3D mechanical stimulation modulates endothelial exosomes to promote fibroblast activation

Mechanical forces play a crucial role in regulating cellular communication during tissue repair; however, it remains unclear how mechanical stimulation modulates endothelial exosome secretion and its effects on fibroblast activation.

In this study, endothelial cells were incorporated into 3D bioprinted tissue-engineered dermal constructs and cultured under static or mechanically stretched conditions. Exosomes were isolated from these cells, characterised, and applied to human dermal fibroblasts to assess their influence on proliferation, migration, and extracellular matrix formation. Data-independent acquisition proteomics was performed to analyse exosomal protein cargo and associated signalling pathways.

Mechanical loading increased exosome secretion by approximately 2.5-fold without altering vesicle morphology. Functionally, mechanically stimulated exosomes enhanced fibroblast migration and type I collagen synthesis more significantly than controls. Proteomics profiling identified 4476 proteins in the exosomes, of which 677 were differentially expressed. Enrichment analysis revealed activation of the VEGF, HIF-1, Relaxin, and AGE-RAGE pathways, implicating roles in angiogenesis, metabolic regulation, and extracellular matrix remodelling.

These findings demonstrate that 3D mechanical stimulation not only augments the quantity of endothelial exosomes but also reshapes their molecular cargo, thereby enhancing biomechanical communication between endothelial cells and fibroblasts. Building on prior evidence that fibroblast-derived exosomes promote endothelial angiogenesis, we propose a bidirectional 'mechanical stimulation-exosome-communication-tissue reconstruction' loop, providing a theoretical foundation for optimising exosome-based strategies in skin tissue engineering.