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. 2019 Mar 5;9:3454. doi: 10.1038/s41598-019-40042-6

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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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In vitro isolation, expansion and differentiation of human cremaster muscle-derived myogenic precursor cells. (A) Schematic representation of the male reproductive system anatomy, showing cremaster muscle (red) and the biopsy sample zone is highlighted by a rectangle. Outline of the myosphere suspension culture and myotube differentiation (adhesion culture) steps. (B,C) Optical microscope images of the suspension culture showing cells and tissue fragments (arrow) at day 0 (B) and spheres at day 7 (C). (D–F) Optical microscope images of the adhesion culture at day 2 (D) and day 9 (E,F), where multinucleated myotubes (arrows) become predominant. Scale bars, 100 μm.