Extracellular vesicles (EVs) function as a natural communication system, enabling transfer of biomolecules between cells. Although EVs are produced by nearly all cell types and have shown promise as therapeutic agents, our understanding of how EVs from different cellular origins recognize recipient cells, are taken up, and processed intracellularly remains limited. In particular, it remains unclear whether intracellular EV behavior is primarily governed by EV-intrinsic properties or by the recipient-cell environment. Here, we systematically investigate the relative contributions of EV origin and recipient-cell identity to intracellular EV trafficking. By combining single-vesicle tracking with machine learning-based diffusion-state classification, we analyzed EVs derived from four distinct sources across three recipient cell types over three different time spans. This approach enabled characterization of dynamic transport behaviors, overcoming limitations of traditional ensemble-averaging methods. We observed recipient cell-dependent differences in intracellular trafficking patterns, ranging from relatively uniform and dynamic motion to more heterogeneous and confined behaviors, reflecting variability in intracellular transport environments. Despite these differences, EV origin had only a modest influence on intracellular dynamics following uptake. These findings suggest a model in which intracellular transport arises from an interplay between general EV-associated properties and cell-specific environment.
Brochner, B. H., Song, P., Yan, Y., Weissinger, H., Sorensen, E. A., Kjems, J., Malle, M. G.
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