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. 2017 Oct 10;7:12879. doi: 10.1038/s41598-017-12861-y

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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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ERK and JNK activity are necessary for TNFα-induced DUSP5 gene expression. Adipocytes were pre-treated with the MEK inhibitor (U0126; ERKi)), JNK inhibitor (SP600125; JNKi) or p38 inhibitor (SB203580; p38i) for 30 min prior to stimulation with 100 pM TNFα. Protein was collected 15 min and RNA isolated 30 min post-TNFα stimulation and (A) immunoblotted for phospho-ERK, JNK, p38, c-Jun and MAPKAPK2 as well as total ERK, JNK, p38 and c-Jun. (B) DUSP5 mRNA expression was assessed via qPCR. Adipocytes were pre-treated with two independent MEK inhibitors U0126 and PD98059 for 30 min prior to stimulation with 100 pM TNFα. Protein was collected 15 min and RNA isolated 30 min post-TNFα stimulation and (C) immunoblotted for phospho-ERK as well as total ERK. (D) DUSP5 mRNA expression was assessed via qPCR. Adipocytes were pre-treated with the JNK inhibitors SP600125 or JNK-IN-8 for 30 min prior to stimulation with 100 pM TNFα. (E) Protein was collected 15 min post-TNFα stimulation and immunoblotted for phospho-JNK and total JNK. (F) RNA was isolated 30 min post-TNFα and qPCR used to examine DUSP5 mRNA expression. ANOVA followed by Tukey’s post-hoc using GraphPad Prism software was used to determine significance p < 0.05.