The Repair Role And Mechanism Of Fibroblasts Phenotype Transformation Promoted By Human Umbilical Cord Mesenchymal Stem Cells Derived Exosomes During Inflammatory Phases For Myocardial Infarction

Objective: Our previous studies showed that human umbilical cord mesenchymal stem cell-derived exosomes(hucMSC-exosomes)can promote the repair of myocardial injury post myocardial infarction(MI),but its mechanism had not been fully elucidated.This study will investigate whether hucMSC-exosomes can promote cardiac fibroblast-to-myofibroblast differentiation during inflammatory phases after MI,attenuate inflammatory responses and repair myocardial injury.Methods: HucMSCs and hucMSC-exosomes were identified by flow cytometry analysis,adipogenic and osteogenic differentiation assay,transmission electron microscopy,NTA(nanoparticle tracking analysis)technique and Western blot.The model of MI was established by permanent ligation of the left anterior descending(LAD)coronary artery in SD rats and injected intramyocardially hucMSC-exosomes immediately after ligation.In vitro,cardiac fibroblasts were treated by lipopolysaccharide(LPS)plus hucMSC-exosomes.Western blot,immunohistochemical staining,immunofluorescence staining,collagen gel contraction assay,Transwell migration assay,CCK-8 assay and qRT-PCR were used to evaluate the effect of hucMSC-exosomes on cardiac fibroblast phenotypic transformation and functional changes which included cell proliferation,migration and the expression of inflammatory factors.Hypoxic experiments in vitro,TUNEL staining and Western blot were used to investigate whether myofibroblasts that underwent phenotypic transformation during inflammatory phases can improve cardiomyocyte apoptosis and promote the repair of myocardial injury.Results: HucMSCs and hucMSC-exosomes were successfully isolated in vitro.HucMSCs expressed CD29,CD90 and CD105,but didnot express CD19,CD34 and CD45.In addition,hucMSCs differentiated into adipocytes and osteocytes successfully.HucMSC-exosomes showed a saucer-like shape that had double membrane structure.HucMSC-exosomes were a homogeneous population about 100 nm and expressed CD9,CD63.In vivo,the quantity of myofibroblasts in infarct areas in hucMSC-exosomes groups was higher than that in PBS groups during inflammatory phases post-MI.In vitro,hucMSC-exosomes also promoted fibroblast-to-myofibroblast differentiation under inflammatory conditions.When fibroblasts differentiated into myofibroblasts,their contractile ability was enhanced and migration ability was decreased.Simultaneously,the expression of pro-inflammatory factors was decreased and anti-inflammatory factor expression was increased.In vitro,cardiomyocyte apoptosis was reduced when treated with culture medium derived from fibroblasts pretreated with LPS plus hucMSC-exosomes.In vivo,cell apoptosis in hucMSC-exosomes groups was also less than that in PBS groups.Conclusions: HucMSC-exosomes promote fibroblasts differentiate into myofibroblasts during inflammatory phases after MI,attenuate inflammatory responses,improve cardiomyocyte apoptosis and repair myocardial injury.