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. 2018 Feb 6;8:2477. doi: 10.1038/s41598-018-20821-3

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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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Regulation of ROCKs during adipogenesis. (A–C) 3T3-L1 cells were differentiated through incubation in DMI with or without KD025 for the indicated time points. (A) mRNA expression levels of ROCK1 and ROCK2 were measured by qRT-PCR. (B) Protein expression levels of ROCK1, ROCK2, p-cofilin, p-MLC, p-MYPT1, p-ERM, ERM and RhoA were analyzed by immunoblot. PPARγ was used as a marker of adipogenesis and β-tubulin as a loading control. (C) The relative intensity of phosphorylation levels of cofilin, ERM, and MLC were compared between vehicle- and KD025-treated conditions. *p < 0.05; **p < 0.01; ***p < 0.001 vs. the corresponding control. (D,E) 3T3-L1 pre-adipocytes were treated with KD025 (7 µM), Y-27632 (10 µM), fasudil (10 µM), H-1152P (3 µM), or SR3677 (5 µM) for one day. The phosphorylation levels of cofilin, ERM, and MLC were measured by immunoblot. β-tubulin was used as a loading control. *p < 0.05; **p < 0.01; ***p < 0.001 vs. vehicle-treated. The data are the representative from more than three independent experiments. Data are expressed as means ± S.E. based on triplicate.