Priming strategies to enhance the therapeutic efficacy of mesenchymal stromal/stem cell-derived vesicles in regenerative medicine.

Mesenchymal stromal/stem cell-derived extracellular vesicles (MSC-EVs) have emerged as promising acellular therapeutics in regenerative medicine, offering a safer and more controllable alternative to whole-cell therapies. Their therapeutic efficacy, however, is highly dependent on their molecular cargo, which reflects the physiological state and environmental conditions of the parent MSCs. Priming of mesenchymal stromal/stem cells (MSCs) with defined stimuli such as hypoxia, inflammatory cytokines, 3D culture systems, biomaterials, or pharmacological agents has been increasingly employed to enhance extracellular vesicle (EV) bioactivity. These strategies modulate EV content, enriching vesicles with regenerative, immunomodulatory, angiogenic, and antioxidant factors. For instance, hypoxic priming activates hypoxia-inducible factor-1α-driven gene expression, promoting the packaging of angiogenic and anti-inflammatory molecules, while cytokine-based priming upregulates immunosuppressive proteins and regulatory microRNAs. Similarly, 3D culture mimics aspects of the native tissue microenvironment, augmenting the secretion of EVs with enhanced reparative potential. Emerging combination-based approaches synergize these effects, generating EVs with superior therapeutic profiles. Despite encouraging preclinical data, translation to clinical application is challenged by variability in MSC sources, priming conditions, and EV isolation methods. Standardization of protocols, validated potency assays, and regulatory harmonization are critical for clinical advancement. This mini-review summarizes current priming strategies, the underlying mechanisms influencing EV cargo, and their functional implications in disease models, while highlighting key barriers and future directions for the clinical translation of primed MSC-EV therapies.