The influence of endurance training on oxidative phosphorylation and the susceptibility of mitochondrial oxidative function to reactive oxygen species (ROS) was investigated in skeletal muscle of four men and four women. Mitochondria were isolated from muscle biopsies taken before and after 6 weeks of endurance training. Mitochondrial respiration was measured before and after exposure of mitochondria to exogenous ROS (H2O2+ FeCl2).
Endurance training increased peak pulmonary O2 uptake (V̇O2,peak) by 24 % and maximal ADP‐stimulated mitochondrial oxygen consumption (state 3) by 40 % (P < 0.05). Respiration in the absence of ADP (state 4), the respiratory control ratio (RCR = state 3/state 4) and the ratio between added ADP and consumed oxygen (P/O) remained unchanged by the training programme.
Exposure to ROS reduced state 3 respiration but the effect was not significantly different between pre‐ and post‐training samples. State 4 oxygen consumption increased after exposure to ROS both before (+189 %, P < 0.05) and after training (+243 %, P < 0.05) and the effect was significantly higher after training (P < 0.05, pre‐ vs. post‐training). The augmented state 4 respiration could in part be attenuated by atractyloside, which indicates that ADP/ATP translocase was affected by ROS. The P/O ratio in ROS‐treated mitochondria was significantly lower (P < 0.05) compared to control conditions, both before (−18.6 ± 2.2 %) and after training (−18.5 ± 1.1 %).
Muscle activities of superoxide dismutase (mitochondrial and cytosolic), glutathione peroxidase and muscle glutathione status were unaffected by training. There was a positive correlation between muscle superoxide dismutase activity and age (r= 0.75; P < 0.05; range of age 20–37 years), which may reflect an adaptation to increased generation of ROS in senescent muscle. The muscle glutathione pool was more reduced in subjects with high activity of glutathione peroxidase (r= 0.81; P < 0.05).
The influence of short‐term training on mitochondrial oxygen consumption has for the first time been investigated in human skeletal muscle. The results showed that maximal mitochondrial oxidative power is increased after endurance training but that the efficiency of energy transfer (P/O ratio) remained unchanged. Antioxidative defence was unchanged after training when expressed relative to muscle weight. Although this corresponds to a reduced antioxidant protection per individual mitochondrion, the sensitivity of aerobic energy transfer to ROS was unchanged. However, the augmented ROS‐induced non‐coupled respiration after training indicates an increased susceptibility of mitochondrial membrane proton conductance to oxidative stress.
A novel hypothesis-free multivariate screening methodology for the study of human exercise metabolism in blood serum is presented. Serum gas chromatography/time-of-flight mass spectrometry (GC/TOFMS) data was processed using hierarchical multivariate curve resolution (H-MCR), and orthogonal partial least-squares discriminant analysis (OPLS-DA) was used to model the systematic variation related to the acute effect of strenuous exercise. Potential metabolic biomarkers were identified using data base comparisons. Extensive validation was carried out including predictive H-MCR, 7-fold full cross-validation, and predictions for the OPLS-DA model, variable permutation for highlighting interesting metabolites, and pairwise t tests for examining the significance of metabolites. The concentration changes of potential biomarkers were verified in the raw GC/TOFMS data. In total, 420 potential metabolites were resolved in the serum samples. On the basis of the relative concentrations of the 420 resolved metabolites, a valid multivariate model for the difference between pre- and post-exercise subjects was obtained. A total of 34 metabolites were highlighted as potential biomarkers, all statistically significant (p < 8.1E-05). As an example, two potential markers were identified as glycerol and asparagine. The concentration changes for these two metabolites were also verified in the raw GC/TOFMS data. The strategy was shown to facilitate interpretation and validation of metabolic interactions in human serum as well as revealing the identity of potential markers for known or novel mechanisms of human exercise physiology. The multivariate way of addressing metabolism studies can help to increase the understanding of the integrative biology behind, as well as unravel new mechanistic explanations in relation to, exercise physiology.
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