Key points: Several biochemical measures of mitochondrial components are used as biomarkers of mitochondrial content and muscle oxidative capacity. However, no studies have validated these surrogates against a morphological measure of mitochondrial content in human subjects. The most commonly used markers (citrate synthase activity, cardiolipin content, mitochondrial DNA content (mtDNA), complex I–V protein, and complex I–IV activity) were correlated with a measure of mitochondrial content (transmission electron microscopy) and muscle oxidative capacity (respiration in permeabilized fibres). Cardiolipin content followed by citrate synthase activity and complex I activity were the biomarkers showing the strongest association with mitochondrial content. mtDNA was found to be a poor biomarker of mitochondrial content. Complex IV activity was closely associated with mitochondrial oxidative phosphorylation capacity. Abstract  Skeletal muscle mitochondrial content varies extensively between human subjects. Biochemical measures of mitochondrial proteins, enzyme activities and lipids are often used as markers of mitochondrial content and muscle oxidative capacity (OXPHOS). The purpose of this study was to determine how closely associated these commonly used biochemical measures are to muscle mitochondrial content and OXPHOS. Sixteen young healthy male subjects were recruited for this study. Subjects completed a graded exercise test to determine maximal oxygen uptake () and muscle biopsies were obtained from the vastus lateralis. Mitochondrial content was determined using transmission electron microscopy imaging and OXPHOS was determined as the maximal coupled respiration in permeabilized fibres. Biomarkers of interest were citrate synthase (CS) activity, cardiolipin content, mitochondrial DNA content (mtDNA), complex I–V protein content, and complex I–IV activity. Spearman correlation coefficient tests and Lin's concordance tests were applied to assess the absolute and relative association between the markers and mitochondrial content or OXPHOS. Subjects had a large range of (range 29.9–71.6 ml min⁻¹ kg⁻¹) and mitochondrial content (4–15% of cell volume). Cardiolipin content showed the strongest association with mitochondrial content followed by CS and complex I activities. mtDNA was not related to mitochondrial content. Complex IV activity showed the strongest association with muscle oxidative capacity followed by complex II activity. We conclude that cardiolipin content, and CS and complex I activities are the biomarkers that exhibit the strongest association with mitochondrial content, while complex IV activity is strongly associated with OXPHOS capacity in human skeletal muscle.

Obesity is associated with low-grade inflammation, insulin resistance, type 2 diabetes, and cardiovascular disease. This study investigated the effect of a 15-wk lifestyle intervention (hypocaloric diet and daily exercise) on inflammatory markers in plasma, adipose tissue (AT), and skeletal muscle (SM) in 27 severely obese subjects (mean body mass index: 45.8 kg/m²). Plasma samples, subcutaneous abdominal AT biopsies, and vastus lateralis SM biopsies were obtained before and after the intervention and analyzed by ELISA and RT-PCR. The intervention reduced body weight ( P < 0.001) and increased insulin sensitivity (homeostasis model assessment; P < 0.05). Plasma adiponectin ( P < 0.001) increased, and C-reactive protein ( P < 0.05), IL-6 ( P < 0.01), IL-8 ( P < 0.05), and monocyte chemoattractant protein-1 ( P < 0.01) decreased. AT inflammation was reduced, determined from an increased mRNA expression of adiponectin ( P < 0.001) and a decreased expression of macrophage-specific markers (CD14, CD68), IL-6, IL-8, and tumor necrosis factor-α ( P < 0.01). After adjusting for macrophage infiltration in AT, only IL-6 mRNA was decreased ( P < 0.05). Only very low levels of inflammatory markers were found in SM. The intervention had no effect on adiponectin receptor 1 and 2 mRNA in AT or SM. Thus hypocaloric diet and increased physical activity improved insulin sensitivity and reduced low-grade inflammation. Markers of inflammation were particularly reduced in AT, whereas SM does not contribute to this attenuation of whole body inflammation.

MicroRNAs (miRNA), small noncoding RNA molecules, may regulate protein synthesis, while resistance exercise training (RT) is an efficient strategy for stimulating muscle protein synthesis in vivo. However, RT increases muscle mass, with a very wide range of effectiveness in humans. We therefore determined the expression level of 21 abundant miRNAs to determine whether variation in these miRNAs was able to explain the variation in RT-induced gains in muscle mass. Vastus lateralis biopsies were obtained from the top and bottom ∼20% of responders from 56 young men who undertook a 5 day/wk RT program for 12 wk. Training-induced muscle mass gain was determined by dual-energy X-ray absorptiometry, and fiber size was evaluated by histochemistry. The expression level of each miRNA was quantified using TaqMan-based quantitative PCR, with the analysis carried out in a blinded manner. Gene ontology and target gene profiling were used to predict the potential biological implications. Of the 21 mature miRNAs examined, 17 were stable during RT in both groups. However, miR-378, miR-29a, miR-26a, and miR-451 were differentially expressed between low and high responders. miR-378, miR-29a, and miR-26a were downregulated in low responders and unchanged in high responders, while miR-451 was upregulated only in low responders. Interestingly, the training-induced change in miR-378 abundance was positively correlated with muscle mass gains in vivo. Gene ontology analysis of the target gene list of miR-378, miR-29a, miR-26a, and miR-451, from the weighted cumulative context ranking methodology, indicated that miRNA changes in the low responders may be compensatory, reflecting a failure to “activate” growth and remodeling genes. We report, for the first time, that RT-induced hypertrophy in human skeletal muscle is associated with selected changes in miRNA abundance. Our analysis indicates that miRNAs may play a role in the phenotypic change and pronounced intergroup variation in the RT response.

Transcription of metabolic genes is transiently induced during recovery from exercise in skeletal muscle of humans. To determine whether pre‐exercise muscle glycogen content influences the magnitude and/or duration of this adaptive response, six male subjects performed one‐legged cycling exercise to lower muscle glycogen content in one leg and then, the following day, completed 2.5 h low intensity two‐legged cycling exercise. Nuclei and mRNA were isolated from biopsies obtained from the vastus lateralis muscle of the control and reduced glycogen (pre‐exercise glycogen = 609 ± 47 and 337 ± 33 mmol kg⁻¹ dry weight, respectively) legs before and after 0, 2 and 5 h of recovery. Exercise induced a significant (P < 0.05) increase (2‐ to 3‐fold) in transcription of the pyruvate dehydrogenase kinase 4 (PDK4) and uncoupling protein 3 (UCP3) genes in the reduced glycogen leg only. Although PDK4, lipoprotein lipase (LPL) and hexokinase II (HKII) mRNA were elevated in the reduced glycogen leg before exercise, no consistent difference was found between the two legs in response to exercise. In a second study, six subjects completed two trials (separated by 2 weeks) consisting of 3 h of two‐legged knee extensor exercise with either control (398 ± 52 mmol kg⁻¹ dry weight) or low (240 ± 38 mmol kg⁻¹ dry weight) pre‐exercise muscle glycogen. Exercise induced a significantly greater increase in PDK4 transcription in the low glycogen (> 6‐fold) than in the control (< 3‐fold) trial. Induction of PDK4 and UCP3 mRNA in response to exercise was also signficantly higher in the low glycogen (11.4‐ and 3.5‐fold, respectively) than in the control (5.0‐ and 1.7‐fold, respectively) trial. These data indicate that low muscle glycogen content enhances the transcriptional activation of some metabolic genes in response to exercise, raising the possibility that signalling mechanisms sensitive to glycogen content and/or FFA availability may be linked to the transcriptional control of exercise‐responsive genes.

In this study, the hypothesis that the release of interleukin (IL)‐6 from human muscle is linked to exercise intensity and muscle glucose uptake was investigated. In the overnight fasted state, seven healthy males performed knee extension exercise, kicking with both legs, each at 25 % of maximal power (W_max) for 45 min (eliciting 23 ± 1 % of pulmonary maximal oxygen uptake, V̇_O2,max) and then simultaneously with one leg at 65 % and the other leg at 85 % W_max for 35 min (40 ± 1 % of pulmonary V̇_O2,max). Blood was sampled from a femoral artery and both femoral veins, and blood flow was determined by thermodilution. Thigh plasma flow (0.15 ± 0.01, 1.4 ± 0.2, 2.0 ± 0.1 and 2.3 ± 0.2 l min⁻¹ thigh⁻¹ at rest and 25 %, 65 % and 85 % W_max, respectively) and thigh oxygen uptake (0.02 ± 0.01, 0.27 ± 0.03, 0.48 ± 0.04 and 0.55 ± 0.05 l min⁻¹ thigh⁻¹ at rest and 25 %, 65 % and 85 % W_max, respectively) increased with increasing exercise intensity (P < 0.05). Also, thigh IL‐6 release (0.4 ± 0.1, 1.3 ± 0.5, 1.5 ± 0.6 and 2.5 ± 0.7 ng min⁻¹ thigh⁻¹ at rest and 25 %, 65 % and 85 % W_max, respectively) and thigh glucose uptake (0.05 ± 0.01, 0.3 ± 0.05, 0.75 ± 0.16, 1.07 ± 0.15 mmol min⁻¹ thigh⁻¹ at rest and 25 %, 65 % and 85 % W_max, respectively) increased with increasing exercise intensity (P < 0.05). During the last 35 min of exercise, arterial catecholamine concentrations were higher (P < 0.05) than at rest and during low‐intensity exercise. During exercise, thigh IL‐6 release was positively related to both thigh glucose uptake (P < 0.001) and thigh glucose delivery (P < 0.005), but not to thigh glucose extraction. Thigh IL‐6 release was also positively related to arterial plasma adrenaline concentration. The pre‐exercise muscle glycogen concentration tended to correlate with the arteriovenous IL‐6 concentration difference at rest, and the postexercise glycogen concentration was inversely correlated with IL‐6 release during the final 35 min of exercise. In conclusion, the study indicates that IL‐6 release from human muscle is positively related to exercise intensity, arterial adrenaline concentration and muscle glucose uptake. This supports the hypothesis that IL‐6 may be linked to the regulation of glucose homeostasis during exercise. The observation of a relationship between IL‐6 release and muscle glycogen store both at rest and after exercise suggests that IL‐6 may act as a carbohydrate sensor.