The role of biomechanical stress in extracellular vesicle formation, composition and activity.

Extracellular vesicles (EVs) are cornerstones of intercellular communication with exciting fundamental, clinical, and more broadly biotechnological applications. However, variability in EV composition, which results from the culture conditions used to generate the EVs, poses significant fundamental and applied challenges and a hurdle for scalable bioprocessing. Thus, an understanding of the relationship between EV production (and for clinical applications, manufacturing) and EV composition is increasingly recognized as important and necessary. While chemical stimulation and culture conditions such as cell density are known to influence EV biology, the impact of biomechanical forces on the generation, properties, and biological activity of EVs remains poorly understood. Given the omnipresence of these forces in EV preparation and in biomanufacturing, expanding the understanding of their impact on EV composition-and thus, activity-is vital. Although several publications have examined EV preparation and bioprocessing and briefly discussed biomechanical stresses as variables of interest, this review represents the first comprehensive evaluation of the impact of such stresses on EV production, composition and biological activity. We review how EV biogenesis, cargo, efficacy, and uptake are uniquely affected by various types, magnitudes, and durations of biomechanical forces, identifying trends that emerge both generically and for individual cell types. We also describe implications for scalable bioprocessing, evaluating processes inherent in common EV production and isolation methods, and propose a path forward for rigorous EV quality control.