Fig. 4: JQ1 decreases oxidative stress in the mdx muscle and in H₂O₂-treated cells.

a Representative images of 8-OHdG staining of muscle cross-sections in control, vehicle- and JQ1-treated TAs (n = 6 sections examined from n = 3 animals for each experimental group). Scale bar: 50 µm. Right panel: quantification of the staining. Data are expressed as a mean ± SD. a indicates statistical significance compared to Control group; b indicates statistical significance compared to the mdx mice animal group. b Representative images of 8-OHdG staining of C2C12 cells. Cells were pretreated with JQ1 (200 nM) for 24 h and then stimulated with H₂O₂ for 24 h. Scale bar: 50 µm. Right panel: quantification of the staining derived from three independent experiments. n = 6 fields examined from n = 3 independent experiments. Data are expressed as a mean ± SD. a indicates statistical significance compared to Control cells; b indicates statistical significance compared to the H₂O₂-treated C2C12. c Representative western blot for LC3 (c, n = 4; H₂O₂, n = 3; H₂O₂ + JQ1, n = 4) and p62 (n = 3 independent experiments) in C2C12 myotube extracts of control and H₂O₂- and H₂O₂/JQ1-treated cells for 24 h. Myotubes were pretreated with JQ1 (200 nM) for 24 h and then stimulated with H₂O₂ for 24 h. In order to study the autophagy flux, the experiment was performed by pre-treating cells with 30 µM of the lysosomotropic agent chloroquine. Vinculin serves as a loading control. Right panel: quantification of normalized band intensity derived from three different experiments. Data represent means ± SD. a indicates statistical significance compared to Control cells; b indicates statistical significance compared to the H₂O₂-treated C2C12. d Representative western blot for Sirt1, p-AMPK, AMPK, Ulk1, p-Ulk1 (Ser555), Akt, p-Akt (Ser473) in C2C12 myoblast extracts of control and H₂O₂- and H₂O₂/JQ1-treated cells. Myotubes were pretreated with JQ1 (200 nM) for 24 h and then stimulated with H₂O₂ for 24 h. Vinculin serves as a loading control. Right panel: quantification of normalized band intensity derived from at least four different experiments. Data represent means ± SD, n = 4 independent experiments. a indicates statistical significance compared to Control cells; b indicates statistical significance compared to the H₂O₂-treated C2C12. In all panels, statistical analysis was assessed by using one-way ANOVA followed by Tukey’s post hoc test. *P < 0.05; **P < 0.01; ***P < 0.001.