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Injecting human mesenchymal stem cells (hMSCs) at wound sites is known to have a therapeutic effect; however, hMSCs have several limitations, such as low viability and poor engraftment after injection, as well as a potential risk of oncogenesis. The use of a conditioned medium (CM) was suggested as an alternative method for treating various wounds instead of direct hMSC administration. In addition to not having the adverse effects associated with hMSCs, a CM can be easily mass produced and can be stored for long-term, thereby making it useful for clinical applications. In general, a CM is collected from hMSCs with low passage number; whereas, the hMSCs with high passage number are usually discarded because of their low therapeutic efficacy as a result of reduced angiogenic factor secretion. Herein, we used a CM collected from high passage number (passage 12, P12) hMSCs treated with gold-iron nanoparticles (AuFe NPs). Our AuFe NPs were designed to release the iron ion intracellularly via endocytosis. Endosomes with low pH can dissolve iron from AuFe NPs, and thus, the intracellularly released iron ions up-regulate the hypoxia-inducible factor 1α and vascular endothelial growth factor (VEGF) expression. Through this mechanism, AuFe NPs improve the amount of VEGF expression from P12 hMSCs so that it is comparable to the amount of VEGF expression from low passage number (passage 6, P6), without treatment. Furthermore, we injected the CM retrieved from P12 MSCs treated with AuFe NPs in the mouse skin wound model (AuFe P12 group). AuFe P12 group revealed significantly enhanced angiogenesis in the mouse skin wound model compared to the high passage hMSC CM-injected group. Moreover, the result from the AuFe P12 group was similar to that of the low passage hMSC CM-injected group. Both the AuFe P12 group and low passage hMSC CM-injected group presented significantly enhanced re-epithelization, angiogenesis, and tissue remodeling compared to the high passage hMSC CM-injected group. This study reveals a new strategy for tissue regeneration based on CM injection without considering the high cell passage count.

international journal of molecular sciences

Enhancing the Wound Healing Effect of Conditioned Medium Collected from Mesenchymal Stem Cells with High Passage Number Using Bioreducible Nanoparticles