Background: Muscle regeneration is a highly coordinated process involving interactions between various cell types, including macrophages and muscle stem cells (satellite cells). Following injury, macrophages migrate to the damaged site and regulate satellite cell activation, proliferation, and differentiation into myotubes through direct contact or the release of soluble factors. Extracellular vesicles (EVs), tiny membrane-enclosed particles that transport regulatory molecules such as microRNA and growth factors, are key mediators of intercellular communication. It remains unclear whether macrophages influence satellite cell activities through EVs. A significant challenge in this field is that most EV research has been conducted using fixed tissues, limiting our understanding of the origin, destination, and real-time dynamics of EVs within living organisms. This study aims to investigate the role of macrophage-derived EVs in regulating satellite cell behaviour during muscle regeneration.
Results: To enable real-time observation of macrophage-satellite cell interactions via EVs, a zebrafish macrophage-specific EV reporter line was generated. Live imaging revealed that macrophage-derived EVs bind to and are internalised by satellite cells, subsequently inducing their division. Isolation and analysis of macrophage EVs identified Nampt, a pro-regenerative factor. The functional role of EVs in muscle regeneration was assessed using an EV inhibitor compound and a macrophage-specific genetic knockout, which impairs EV production and secretion. Zebrafish larvae lacking macrophage-derived EVs exhibited defective muscle regeneration, with the most pronounced effects observed during the early stages of regeneration. Further analysis showed a significant decrease in satellite cell proliferation and differentiation, while macrophage migration remained unaffected in the absence of EVs.
Conclusion: The macrophage-derived EV reporter line provides a valuable tool to study EV-mediated communication between macrophages and satellite cells in real time. This study demonstrates that macrophage-derived EVs are essential for promoting satellite cell proliferation and differentiation during muscle regeneration.