脐带间充质干细胞来源外泌体的规模化制备及鼻腔递送研究OA

OBJECTIVE To investigate a scalable preparation method for exosomes derived from human umbilical cord-derived mesenchymal stem cells(hUC-MSCs),and to observe the local dynamic distribution of exosomes following intranasal delivery.METHODS Human umbilical cord mesenchymal stem cells(hUC-MSCs)were isolated and cultured using the tissue explant method and expanded in a serum-free medium.The cell culture supernatant was collected,and exosomes were isolated and purified via traditional ultracentrifugation and ultrafast-isolation system,respectively.The products were characterized by transmission electron microscopy(TEM),nanoflow cytometry,nanoparticle tracking analysis(NTA),and Western blotting.To evaluate distribution,exosomes were labeled with DiR fluorescent dye(DiR-exo)and administered intranasally to mice.An in vivo imaging system was used to observe the distribution of DiR-exo in the nasal cavity at different time points.RESULTS We successfully established a serum-free culture system for hUC-MSCs.Exosomes isolated by both methods displayed typical bilayer vesicular structures and expressed CD9 and TSG101 without Calnexin expression.In vivo imaging revealed persistent fluorescent signals in the nasal cavities of mice after intranasal delivery,which remained clearly detectable up to 6 hours post-administration.CONCLUSION High-yield preparation of hUC-MSC-derived exosomes was realized using ultrafast-isolation system.Following intranasal delivery,fluorescently labeled exosomes showed stable retention in the mouse nasal cavity,providing a technical reference for the future development and clinical translation of exosome-based formulations.