Preparation of cell-derived vesicles from eukaryotic and prokaryotic origins for the delivery of biomolecules

Cell membrane-derived vesicles play essential roles in intercellular communication, material transport, and waste disposal. Despite their biomedical and industrial potential, isolating extracellular vesicles from natural sources remains technically challenging, limiting purification efficiency and scalability. This study introduces cell membrane extrusion as an alternative approach to optimize the production of cell membrane-derived vesicles (CSMs), from eukaryotic and prokaryotic cells. CSMs, generated from HeLa and SH-SY5Y cells exhibited a distinctive cup-shaped morphology and sizes of 151.36 ± 72.36 nm, and 416.86 ± 108.49 nm at 20 °C by DLS respectively, showing remarkable thermal stability at 4-70 °C range. Furthermore, loaded vesicles interacted with mammalian cells and achieved successful cargo internalization. CSMs were also produced from <i>E. coli</i> membranes, forming unilamellar vesicles of approximately 100 nm, as observed by Cryo-TEM. These vesicles displayed an inverse correlation between vesicle size and thermal stability and efficient cargo incorporation detected in 85% ± 3% of CSMs. However, under tested conditions, no interaction with prokaryotic cells occurred, and consequently, no delivery of the loaded molecule was observed. Overall, thesefindings highlight the potential of generating cell membrane-derived nanovesicles through extrusion, offering a promising strategy to mimic extracellular vesicles for innovative biomedical and industrial applications, including targeted drug delivery system.