The Effects And Mechanisms Of Empagliflozin Pretreated Exosomes Derived From Mesenchymal Stem Cells On Repair After Myocardial Infarction

Objective:Exosomes derived from mesenchymal stem cells（MSCs）play an important rol-e in the cardioprotection of acute myocardial infarction（AMI）.MSCs are a kind of pluri-potent stem cell population,which can be isolated from bone marrow,adipose tissue,um-bilical cord blood and other sources.MSCs have the characteristics of multidirectional dif-ferentiation potential and low immunogenicity.The therapeutic effects of MSCs are mainl-y mediated by their paracrine effects.Many studies have shown that exosomes derived fr-om MSCs can improve cardiac function after myocardial infarction by inhibiting cardiom-yocyte apoptosis and reducing fibrosis.Methods such as genetic engineering and pharmaco-logical compounds to optimize exosomes can further enhance the cardioprotective effect o-f exosomes.Sodium-glucose cotransporter 2 inhibitor（SGLT-2i）is a new oral hypoglyce-mic drug.Studies have shown that SGLT-2i can reduce cardiovascular disease mortality,a-nd its cardiovascular protective effect is independent of the hypoglycemic effect.The aim of this study is to investigate whether exosomes from MSCs pretreated with empagliflozi-n（EMPA）have enhanced cardioprotection after myocardial infarction and to explore the underlying mechanisms.Methods:Human bone marrow mesenchymal stem cells（hMSCs）were cultured in mediu-m containing different concentrations of EMPA to determine the optimal concentration of EMPA.Ed U staining assay,senescenceβ-Galactosidase staining assay,and scratch wound assay were used to evaluate the effect of EMPA on hMSCs.To extract,characterize and identify exosomes derived from hMSCs.In vitro,the H9c2 cells model was established b-y using serum-free edium containing 120μM H₂O₂,which was co-cultured with hMSCs derived exosome.Caspase-3/7 staining assay and PI staining assay were used to evaluate the effect of EMPA pretreated hMSCs-derived exosomes（EMPA-Exo）on the apoptosis o-f cardiomyocytes after ischemia-hypoxia.In vivo,myocardial infarction was induced in S-prague-Dawley（SD）rats by ligation of the left anterior descending coronary artery,and h-MSCs derived exosomes were injected into the infarct area.Echocardiography,masson sta-ining,sirius red staining,HE staining,immunofluorescence and immunohistochemistry wer-e used to evaluate the effects of EMPA-Exo on cardiac function,infarct size,myocardial fibrosis and myocardial apoptosis in rats with myocardial infarction.Results:Firstly,pretreatment with EMPA could improve the viability and migration abilit-y of hMSCs,and inhibit the apoptosis and senescence of hMSCs.EMPA pretreated h MS-Cs-derived exosomes had a typical structure and particle size.Secondly,to evaluate the pr-otective effect of EMPA pretreated hMSCs-derived exosomes on H9c2 cells.In vitro,we found that EMPA-Exo could significantly inhibit the apoptosis of H9c2 cells compared with the control group（MSC-Exo）.In vivo,EMPA-Exo and MSC-Exo were injected into the infarct area,respectively.Compared with MSC-Exo,EMPA-Exo inhibited myocardial apoptosis and promoted angiogenesis in the infarct border zone more significantly.At the same time,EMPA-Exo reduced myocardial fibrosis,infarct size and improved cardiac func-tion.Finally,by sequencing exosomal miRNAs,we identified exosomal differentially expr-essed miRNAs,among which miR-214-3p was significantly elevated in EMPA-Exo.HMS-Cs derived exosomes with high miR-214-3p expression showed similar protective effects t-o EMPA-Exo when co-cultured with H9c2 cells.In addition,miR-214-3p was also found to promote AKT phosphorylation in H9c2 cells.Conclusions:Our data suggest that EMPA-Exo significantly improve cardiac repair after MI by inhibiting myocardial apoptosis.Mi R-214-3p may mediate the myocardial protective effect of EMPA-Exo at least in part through AKT signaling pathway.