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. 2018 Nov 1;8:16195. doi: 10.1038/s41598-018-34324-8

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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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Mg deficiency accelerates osteogenic differentiation. (A) Alizarin Red S staining was performed on hMSC cultured in 0.1 or 1.0 mM Mg with or without the osteogenic cocktail for 14 days. Whole well image (upper left panel) and photographs taken at 10X magnification (lower panel) are shown. Absorbance was measured at 550 nm after acid extraction (upper right panel). (B) Real-Time PCR was performed on RNA extracted from hMSC cultured in 1.0 or 0.1 mM Mg, additioned or not with the osteogenic cocktail, treated or untreated with bafilomycin A1 (10 nM) for 3 days. Primers designed on RUNX2 sequence were used. (C) Total Mg was measured using the fluorescent chemosensor DCHQ5 as described. (D) Western blot was performed on extracts from hMSC cultured in 1.0 or 0.1 mM Mg for 3 and 6 days using antibodies against TRPM7 or MagT1. Actin was used as a control of loading. A representative blot is shown and quantification is provided in the Supplementary information (Fig. S6A).