Exsomes From Human-bone-marrow-derived Mesenchymal Stem Cells Protect Against Renal Ischemia/Reperfusion Injury Via Transferring MiR-199a-3p

Background: Renal ischemia/reperfusion injury(IRI)is the main cause for acute kidney injury.Previous studies have found that bone marrow-derived mesenchymal stem cells(BMSCs)play a protective role in myocardial,brain,and kidney IRI,and confirmed that this protective effect is mediated by exosomes.Exosomes are one of extracellular vesicles with a diameter of about 40-200 nm,which can selectively encapsulate a variety of biomolecules,including microRNA(miRNA).A recent study has found that exosomes secreted from human BMSCs(hBMSC-Exos)are rich in miR-199a-3p which is reported to play an important regulatory role in the process of IRI.However,the effect of miR-199a-3p on renal IRI is unclear.Objectives: To explore the protective effect of hBMSC-Exos on renal IRI,and investigate the function of miR-199a-3p contained in hBMSC-Exos in the process of renal IRI,and finally elucidate its underlying mechanism.Methods: hBMSC-Exos were isolated by ultracentrifugation,and identified by transmission electron microscopy,nanoparticle trackinganalysis and western blot.Laser scanning confocal microscopy was used to observe hBMSC-Exos internalization in vitro and in vivo.An in vitro hypoxia/reoxygenation(H/R)model was estabulished by oxygen and glucose deprivation and reoxygenation(OGD/R),and the effect of hBMSC-Exos on renal IRI was investigated using flow cytometry and western blot.In vivo,the function of hBMSC-Exos was explored using H&E staining and immunohistochemistry.Real-time fluorescent quantitative polymerase chain reaction(qPCR)was used to detect the expression of miR-199a-3p in hBMSC-Exos.hBMSCs transfected with miR-199a-3p mimics or inhibitor using lipofectamine 2000 were used for flow cytometry and western blot to investigate the effect of exosomal miR-199a-3p on renal IRI and further explore its potential mechanism.Results: This study found that hBMSC-Exos can inhibit apoptosis induced by H/R in vitro.Meanwhile,we found that hBMSC-Exos can significantly promote the recovery of renal function in mice after IRI.In addition,we found that miR-199a-3p is highly expressed in hBMSC-Exos,and hBMSC-Exos treatment can significantly increase the expression of miR-199a-3p in the HK-2 cells.We further found that miR-199a-3p can inhibit apoptosis induced by H/R in vitro,and can inhibit the increase of Semaphorin 3A(Semaphorin 3A)expression caused by H/R,and further activate protein kinase B(AKT)and extracellular-signal-regulated protein kinase(ERK)signaling pathways.Conclusion: On the whole,this study demonstrated an antiapoptotic effect of hBMSC‐Exos,which protected against I/R injury,via delivering miR‐199a‐3p to renal cells,downregulating Sema3 A expression and thereby activating the AKT and ERK pathways.