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. 2019 Oct 17;9:14910. doi: 10.1038/s41598-019-51410-7

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© The Author(s) 2019

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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SM suppresses C2C12 myogenic differentiation at an early phase of myogenesis. (A,B) C2C12 myoblasts were induced to differentiate either under 1 g or SM for 12 h. Cells were lysed and subjected to western blotting (A) or a quantitative RT-PCR analysis of MHC and myogenin expression (n = 6) (B). **P < 0.01 versus undiff. con; ^##P < 0.01 versus diff. con. (C,D) The cells were differentiated either under 1 g, or under SM for 48 h. (C) The cells were stained for MHC (green) and DAPI (red). scale bar = 50 μm (D) The stained images were quantified as described in Fig. 2C (n = 9). ^##P < 0.01 versus diff.con. (E–H) C2C12 myoblasts were differentiated for 48 h and shifted either to 1 g or SM for 36 h. (E) The cells were stained for MHC (green) and DAPI (red). scale bar = 50 μm. (F) The quantification of images was determined as described in Fig. 2C (n = 9). (G,H) Cells were lysed and subjected to western blotting (G) or a quantitative RT-PCR analysis of MHC and myogenin expression (n = 4) (H). *P < 0.05 versus undiff. con. Mouse GAPDH was used to normalize gene expression. Abbreviations: simulated microgravity (SM); differentiation (Diff); undifferentiated control (undiff. con); differentiated control (diff. con); differentiated under SM condition (diff.SM). Data are expressed as mean ± SD, with Mann-Whitney U test performed as indicated.