Epigenetic age deceleration reflects exercise-induced cardiorespiratory fitness improvements.

Epigenetic clocks are emerging as promising biomarkers of biological aging, yet their sensitivity to short-term interventions remains unclear. This pilot study investigates whether the GrimAge clock can capture the effects of a 6-month cycling-based endurance exercise training intervention, with cardiorespiratory fitness (VO2 max) and body composition as primary outcomes. We enrolled 42 adults aged 35-65, of whom 38 completed the study and 33 adhered to the protocol (> 66% adherence). Participants demonstrated significant improvements in VO2 max (+ 20%, P < 0.001) and body composition (P < 0.001). High-quality epigenetic data preprocessing yielded highly reproducible GrimAge estimates (< 2 months measurement error), which strongly correlated with chronological age (R2 = 0.86, P < 0.001). On average, GrimAge decreased by 7.44 months relative to the expected trajectory (P = 0.012), reflecting improvements in VO2 max (R2 = 0.27, P = 0.002) but not body composition changes. Notably, GrimAge changes strongly correlated with fluctuations in leukocyte composition, particularly neutrophil fraction (R2 = 0.74, P < 0.001). Adjusting for leukocyte composition improved consistency in GrimAge changes, aligning them with additional intervention outcomes and explaining up to 81% of variance. These findings demonstrate that GrimAge is responsive to short-term endurance training, serving as a meaningful biomarker of improved cardiorespiratory fitness, while also capturing immune system variability. This study supports the use of GrimAge in evaluating longevity interventions and highlights the importance of accounting for leukocyte composition in epigenetic aging research.