Hydroxyproline colorimetric assay kit

Gastrocnemius muscle samples were embedded in optimal cutting temperature mounting medium (Histolab Products, Gothenburg, Sweden) and frozen in liquid nitrogen followed by cryosectioning and staining with haematoxylin-eosin (H-E; Histolab Products), Nile Red (Sigma-Aldrich, St Louis, MO, USA) or MitoTracker Red (Thermo Fisher Scientific, Waltham, MA, USA). Enzymatic stainings were performed as previously described [20 ]. For immunofluorescence, sections were incubated with primary antibodies followed by incubation with secondary antibodies (see ESM Table 1). Gastrocnemius muscle samples were also fixed with 4% formaldehyde in phosphate buffer (Histolab Products), embedded in paraffin, sectioned and stained with Picrosirius Red (Histolab Products) or Periodic acid–Schiff (PAS; Sigma-Aldrich). Ultrastructural analysis of gastrocnemius muscle was performed by transmission electron microscopy (TEM; LEO 912AB; Omega; Carl Zeiss NTS, Oberkochen, Germany) as previously described [21 (link)]. Gastrocnemius muscle homogenates were analysed using a Hydroxyproline Colorimetric Assay Kit (Sigma-Aldrich) and a Triglyceride Calorimetric Assay Kit (Biovision, Mountain View, CA, USA).