Ultramarathon Race on Circulating Extracellular Vesicles Release and their Potential Role as Exercise Biomarkers

Introduction: Regular physical exercise promotes systemic adaptations which enhance skeletal muscle health, prevent atrophy, and promote regeneration, a highly coordinated process involving inflammation, regeneration and remodeling. Extracellular vesicles (EVs) have recently emerged as key mediators of the inflammatory processes associated with exercise, essential for initiating tissue repair. EVs activate resident and circulating immune cells, mostly macrophages, at the injury site by transferring inflammatory cytokines and signaling molecules, thus modulating the immune response. Moreover, EVs are also involved in the activation of satellite cells, essential for muscle repair and remodeling, by delivering signals that promote their proliferation and differentiation, thereby enhancing muscle regeneration. In this context, the present study aims to investigate the modulation of circulating EVs in response to high intensity and prolonged exercise in order to predict or detect systemic exercise adaptations. Methods: Serum samples of 19 ultramarathoners were collected (pre- and post-race) and processed by Size Exclusion Chromatography (SEC). SEC fractions were characterized with BCA assay for total protein content, Nanoparticle Tracking analysis (NTA) for particle counting and size distribution, TEM for EV imaging and Dot blot assay for EV markers detection. Results: EV size and concentration resulted comparable with those reported in literature and EVs showed structural integrity with no morphological differences between pre- and post-race. In addition, there is a significant post-race increase in the expression of tetraspanin CD9, muscle related marker CAV3 and stress-related mitochondrial marker HSP60. Conclusion: The obtained results suggest that post-exercise adaptations may promote a shift of the EV phenotype towards an increase in CD9-enriched EVs. Moreover, the increase of CAV3 and HSP60 expression indicates that circulating EVs might mediate muscle remodeling in response to the metabolic stress of high-intensity, prolonged exercise. EVs could therefore represent novel exercise biomarkers that could lead to personalized treatments for muscle injuries and degenerative conditions.