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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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Inhibitory effects of the MALT1 inhibitor MI-2 on osteoclast formation. The inhibitory effects of MI-2 on osteoclast formation from monocytes was determined by performing in vitro osteoclast-differentiation assays. (a) Structure of the small molecule MI-2, a MALT1 inhibitor. (b) Representative images from more than 3 independent experiments are shown. (c) Relative value (%) of the number of TRAP-positive osteoclasts per well. Pooled results are shown from 3 independent experiments. (d) TRAP enzyme activities (%) in untreated cells and cells treated with the indicated concentrations of MI-2. (e) Human monocytes were incubated in differentiation medium (α-MEM containing 10% FBS, 20 ng/mL MCSF, and 50 ng/mL RANKL). Protein expression in the cell lysates was analysed by immunoblotting with an anti-MALT1 antibody. The level of β-actin was analysed as a loading control. Full-length blots are presented in Supplementary Figure 1. The results shown are representative of 3 different independent experiments. *, **, and *** indicate significant differences from control (untreated) cells based on 2-tailed unpaired Student’s t-tests at P < 0.05, P < 0.01, and P < 0.001, respectively. The error bars show standard error of the mean (s.e.m.) values.