The order of exercise during concurrent training for rehabilitation does not alter acute genetic expression, mitochondrial enzyme activity or improvements in muscle function

Abstract

Concurrent exercise combines different modes of exercise (e.g., aerobic and resistance) into one training protocol, providing stimuli meant to increase muscle strength, aerobic capacity and mass. As disuse is associated with decrements in strength, aerobic capacity and muscle size concurrent training is an attractive modality for rehabilitation. However, interference between the signaling pathways may result in preferential improvements for one of the exercise modes. We recruited 18 young adults (10 ♂, 8 ♀) to determine if order of exercise mode during concurrent training would differentially affect gene expression, protein content and measures of strength and aerobic capacity after 2 weeks of knee-brace induced disuse. Concurrent exercise sessions were performed 3x/week for 6 weeks at gradually increasing intensities either with endurance exercise preceding (END>RES) or following (RES>END) resistance exercise. Biopsies were collected from the vastus lateralis before, 3 h after the first exercise bout and 48 h after the end of training. Concurrent exercise altered the expression of genes involved in mitochondrial biogenesis (PGC-1α, PRC, PPARγ), hypertrophy (PGC-1α4, REDD2, Rheb) and atrophy (MuRF-1, Runx1), increased electron transport chain complex protein content, citrate synthase and mitochondrial cytochrome c oxidase enzyme activity, muscle mass, maximum isometric strength and VO 2peak. However, the order in which exercise was completed (END>RES or RES>END) only affected the protein content of mitochondrial complex II subunit. In conclusion, concurrent exercise training is an effective modality for the rehabilitation of the loss of skeletal muscle mass, maximum strength, and peak aerobic capacity resulting from disuse, regardless of the order in which the modes of exercise are performed.

Figure 1. Concurrent exercise program design with weekly intensities for aerobic and resistance exercise protocols.

Figure 2. Longitudinal study design.

(Bx) - Timing of muscle biopsy.

Figure 3. Concurrent exercise alters mRNA content of genes involved in mitochondrial biogenesis and metabolism and increases protein content of subunits of the ETC.

A - Gene expression changes in skeletal muscle 3 hours after concurrent rehabilitative exercise for genes involved in aerobic exercise adaption. Peroxisome proliferative activated receptor (PPAR) gamma coactivator-1α (PGC-1α), PGC-1-related coactivator (PRC), PGC-1β, and PPARγ. B - Fold changes in protein content from pre-exercise for subunits of the 5 complexes of the ETC (CI-CV) following 6 weeks of concurrent rehabilitative exercise. Light bars indicate END>RES group, dark bars indicate RES>END group. Representative western blotting image for pre-exercise and post included. *Significant difference from baseline (P<0.05). **Significant difference from baseline (P<0.01). ***Significant difference from baseline (P<0.001). Bar indicates main effect for time and not exercise group. Mean ± SD.

Figure 4. Concurrent exercise alters mRNA content of genes involved in the regulation of mTOR signaling, and proposed IGF-1 and myostatin gene expression.

Gene expression changes in skeletal muscle 3 hours after concurrent rehabilitative exercise for genes involved in resistance exercise adaption. Light bars indicate END>RES group, dark bars indicate RES>END group. Regulated in DNA damage 1 (REDD1) and 2 (REDD2), Ras homolog enriched in brain (Rheb), and peroxisome proliferative activated receptor (PPAR) gamma coactivator-1α, isoform 4 (PGC-1α4), ***Significant difference from baseline (P<0.001). Bar indicates main effect for time and not exercise group. Mean ± SD.

Figure 5. Concurrent exercise acutely increases mRNA content of genes involved in the regulation of muscle atrophy but does not affect chronic protein content of selected E3 ligases or total ubiquitination.

A - Gene expression changes in skeletal muscle 3 hours after concurrent rehabilitative exercise for genes involved in regulation of protein breakdown. Atrogin-1, muscle-specific RING finger-1 (MuRF-1), neural precursor cell expressed developmentally downregulated protein 4 (Nedd4), and Runt-related transcription factor 1 (Runx1). B - Fold changes in protein content from pre-exercise levels for muscle specific (Atrogin-1 and MuRF-1) and ubiquitously expressed (Nedd4) E3 ligases, and total ubiquitination (Total Ub) following 6 weeks of concurrent rehabilitative exercise. Light bars indicate END>RES group, dark bars indicate RES>END group. Representative western blotting images for pre-exercise and post included. ***Significant difference from baseline (P<0.001). Bar indicates main effect for time and not exercise group. Mean ± SD.

Figure 6. Concurrent exercise increases mitochondrial enzyme activity.

Enzyme activities in skeletal muscle before (Pre-exercise) and after 6 weeks of concurrent rehabilitative exercise (Post). A - citrate synthase (CS), B - cytochrome c oxidase (COX) and C – COX/CS ratio. Light bars indicate END>RES group, dark bars indicate RES>END group. *Significant difference from baseline (P<0.05). ***Significant difference from baseline (P<0.001). Bar indicates main effect for time and not exercise group. Mean ± SD.

Figure 7. Concurrent exercise recovers maximum isometric strength loss induced by disuse and increases aerobic performance.

A - Values for maximum isometric strength of the knee extensors before immobilization (BL), before exercise (pre-exercise) and after 6 weeks of rehabilitative concurrent exercise (Post). B - peak oxygen consumption (VO_2peak) before immobilization (BL) and after 6 weeks of concurrent exercise (Post). Light bars indicate END>RES group, dark bars indicate RES>END group. **Significant difference from baseline (P<0.01). ***Significant difference from baseline (P<0.001). Bar indicates main effect for time and not exercise group. Mean ± SD.