The Study Of The Effect And Molecular Mechanism About Exosomes On Tissue Repair

BACKGROUND: Exosomes(Exos),which are one kind of extracellular vesicles(EVs)secreted from cells,have similar effects as their source cells in promoting tissue regeneration via mediating intercellular communication even cross species.Currently,there is no report using the exosomes derived from synovial mesenchymal stem cell(SMSC-Exos)and platelet-rich plasm(PRP-Exos)in preventing osteonecrosis of femoral head(ONFH),or using PRP-Exos in promoting the healing process of chronic wound.OBJECTIVE: This study is aimed at evaluating the effects and molecular mechanism of the SMSC-Exos and PRP-Exos on preventing ONFH and the effects and molecular mechanism of PRP-Exos on promoting healing process of the chonic wound.METHODS: SMSCs was isolated from human synovial membrane and PRP was isolated from blood.Then SMSC-Exos and PRP-Exos were isolated,purified and identified.The effects of exosomes on preventing ONFH and promoting healing process of chronic wound were evaluated by in vivo experiments.And the effects of exosomes on the cell of proliferation,migration,osteogenesis,angiogenesis and antiapoptosis were evaluated by series of in vitro experiments.Furthermore,the osteogenicrelated pathway,angiogenesis-related pathway and apoptosis-related pathway were evaluated in vitro for in-depth studying.RESULTS:(1)SMSCs was isolated from human synovial membrane and PRP was extracted from blood successfully.The SMSC-Exos and PRP-Exos were both isolated and the animal models were built.(2)Through transmission electron microscopy,DLS and analysis of exosomal surface markers,SMSC-Exos and PRP-Exos were identified.(3)SMSC-Exos and PRP-Exos were proven to be effective in preventing ONFH.PRP-Exos were proven to be effective in promoting healing process of the chronic wound.In vitro,Exos could be uptake and integrated by cells,proven by using DiL labelling.The proliferation assays indicated that Exos could promote cell proliferation and block the glucocorticoid(GC)-induced inhibition of proliferation.The results of Annexin V assay and caspase-3 detection suggested that Exos cound block apoptosis induced by GC.The results of vessel formation assay showed that Exos could promote angiogenesis of endothelial cells and block the GC-induced inhibition of angiogenesis.The results of osteogenic assay indicated that PRP-Exos could block the GC-induced inhibitioin of osteogenesis.And in vivo,the results of Ki67 and TUNEL assays showed that Exos rescued the GC-induced inhibition of cell proliferation and apoptosis in accordance with the vitro results.The morphology,pathology and vessel number of femoral head were observed by histological and radiographic technique.(4)The molecular mechanism of PRP-Exos in preventing ONFH and promoting healing of chronic wound were proved as follows.The phosphorylation level of Akt and Erk were evaluated and indicated that PRP-Exos promote vessel formation through Akt and Erk pathway.The protein expression level of Runx2,Type I Collagen and ?-catenin were examined and indicated that PRP-Exos rescued the GC-induced inhibition of osteogenesis by maintaining the stability of ?-catenin.PRP-Exos rescued the GCinduced apoptosis through modulating Bcl-2 expression.And PRP-Exos promoted reepithelization via YAP activation.CONCLUSIONS:(1)SMSC-Exos and PRP-Exos could prevent ONFH,and PRPExos could promote the healing of chronic wound.(2)PRP-Exos performed the regenerative and protective function through regulating Akt,Erk,?-catenin and Bcl-2.(3)PRP-Exos promoted re-epithelization via YAP activation.