differentiation medium (α-MEM containing 10% FBS, and 20 ng/mL MCSF), with or without 10 ng/mL LPS, for 5 min. Protein expression in the cell lysates was analysed by immunoblotting with antibodies against A20. β-actin expression was analysed as a loading control. Full-length blots are presented in Supplementary Figure 3. The results shown are representative of 3 independent experiments. (d–e) The densities of p-p65, p65, p-IkBα, and IkBα bands were measured and normalised to β-actin levels. The p-p65/p65 and p-IkBα/β-actin ratios are shown. Pooled results are shown from 3 independent experiments. * and ** indicate significant differences from control (untreated) cells based on 2-tailed unpaired Student’s t-tests at P < 0.05 and P < 0.01, respectively. The error bars show the s.e.m. values.