Chamber oxygen concentration impacts mitochondrial function and hydrogen peroxide appearance in permeabilized human skeletal muscle fibers

Skeletal muscle mitochondrial respiration is commonly assessed ex vivo using permeabilized fibers in media with high oxygen (O2) concentrations to ensure that O2 availability does not limit respiration. However, high O2 concentrations also increase the production of reactive O2 species that can negatively affect respiration. In this study, we tested the hypotheses that permeabilized fiber mitochondria in a high, compared to low, O2 concentration would (i) not be different at maximal state 3 respiration rate (Vmax), (ii) have lower submaximal respiration rates at submaximal O2 concentrations, and (iii) have greater total cumulative hydrogen peroxide (H2O2) appearance. We continuously monitored mitochondrial state 3 respiration and H2O2 appearance rates using high-resolution respirometry in permeabilized skeletal muscle fibers (12 untrained participants; 22 ± 4 yrs) with either control (~127 mmHg; CON) or high (~327 mmHg; HIGH) partial pressures of O2 (PO2). Vmax was not different between conditions (HIGH: 80.7 ± 16.7 vs. CON: 82.3 ± 18.7 pmol/s/mg, p = 0.695). The PO2 at 80 % Vmax (P80) was greater in HIGH (73.9 ± 25.5 vs. 28.0 ± 7.1 mmHg, p < 0.001) and respiration rates at 5-60 mmHg PO2 were lower for HIGH than CON (all p < 0.001). Additionally, the total cumulative H2O2 appearance was greater in HIGH than CON (n = 11; 51.5 ± 23.2 vs. 18.3 ± 10.3 pmol/mg, p < 0.001), and this difference was directly correlated with the difference in P80 (r = 0.655, p = 0.029). The current findings support that a high O2 concentration, by itself, does not appear to affect Vmax in the permeabilized skeletal muscle fiber preparation, but the corollary increase in H2O2 exposure may diminish mitochondrial state 3 respiratory function.