Harnessing hydrodynamics for high-yield production of extracellular vesicles from stem cells spheroids with specific cargo profiling
Lenoir, S.; Thouvenot, E.; Gropplero, G.; Dec, L.; Loew, D.; Thery, C.; Perez, J. E.; Wilhelm, C.
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This study presents a novel method and device for the hydrodynamic production of extracellular vesicles (EVs) derived from biomimetic multicellular 3D spheroids, enabling high-throughput particle release that is 10 to 20 times higher than in non-stimulated conditions. The device facilitates the formation of spheroids from human mesenchymal stem cells (hMSCs), offering an all-in-one approach for both spheroid generation and EV release. Production times are reduced to just a few hours, with yield further increased by alternating periods of high hydrodynamic flow and spheroid recovery in a sequential production approach. Using this system, we explored the impact of hydrodynamic and starvation conditions on the protein cargo of EVs, identifying distinct protein markers through proteomics. Specifically, hydrodynamic stimulation enriched EVs in plasma membrane-derived and mitochondrial proteins, revealing divergent biogenesis pathways. Importantly, the produced EVs exhibited therapeutic properties, with demonstrated effects in wound healing, angiogenesis, and anti-inflammatory responses, some showing enhanced efficacy under hydrodynamic stimulation.
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