P78 | THE ROLE OF EXTRACELLULAR VESICLES DERIVED FROM MESENCHYMAL STROMAL CELLS IN MYOFIBER REGENERATION

A. Longhin¹, V. Gatta², G. Teti², C. Sassoli³, F. Chellini³, A. Tani³, M. Parigi³, R. Garella³, F. Palmieri³, R. Squecco³, M. Mattioli Belmonte⁴, C. Licini⁴, A. La Contana⁴, S. Zecchi Orlandini³, M. Falconi¹ | ¹Dep. Medical and Surgical Sciences, University of Bologna, Bologna, Italy; ²Dep. Biomedical and Neuromotor Sciences, University of Bologna, Bologna, Italy; ³Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy; ⁴Department of Clinical and Molecular Sciences, Università Politecnica delle Marche, Ancona, Italy

Traumatic injuries can severely impair skeletal muscle, leading to limited regeneration, muscle loss, fibrosis, and compromised function. Despite the exploration of various therapeutic approaches, no fully effective treatment has yet been established. Mesenchymal stem cells (MSCs) have emerged as a promising strategy for muscle repair, largely due to their paracrine signaling and immunomodulatory properties mediated by extracellular vesicles (EVs). Although the cellular and molecular processes involved in skeletal muscle regeneration are well understood, the specific role of EVs in mediating intercellular communication during myofiber repair remains under active investigation. This study aims to clarify the contribution of MSC-derived EVs to muscle repair and regeneration. EVs were isolated from murine myoblasts using a polyethylene glycol (PEG)-based liquid exchange precipitation protocol. EVs from control myoblasts, differentiated myoblasts, and MSCs were characterized in terms of size, surface markers, and myokine content using electron microscopy, western blotting, and ProQuantum immunoassays. These EVs were subsequently added to cultures of damaged, differentiated myoblasts to assess their regenerative potential. Myotube formation was evaluated via inverted light microscopy, while the expression of key myogenic markers was assessed through western blot analysis. The results confirmed the presence of EVs in both myoblasts and MSCs. The expression of EV markers varied depending on the experimental conditions used to simulate muscle damage. Injured myoblasts released EVs containing inflammatory factors, whereas MSC-derived EVs attenuated the inflammatory environment, enhanced myogenic repair, and promoted muscle regeneration. In conclusion, our findings highlight the critical role of MSC-derived extracellular vesicles in regulating myogenic differentiation and supporting the regeneration of skeletal muscle following injury.
Funding: PRIN 2022 programme (20222P2NAJ; CUP I53D23003030006) (funded by the European Union - Next Generation EU)