DNA methylation of exercise-responsive genes differs between trained and untrained men.

BACKGROUND: Physical activity is well known for its multiple health benefits and although the knowledge of the underlying molecular mechanisms is increasing, our understanding of the role of epigenetics in long-term training adaptation remains incomplete. In this intervention study, we included individuals with a history of > 15 years of regular endurance or resistance training compared to age-matched untrained controls performing endurance or resistance exercise. We examined skeletal muscle DNA methylation of genes involved in key adaptation processes, including myogenesis, gene regulation, angiogenesis and metabolism. RESULTS: A greater number of differentially methylated regions and differentially expressed genes were identified when comparing the endurance group with the control group than in the comparison between the strength group and the control group at baseline. Although the cellular composition of skeletal muscle samples was generally consistent across groups, variations were observed in the distribution of muscle fiber types. Slow-twitch fiber type genes MYH7 and MYL3 exhibited lower promoter methylation and elevated expression in endurance-trained athletes, while the same group showed higher methylation in transcription factors such as FOXO3, CREB5, and PGC-1α. The baseline DNA methylation state of those genes was associated with the transcriptional response to an acute bout of exercise. Acute exercise altered very few of the investigated CpG sites. CONCLUSIONS: Endurance- compared to resistance-trained athletes and untrained individuals demonstrated a different DNA methylation signature of selected skeletal muscle genes, which may influence transcriptional dynamics following a bout of acute exercise. Skeletal muscle fiber type distribution is associated with methylation of fiber type specific genes. Our results suggest that the baseline DNA methylation landscape in skeletal muscle influences the transcription of regulatory genes in response to an acute exercise bout.

Circulating immune cell populations at rest and in response to acute endurance exercise in young adults with cerebral palsy.

AIM: The aim of this observational study was to determine the immune status and function in young adults with cerebral palsy (CP) in comparison to typically developing individuals. METHOD: Blood samples from 12 individuals with CP (five males, seven females; mean age: 25 years 1 month (5 years 9 months); age range: 19-38 years) and 17 typically developing individuals (eight males, nine females; mean age: 31 years 4 months (6 years 2 months); age range: 20-40 years) were collected before, immediately after, and 1 hour after 45 minutes of frame running or running respectively. Independent t-tests were used to compare heart rate, level of exertion, and baseline cell proportions between groups. Mixed model analysis of variance was utilized to investigate immune cell responses to exercise across groups. RESULTS: Baseline levels of gamma delta (TCRγδ+) T-cells were significantly higher (absolute percentage: +2.65, p = 0.028) in the individuals with CP. Several cell populations showed similar significant changes after exercise in both CP and typically developing groups. Cytotoxic (CD8+) T-cells were only significantly elevated immediately after exercise in the typically developing participants (p < 0.01). Individuals with CP exhibited significantly lower heart rates (-11.1%, p < 0.01), despite similar ratings of perceived exertion. INTERPRETATION: Elevated baseline TCRγδ+ T-cells may indicate low-grade inflammation in adults with CP. Although most of the cell populations showed typical responses to endurance exercise, the absence of response in CD8+ T-cells in individuals with CP may indicate the need for higher intensity during exercise. WHAT THIS PAPER ADDS: TCRγδ+ T-cell baseline levels are elevated in adults with cerebral palsy (CP). The CD8+ T-cell response to exercise was blunted in adults with CP. Exercise intensity is decisive for CD8+ T-cell responses in individuals with CP.

The effect of neuromuscular electrical stimulation on the human skeletal muscle transcriptome.

AIM: The influence on acute skeletal muscle transcriptomics of neuromuscular electrical stimulation (NMES), as compared to established exercises, is poorly understood. We aimed to investigate the effects on global mRNA-expression in the quadriceps muscle early after a single NMES-session, compared to the effects of voluntary knee extension exercise (EX), and to explore the discomfort level. METHODS: Global vastus lateralis muscle gene expression was assessed (RNA-sequencing) in 30 healthy participants, before and 3 h after a 30-min session of NMES and/or EX. The NMES-treatment was applied using textile electrodes integrated in pants and set to 20% of each participant's pre-tested MVC mean (±SD) 200 (±80) Nm. Discomfort was assessed using Visual Analogue Scale (VAS, 0-10). The EX-protocol was performed at 80% of 1-repetition-maximum. RESULTS: NMES at 20% of MVC resulted in VAS below 4 and induced 4448 differentially expressed genes (DEGs) with 80%-overlap of the 2571 DEGs of EX. Genes well-known to be up-regulated following exercise, for example, PPARGC1A, ABRA, VEGFA, and GDNF, were also up-regulated by NMES. Gene set enrichment analysis demonstrated many common pathways after EX and NMES. Also, some pathways were exclusive to either EX, for example, muscle tissue proliferation, or to NMES, for example, neurite outgrowth and connective tissue proliferation. CONCLUSION: A 30-min NMES-session at 20% of MVC with NMES-pants, which can be applied with an acceptable level of discomfort, induces over 4000 DEGs, of which 80%-overlap with DEGs of EX. NMES can induce exercise-like molecular effects, that potentially can lead to health and performance benefits in individuals who are unable to perform resistance exercise.

Molecular profiling of high-level athlete skeletal muscle after acute endurance or resistance exercise - A systems biology approach.

OBJECTIVE: Long-term high-level exercise training leads to improvements in physical performance and multi-tissue adaptation following changes in molecular pathways. While skeletal muscle baseline differences between exercise-trained and untrained individuals have been previously investigated, it remains unclear how training history influences human multi-omics responses to acute exercise. METHODS: We recruited and extensively characterized 24 individuals categorized as endurance athletes with >15 years of training history, strength athletes or control subjects. Timeseries skeletal muscle biopsies were taken from M. vastus lateralis at three time-points after endurance or resistance exercise was performed and multi-omics molecular analysis performed. RESULTS: Our analyses revealed distinct activation differences of molecular processes such as fatty- and amino acid metabolism and transcription factors such as HIF1A and the MYF-family. We show that endurance athletes have an increased abundance of carnitine-derivates while strength athletes increase specific phospholipid metabolites compared to control subjects. Additionally, for the first time, we show the metabolite sorbitol to be substantially increased with acute exercise. On transcriptional level, we show that acute resistance exercise stimulates more gene expression than acute endurance exercise. This follows a specific pattern, with endurance athletes uniquely down-regulating pathways related to mitochondria, translation and ribosomes. Finally, both forms of exercise training specialize in diverging transcriptional directions, differentiating themselves from the transcriptome of the untrained control group. CONCLUSIONS: We identify a "transcriptional specialization effect" by transcriptional narrowing and intensification, and molecular specialization effects on metabolomic level Additionally, we performed multi-omics network and cluster analysis, providing a novel resource of skeletal muscle transcriptomic and metabolomic profiling in highly trained and untrained individuals.

Remodeling of the human skeletal muscle proteome found after long-term endurance training but not after strength training.

Exercise training has tremendous systemic tissue-specific health benefits, but the molecular adaptations to long-term exercise training are not completely understood. We investigated the skeletal muscle proteome of highly endurance-trained, strength-trained, and untrained individuals and performed exercise- and sex-specific analyses. Of the 6,000+ proteins identified, >650 were differentially expressed in endurance-trained individuals compared with controls. Strikingly, 92% of the shared proteins with higher expression in both the male and female endurance groups were known mitochondrial. In contrast to the findings in endurance-trained individuals, minimal differences were found in strength-trained individuals and between females and males. Lastly, a co-expression network and comparative literature analysis revealed key proteins and pathways related to the health benefits of exercise, which were primarily related to differences in mitochondrial proteins. This network is available as an interactive database resource where investigators can correlate clinical data with global gene and protein expression data for hypothesis generation.

FiNuTyper: Design and validation of an automated deep learning-based platform for simultaneous fiber and nucleus type analysis in human skeletal muscle.

AIM: While manual quantification is still considered the gold standard for skeletal muscle histological analysis, it is time-consuming and prone to investigator bias. To address this challenge, we assembled an automated image analysis pipeline, FiNuTyper (Fiber and Nucleus Typer). METHODS: We integrated recently developed deep learning-based image segmentation methods, optimized for unbiased evaluation of fresh and postmortem human skeletal muscle, and utilized SERCA1 and SERCA2 as type-specific myonucleus and myofiber markers after validating them against the traditional use of MyHC isoforms. RESULTS: Parameters including cross-sectional area, myonuclei per fiber, myonuclear domain, central myonuclei per fiber, and grouped myofiber ratio were determined in a fiber-type-specific manner, revealing that a large degree of sex- and muscle-related heterogeneity could be detected using the pipeline. Our platform was also tested on pathological muscle tissue (ALS and IBM) and adapted for the detection of other resident cell types (leucocytes, satellite cells, capillary endothelium). CONCLUSION: In summary, we present an automated image analysis tool for the simultaneous quantification of myofiber and myonuclear types, to characterize the composition and structure of healthy and diseased human skeletal muscle.

MRI characterization of skeletal muscle size and fatty infiltration in long-term trained and untrained individuals.

This study investigated body composition measures in highly trained and untrained individuals using whole-body magnetic resonance imaging (MRI). Additionally, correlations between these measures and skeletal muscle gene expression were performed. Thirty-six individuals were included: endurance-trained males (ME, n = 8) and females (FE, n = 7), strength-trained males (MS, n = 7), and untrained control males (MC, n = 8) and females (FC, n = 6). MRI scans were performed, and resting M. vastus lateralis (VL) biopsies were subjected to RNA sequencing. Liver fat fraction, visceral adipose tissue volume (VAT), total body fat, and total lean tissue were measured from MRI data. Additionally, cross-sectional area (CSA) and fat signal fraction (FSF) were calculated from Mm. pectoralis, M. erector spinae and M. multifidus combined, Mm. quadriceps, and Mm. triceps surae (TS). Liver fat fraction, VAT, and total body fat relative to body weight were lower in ME and FE compared with corresponding controls. MS had a larger CSA across all four muscle groups and lower FSF in all muscles apart from TS compared with MC. ME had a lower FSF across all muscle groups and a larger CSA in all muscles except TS than MC. FE athletes showed a higher CSA in Mm. pectoralis and Mm. quadriceps and a lower CSA in TS than FC with no CSA differences found in the back muscles investigated. Surprisingly, the only difference in FSF between FE and FC was found in Mm. pectoralis. Lastly, correlations between VL gene expression and VL CSA as well as FSF showed that genes positively correlated with CSA revealed an enrichment of the oxidative phosphorylation and thermogenesis pathways, while the genes positively correlated with FSF showed significant enrichment of the spliceosome pathway. Although limited differences were found with training in females, our study suggests that both regular endurance and resistance training are useful in maintaining muscle mass, reducing adipose tissue deposits, and reducing muscle fat content in males.

Skeletal Muscle Transcriptomic Comparison between Long-Term Trained and Untrained Men and Women.

To better understand the health benefits of lifelong exercise in humans, we conduct global skeletal muscle transcriptomic analyses of long-term endurance- (9 men, 9 women) and strength-trained (7 men) humans compared with age-matched untrained controls (7 men, 8 women). Transcriptomic analysis, Gene Ontology, and genome-scale metabolic modeling demonstrate changes in pathways related to the prevention of metabolic diseases, particularly with endurance training. Our data also show prominent sex differences between controls and that these differences are reduced with endurance training. Additionally, we compare our data with studies examining muscle gene expression before and after a months-long training period in individuals with metabolic diseases. This analysis reveals that training shifts gene expression in individuals with impaired metabolism to become more similar to our endurance-trained group. Overall, our data provide an extensive examination of the accumulated transcriptional changes that occur with decades-long training and identify important "exercise-responsive" genes that could attenuate metabolic disease.

Exercise Induces Different Molecular Responses in Trained and Untrained Human Muscle.

INTRODUCTION: Human skeletal muscle is thought to have heightened sensitivity to exercise stimulus when it has been previously trained (i.e., it possesses "muscle memory"). We investigated whether basal and acute resistance exercise-induced gene expression and cell signaling events are influenced by previous strength training history. METHODS: Accordingly, 19 training naïve women and men completed 10 wk of unilateral leg strength training, followed by 20 wk of detraining. Subsequently, an acute resistance exercise session was performed for both legs, with vastus lateralis biopsies taken at rest and 1 h after exercise in both legs (memory and control). RESULTS: The phosphorylation of AMPK and eEF2 was higher in the memory leg than that in the control leg at both time points. The postexercise phosphorylation of 4E-BP1 was higher in the memory leg than that in the control leg. The memory leg had lower basal mRNA levels of total PGC1α and, unlike the control leg, exhibited increases in PGC1α-ex1a transcripts after exercise. In the genes related to myogenesis (SETD3, MYOD1, and MYOG), mRNA levels differed between the memory and the untrained leg; these effects were evident primarily in the male subjects. Expression of the novel gene SPRYD7 was lower in the memory leg at rest and decreased after exercise only in the control leg, but SPRYD7 protein levels were higher in the memory leg. CONCLUSION: In conclusion, several key regulatory genes and proteins involved in muscular adaptations to resistance exercise are influenced by previous training history. Although the relevance and mechanistic explanation for these findings need further investigation, they support the view of a molecular muscle memory in response to training.

Expression of striated activator of rho-signaling in human skeletal muscle following acute exercise and long-term training.

The striated activator of rho-signaling (STARS) protein acts as a link between external stimuli and exercise adaptation such as muscle hypertrophy. However, the acute and long-term adaptational response of STARS is still unclear. This study aimed at investigating the acute and long-term endurance training response on the mRNA and protein expression of STARS and its related upstream and downstream factors in human skeletal muscle. mRNA and protein levels of STARS and related factors were assessed in skeletal muscle of healthy young men and women following an acute bout of endurance exercise (n = 15) or 12 weeks of one-legged training (n = 23). Muscle biopsies were obtained before (acute and long-term), at 30 min, 2, and 6 h following acute exercise, and at 24 h following both acute exercise and long-term training. Following acute exercise, STARS mRNA was significantly elevated 3.9-fold at 30 min returning back to baseline 24 h after exercise. STARS protein levels were numerically but nonsignificantly increased 7.2-fold at 24 h. No changes in STARS or ERRα mRNA or STARS protein expression were seen following long-term training. PGC-1α mRNA increased 1.7-fold following long-term training. MRTF-A mRNA was increased both following acute exercise and long-term training, in contrast to SRF mRNA and protein which did not change. STARS mRNA is acutely upregulated with exercise, but there is no cumulative effect to long-term training as seen in PGC-1α mRNA expression. Exercise intensity might play a role in manifestation of protein expression, suggesting a more complex regulation of STARS.