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. 2018 Jul 6;8:10289. doi: 10.1038/s41598-018-28213-3

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© The Author(s) 2018

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 YM155 on the protein expression of p65 NF-κB. RCC786.0 was treated with 40 nM of YM155 in the absence or presence of TNF-α stimulation. Using western immunoblotting analysis, the expression levels of the phosphorylated (phospho-) and total p65 were probed at 6 h and 24 h. In the nuclear extract of YM155 treated cells, (A) there was an increase in p65 NF-κB at 6 h, (B) there was a decrease in p65 NF-κB at 24 h. The increased/decreased was greater in cells stimulated with TNF-α. There was no concurrent increase in levels of phosphorylated p65. In the cytoplasmic extract of YM155 treated cells, (C) there was an increase in phospho-p65 but total p65 was not affected at 6 h, (D) there was no significant change in total p65 at 24 h and phospho- p65 was maintained at basal levels. TATA-box binding protein (TBP) and beta actin (β-actin) were used as loading control for nuclear and cytoplasmic fraction respectively.