The effect of mesenchymal stem cells-derived exosomes on the formation of intracranial aneurysm in rats by regulating miR143
Objective This study aims to explore the effect and mechanism of mesenchymal stem cells(MSCs)-derived exosomes regulating miR-143 on the formation of intracranial aneurysm(IA).Methods Primary rat bone marrow mesenchymal stem cells(BM-SCs)and brain vascular smooth muscle cells(VSMCs)were isolated.VSMCs were divided into PBS Group,Exosome-miR-NC Group,Exo-some-miR-143 Group,Exosome-miR-143+GW4869 Group,Overex-pression Control Group(oeNC Group),KLF5-overexpression Group(oeKLF5 Group)and oeKLF5+Exosome-miR-143 Group.BMSCs-derived exosomes were isolated by ultracentrifugation and identified by transmission electron microscopy and nano-tracer analysis.PKH26 staining was used to detect exosome transfer.Transfection of miR-143 mimic and miR-NC to BMSCs and VSMCs was performed.qRT-PCR was performed to determine miR-143 expression.West-ern blot was used to detect protein expressions.CCK8,EdU,flow cytometry,wound healing and Transwell assays were conducted to measure the viability,proliferation,apoptosis,and migration abilities of VSMCs.The dual luciferase reporter gene system was used to analyze the relationship between miR-143 and KLF5.A rat IA model was established to study the effects of BMSCs-derived exosome-miR-143 on the formation of IA in vivo.Results Compared with cell lysate Group,miR-143 expression in BMSCs exosomes was sig-nificantly increased(P＜0.01),and it could be transferred into VSMCs through exosomes.Compared with the exosome-miR-NC Group,the viability of VSMCs in the Exosome-miR-143 Group was increased(P＜0.01),while the number of EdU-positive cells,S-phase cells,apoptotic and migratory cells were significantly decreased(P＜0.01).Compared with the exosome-miR-143 group,the cell viability in exosome-miR-143+GW4869 group was decreased(P＜0.01),while the number of EdU-positive cells,S-phase cells,apoptotic and mi-gratory cells were significantly increased(P＜0.01).Compared with the miR-NC or the control group,the luciferase reporter gene activi-ty of WT-KLF5 and the mRNA and protein levels of KLF5 in the miR-143 group,the exosome-miR-143 group and the exosome-miR-NC groups were decreased(P＜0.01),while the luciferase reporter activity of MUT-KLF5 was unchanged(P＞0.05).Compared with the oeNC group,the cell viability in the oeKLF5 group was significantly decreased(P＜0.01),and the number of EdU-positive cells,S-phase cells,apoptotic and migratory cells were significantly increased(P＜0.01).Compared with the oeKLF5 group,the cell activity in the oeKLF5+Exosome-miR-143 group was significantly increased(P＜0.01),while the number of EdU-positive cells,S-phase cells,apoptotic and migratory cells were significantly decreased(P＜0.01).The results of in vivo experiments were consistent with those of cell experiment.Conclusion BMSCs-derived exosome-miR-143 inhibits the formation of rat IA by targeting KLF5.
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