Load-responsive release of mesenchymal stromal cell extracellular vesicles from polycaprolactone scaffolds

Abstract Background Extracellular vesicles (EVs) are increasingly recognised as potent acellular mediators of tissue regeneration; however, strategies that enable their controlled and sustained delivery from biomaterial scaffolds remain limited. This study aimed to evaluate the load-responsive release of mesenchymal stromal cell-derived EVs from architecturally distinct polycaprolactone (PCL) scaffolds under in vitro mechanical stimulation for cartilage tissue regeneration applications. Results Electrowritten (1EW, aligned) and melt electrospun (1ME, random) PCL scaffolds were plasma-treated, coated with polydopamine (PDA), functionalised with carboxyfluorescein diacetate (CFDA)-labelled EVs, and subjected to cyclic compressive loading for up to 7 days. EV release into surrounding dPBS was quantified by flow cytometry based on particle concentration, fluorescence intensity, and tetraspanin expression. Scaffolds were also visualised by confocal microscopy. After 1 day of loading, both scaffold types showed increased EV release compared with unloaded controls, however, the load-to-control release ratio was significantly higher for 1EW scaffolds. After 7 days, 1EW scaffolds exhibited sustained and significantly elevated EV release, as evidenced by increased particle counts, whereas 1ME scaffolds showed a weaker response. Released particles from all groups expressed canonical EV tetraspanins, supporting their EV-associated identity. Moreover, chondrocytes cultured on 1EW scaffolds for 7 days remained viable, supporting scaffold cytocompatibility. Conclusions This study demonstrates the feasibility of a load-responsive EV delivery strategy using PCL scaffolds. The aligned 1EW scaffold exhibited greater EV release than the 1ME scaffold under dynamic mechanical loading and demonstrated good cytocompatibility, supporting its further development for cartilage tissue engineering.