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. 2018 Jun 20;8:9408. doi: 10.1038/s41598-018-27614-8

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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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Hesperetin activates Nrf2 and reduces oxidative stress. (a–d) Primary differentiated myotubes were treated with hesperetin or DMSO. (a) Taqman analysis of Nrf2 (gene symbol Nfe2l2) gene expression (n = 4)). (b) Taqman analysis of Nrf2-ARE target genes (n = 6). (c) DHE live cell imaging of oxidative stress 30 min after stimulation with 100 µM TBHP. Images were taken with ImageXPress and % positive DHE area was analyzed (2 independent experiments, n = 8). (d) Measurement of reduced (GSH) and oxidized (GSSG) forms of glutathione in myotubes treated with 150 µM TBHP for 1 h (n = 6). (e) ATP content was measured after 24 h incubation of differentiated immortalized human myotubes with hesperetin or DMSO (control) with or without BCNU, an inhibitor of glutathione reductase activity, in low glucose medium (3 independent experiments, n ≥ 24). Values represent means ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 (two-tailed t-test (a–c), one-Way Anova with Dunnett’s test (d), and 2-way ANOVA with Sidak’s test (e). Significant interaction (P < 0.01) between treatment factors indicates that BCNU reduced hesperetin-induced stimulation ATP content.