Optimizing mesenchymal stromal cells priming strategies for tailored effects on the secretome.

Mesenchymal stromal cells (MSCs) have emerged as a promising tool in regenerative medicine, with recent focus shifting towards their secretome as a cell-free therapeutic approach. This study investigates the impact of various priming strategies on the immunomodulatory, anti-inflammatory, and regenerative potential of adipose-derived MSCs (ASCs) secretomes. We evaluated the effects of hypoxia, pro-inflammatory cytokines, and spheroid culture conditions on ASC secretome composition and functionality. Gene expression analysis, nanoparticle tracking, protein quantification, and functional assays were performed to characterize the secretomes. RNA sequencing revealed significant differences in gene expression profiles across priming conditions, particularly in pathways related to osteogenesis, angiogenesis, inflammation, and neurotrophic factors. Notably, spheroid culture combined with hypoxia and inflammation resulted in a substantial increase in extracellular vesicle production and altered protein content. Functional assays demonstrated enhanced neutrophil inhibition by secretomes from hypoxia-primed ASCs. Our findings indicate that tailored priming strategies can significantly modulate the therapeutic properties of ASC secretomes, potentially enhancing their efficacy in various clinical applications. This study provides valuable insights for optimizing cell-free therapies in regenerative medicine and offers a basis for developing more targeted and effective treatments.