Fig. 6
S1P pre-conditioning boosts ATP production by enhancing mitochondrial density; 0.2 × 105/cm2 C2C12 cells were seeded and maintained in DMEM-HG media and induced to differentiation at 90–95% confluence by shifting to DM (DMEM HG with 2% HS). a S1P (1 μM) and VC pretreated differentiating C2C12 cells under normoxia (N) and 0.5% hypoxia (H) at pre-defined study time points were stained by MitoTracker and imaged by an inverted fluorescence microscope at ×10 magnification. Scale bar 100 µm. b Mitochondrial mass (mitotracker intensity) was also quantified by a multimode plate reader (FLUOstar Omega), and average values relative to 0D were reported. c ATP was measured in differentiating C2C12 cell lysate using a commercial kit as per the manufacturer’s protocol. Data are reported as mean ± SD from three independent experiments carried out in triplicate after normalization with cell lysate (mg/ml). Statistical analysis was performed using the one-way ANOVA/post hoc Bonferroni’s analysis method. Statistical significance is denoted by NVC vs. HVC ($p < 0.05, *p ≤ 0.005) and HVC vs. HS1P (@p < 0.05, #p ≤ 0.005) at respective time points