Figure 2

Characterization of the MyHC isoforms in the muscles of BCAAem-treated and untreated mdx mice. Cropped image of a representative Coomassie-Blue-stained protein gel showing electrophoretic separation of the MyHC IIx and IIb isoforms from the total protein lysates of VM and TA muscles of male and female mdx mice. (a) Cropped image of a western blot of β-actin from the total protein lysates of VM and TA muscles of male and female mdx mice. (a) Full-length Coomassie-stained gels and blots are presented in Supplementary Fig. S8. Quantification of the myosin/actin ratio to describe the different amounts of myosin isoforms. (a) Whole field (b) (scale bar, 500 µm) and higher magnification (c) (scale bar, 50 µm) of ATPase (pH 4.3) (left) and SDH-stained (right, scale bar, 100 µm) sections for detection of the distribution and composition of myosin heavy chain (MyHC) isoforms (d) of BCAAem-treated and untreated mdx groups. qRT-PCR quantification analysis of expression of the MYH1, MYH2 and MYH4 genes in the VM and TA muscles of BCAAem-treated and untreated male and female mdx mice (n = 5 per group) normalized to expression of the GAPDH housekeeping gene. (e) Fast MyHC isoform immunofluorescence staining of BCAAem-treated and untreated VM and TA mdx male and female muscle sections (f) (scale bar 100 µm). The fast MyHC isoform fluorescence was quantified with the ImageJ software 6.0 and the corresponding histograms are reported as percentage of positive fibers per section. (g) All the experiments were conducted in triplicate. All data are presented as the mean ± s.e.m. Statistical error analysis was performed by two-way ANOVA with Bonferroni correction; *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001 indicate comparisons that reflect significant differences relative to the untreated group.