Epigenetic Modulation of the ACE System Underlies the Slow Aerobic Muscle Phenotype and Metabolic Exercise Response.

ACE promoter methylation in skeletal muscle was higher than in blood and increased with physical activity levels.

• ACE promoter methylation doubled with heart rate-elevating physical activity exceeding 6 hours weekly (p < 0.001, η² = 0.090)
• ACE promoter methylation also doubled with endurance exercise (p < 0.001, η² = 0.059)
• Muscle ACE promoter methylation was lowest in inactive, aerobically unfit ACE I-allele carriers
• Mean ACE promoter methylation across participants was 0.37 ± 0.18 across 23 CpG sites
• Muscle methylation was compared to capillary blood methylation in a second group of 24 VO2max- and genotype-matched male subjects

ACE promoter methylation was inversely correlated with ACE activity, angiotensin 2 levels, and post-exercise ACE mRNA expression.

• Inverse correlation between ACE promoter methylation and ACE activity: r = -0.406
• Inverse correlation between ACE promoter methylation and angiotensin 2: r = -0.447
• Inverse correlation between ACE promoter methylation and post-exercise ACE mRNA expression: r = -0.745
• These correlations suggest that higher methylation suppresses ACE transcriptional and enzymatic activity

Endurance exercise induced significant increases in blood angiotensin 2 levels.

• Blood angiotensin 2 rose from 18.1 pg/mL at rest to 45.6 pg/mL following exercise
• This rise was part of significant metabolic shifts induced by exercise
• Angiotensin 2 levels were inversely correlated with ACE promoter methylation (r = -0.447)

The ACE I/D gene polymorphism interacted with aerobic fitness state to influence ACE system regulation.

• Interaction between ACE I/D polymorphism and aerobic fitness state was statistically significant (p = 0.046, η² = 0.297)
• Low promoter methylation exerted pronounced effects specifically in ACE-II homozygotes
• Low ACE-promoter methylation may override I-allele-driven transcriptional silencing

Regression models incorporating ACE regulatory parameters significantly explained variance in slow-twitch muscle fiber characteristics and metabolic markers.

• Models significantly explained variance in slow-twitch fiber percentage and cross-sectional area, mitochondrial volume density, and key lipid/glucose metabolites
• Mean r = 0.493 across regression models with statistical power > 0.8
• Seven metabolites including acetyl-CoA, adenosine monophosphate, and phosphocreatine displayed opposing associations with ACE genotype and methylation, indicating divergent regulatory pathways

The study sample exhibited wide interindividual variability in aerobic capacity and ACE promoter methylation.

• Participants were 114 healthy, physically diverse white Caucasian men (age: 29.5 ± 8.1 years; weight: 76.9 ± 14.5 kg)
• VO₂max ranged from 2.1 to 5.4 L·min⁻¹
• ACE promoter methylation ranged broadly with a mean of 0.37 ± 0.18 across 23 CpG sites
• Molecular endpoints included ACE genotype, promoter methylation, mRNA expression, enzyme activity, angiotensin 2 levels, and markers of mitochondrial, glycolytic, and lipid metabolism

Network analysis identified ACE promoter methylation as a contributor to the slow aerobic muscle phenotype integrated within a broader ACE regulatory network.

• Statistical analyses comprised Pearson correlations, regression modeling (α = 0.05), network analysis, and ANOVA
• ACE promoter methylation was integrated with other regulatory indexes of the ACE system to explain the slow aerobic muscle phenotype
• The regulatory network linked methylation to mitochondrial volume density, slow-twitch fiber composition, and lipid/glucose metabolites