Potential upscaling protocol establishment and wound healing bioactivity screening of exosomes isolated from canine adipose-derived mesenchymal stem cells

Mesenchymal stem cell-derived exosomes exhibit promising potential in tissue regeneration. Recent studies highlight its significant therapeutic potential in various stages of wound healing. However, the clinical translation of exosome-based therapy was hindered due to issues regarding low productivity and the lack of efficient production protocol to obtain a clinically relevant exosome quantity. Therefore, this study established a potential upscaling protocol to produce exosomes derived from canine adipose-derived mesenchymal stem cells (cAD-MSCs) and explored its potential for wound treatment. The potential upscaling protocol, termed VSCBIC-3-3D, was carried out using VSCBIC-3 in-house serum-free exosome-collecting solution in a three-dimensional (3D) culture system followed by the tangential flow filtration (TFF) isolation. Our findings suggest that culturing cAD-MSCs with VSCBIC-3 maintained cell morphology and viability. Compared to conventional two-dimensional (2D) protocols, The potential upscaling protocol increased exosome yield and concentration in conditioned medium by 2.4-fold and 3.2-fold, respectively. The quality assessment revealed enhanced purity and bioactivity of exosomes produced using the VSCBIC-3-3D protocol. In addition, the cAD-MSCs-derived exosomes were shown to significantly improve fibroblast migration, proliferation, and wound healing-related gene expression in vitro. This study collectively demonstrates that potential upscaling protocol establishment allowed robust production of exosomes from cAD-MSCs, which exhibit therapeutic potential for wound healing in vitro.