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. 2018 Sep 17;8:13932. doi: 10.1038/s41598-018-32163-1

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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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Effect of EPS-induced continuous contraction on maturation of myofiber sheets. (A) Contraction displacement of myofiber sheets cultured with no stimulation for 10 days (EPS(−)) and with no stimulation for 7 days and then continuous stimulation for 3 days (EPS(+)). Continuous EPS (voltage: 10 V, frequency: 1 Hz, duration: 3 ms) was applied for 1 h with 3 h rest. This procedure started at day 7 after induction of differentiation and was repeated for 3 days. To compare their contraction abilities, EPS was applied at 1 Hz frequency for 5 seconds. (n = 3) (B) Distribution and average value of myofiber diameters with and without continuous EPS for 2 weeks. The data were summarized from three myofiber sheets for each group (total number of myofibers: more than 150). (**P < 0.01) (C) Effect of continuous EPS on contractile ability of myofiber sheets for 2 weeks. Two different continuous EPS (triangle: 3 ms, circle: 10 ms) were applied for 1 h with 3 h rest. This procedure started at day 7 after induction of differentiation (Day 0 for EPS exercise) and was repeated for 2 weeks (Day 14). As the control, myofiber sheets were cultured without continuous EPS (square). The displacements of the shortening myofibers were monitored at Day 0, 7, and 14. (n = 5) (*P < 0.05 vs other samples at the same time point).