Engineering Strategies for Enhanced Extracellular Vesicle Production: Mechanisms and Applications

Extracellular vesicles (EVs) have emerged as key mediators of intercellular communication and hold great promise for liquid biopsy and therapeutic applications. Their molecular composition and abundance dynamically reflect the pathophysiological state of their parental cells, endowing EVs with significant value for biomarker discovery and precision medicine. However, inefficient biogenesis, limited production yield, and considerable heterogeneity remain core bottlenecks to their clinical translation. This review systematically surveys recent engineering strategies aimed at enhancing EV production, encompassing diverse physical stimuli and biochemical interventions, with a particular focus on the underlying molecular mechanisms that govern EV quantity, composition, and functionality. Despite their varied modes of action, these approaches share a convergent regulatory principle-remodeling the fate of multivesicular bodies (MVBs) by promoting MVB biogenesis, intracellular trafficking, and plasma membrane fusion, while suppressing their diversion toward lysosomal degradation. Importantly, yield-enhancing strategies not only facilitate the scalable manufacturing of therapeutic EVs, but also enable the functional activation of EV-producing cells, enriching disease-relevant molecular signatures and directing the cargo composition for improved diagnostic performance. Crucially, the synergy between yield-enhancing strategies and advanced biosensing platforms is reshaping the entire EV research workflow. Emerging integrated analytical systems-encompassing microfluidics, electrochemical sensors, and acoustic detectors-now enable real-time monitoring of EV secretion kinetics, providing dynamic feedback to optimize stimulation parameters and harvest windows. Moreover, lab-on-a-chip platforms are beginning to merge EV production, in-line purification, and multiplexed detection into a single automated workflow, facilitating simultaneous quality control and functional characterization. Highly sensitive biosensing technologies further allow precise assessment of EV cargo composition, surface marker profiles, and biological activity, ensuring product consistency and potency. Looking forward, the deep integration of EV biogenesis engineering with these smart detection platforms holds the potential to overcome current translational bottlenecks, accelerating the development of scalable EV therapeutics and next-generation liquid biopsy systems for early diagnosis, dynamic disease monitoring, and precision medicine.