Extracellular Vesicles From Mesenchymal Stromal Cells Drive Muscle and Neuronal Regeneration Through TNFα Modulation

Muscle defects caused by accidents, tumour resection and congenital malformations affect paediatric and adult patients. In this context, the neural-muscle regeneration potential of mesenchymal stromal/stem cells derived from extracellular vesicles (EV) has been demonstrated by our group and others, but the mechanism by which EVs act remains unknown. This work aimed to investigate the neural-muscle regeneration mechanism shown by EVs in vivo using three-dimensional (3D) multicellular in vitro models. We used (1) human muscle decellularised tissue (ECM) engineered with human muscle precursor cells (hMPC) together with macrophages THP-1 (M0) and (2) organotypic spinal cord from rat foetuses. We also studied neuroinflammation in 2D with primary microglia cells stimulated with lipopolysaccharide (LPS). Samples treated with good manufacturing practices (GMP)-grade EVs were assessed, combining functional analyses, protein and gene expression. In the functional muscle model, EVs protect the cells from death after damage, decreasing cCAS3 and stimulating cell proliferation. The protein array and gene results highlighted that EVs act through the downregulation of the TNFα factor. In parallel, in both neuroinflammation-induced microglia and organotypic spinal cord-damaged models, EVs regulated the neuroinflammation by inhibiting TNFα and promoting neural axon sprouting. In summary, EVs guard great potential for tissue regeneration by TNFα modulation, promoting muscle-neural regeneration.