Journal: Frontiers in Physiology
Article Title: Targeting ERRs to counteract age-related muscle atrophy associated with physical inactivity: a pilot study
doi: 10.3389/fphys.2025.1616693
Figure Lengend Snippet: Effects of SLU-PP-332 treatment in myoblasts from inactive subjects on cell viability, cytotoxicity, oxidative stress, and senescence β-galactosidase activity (SA- β-gal). (a–d) : Immunofluorescence for Pax7 and MyoD in myoblasts: (a) Nuclei are stained with DAPI (blue); (b) Immunostaining for Pax7 (red); (c) Immunostaining for MyoD (green); (d) Merge for Pax7 and MyoD signals. 40× images, scale bar represents 100 μm. (e) MTS assay: the half inhibitory concentration (IC50) was obtained at a dosage between 1 × 10 −3 M and 2.5 × 10 −3 M (n = 9 from N = 3 experiments). (f) Lactate dehydrogenase (LDH) cytotoxicity assay: significant reduction of 16.1% of cell damage in SLU-PP-332-treated myoblasts (Inactive_T) compared with untreated cells (Inactive_NT) (p < 0.0001) (n = 25 from N = 5 experiments). (g) Intracellular reactive oxygen species (ROS) levels: significant reduction of 37.7% of oxidative stress in SLU-PP-332-treated myoblasts (Inactive_T) compared with untreated cells (Inactive_NT) (p < 0.0001) (n = 25 from N = 5 experiments). (h) Reduced glutathione (GSH) assay: significant increase of 117.4% in intracellular GSH levels in SLU-PP-332-treated myoblasts (Inactive_T) compared with untreated cells (Inactive_NT) (p < 0.0001) (n = 25 from N = 5 experiments). (i) SA-β-gal assay: significant reduction of 26.1% of enzymatic activity in SLU-PP-332-treated myoblasts (Inactive_T) compared with untreated cells (Inactive_NT) (p < 0.0001) (n = 25 from N = 5 experiments).
Article Snippet: Briefly, after fixation in 4% paraformaldehyde dissolved in 0.9% saline solution for 30 min, cell cultures were pretreated with EDTA citrate, pH 7.8 for 20 min at 95 C, and incubated for 1 h with rabbit polyclonal anti-Pax7 antibody (dilution 1:100; ab187339, AbCam, Cambridge, United Kingdom), mouse monoclonal anti-MyoD antibody (dilution 1:100; InvitrogenTM, ThermoFisher Scientific, United States), rabbit polyclonal anti-ERRα antibody (dilution 1:100; A90033, antibodies.com, Stockholm, Sweden), or mouse monoclonal anti-MyHC antibody (diluition 1:100; ab51263, AbCam, Cambridge, United Kingdom).
Techniques: Activity Assay, Immunofluorescence, Staining, Immunostaining, MTS Assay, Concentration Assay, LDH Cytotoxicity Assay, GSH Assay
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