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. 2018 Aug 17;8:12307. doi: 10.1038/s41598-018-29968-5

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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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In vitro cell density optimization. (A) Live/dead staining images and (B) quantification of bioprinted muscle constructs with cell densities of 10, 20, 30, and 50 × 10⁶ cells/ml at 1 day in culture (n = 6, 5 random fields/sample, no significant difference). (C) TUNEL assay of bioprinted muscle constructs after 6 days in culture. Apoptotic cells were calculated with different cell densities (n = 3, 5 random fields/sample, no significant difference). (D) MHC immunofluorescent images of bioprinted muscle constructs at 6 days in culture (after 5-day differentiation). Representative immunofluorescent images for MHC (red) showed that bioprinted hMPCs in the constructs with different cell densities were formed into longitudinally aligned myofibers.