Objective To explore the effects of M1 macrophage-derived exosome microRNA(miR)-16-5p on electrophysiology of atrial myocytes and explore the potential mechanisms.Methods Mononuclear macrophages(RAW264.7 cells)were divided into lipopolysaccharide(LPS)group(RAW264.7 cells were induced and stimulated by LPS for 24 h to differentiate into M1 macrophages),miR-16-5p negative control(NC)group(miR-16-5p NC was transfected into RAW264.7 cells and induced differentiation),miR-16-5p mimics group(miR-16-5p mimics were transfected into RAW264.7 cells and induced differentiation),miR-16-5p inhibitor group(miR-16-5 p inhibitor was transfected into RAW264.7 cells and induced differentiation)and LPS+neutral sphingomyelin inhibitor(GW4869)group(RAW264.7 cells were induced by LPS and then cultured with 10 μM GW4869).Supernatant was collected after culture of macrophages for 48-72 h in 5 groups,and exosomes were obtained by ultracentrifugation.Real-time fluorescent PCR(qRT-PCR)was used to detect relative expression level of miR-16-5p in transfected macrophages.Atrial myocytes(HL-1 cells)were exposed to rapid electrical stimulation(1.0 V/cm,10 Hz)for 48 h to construct rapid pacing-induced atrial fibrillation cell model,then co-cultured with exosomes from each group.Atrial myocytes were divided into A group(pacing HL-1 cells+exosomes from LPS group),B group(pacing HL-1 cells+exosomes from miR-16-5p NC group),C group(pacing HL-1 cells+exosomes from miR-16-5p mimics group),D group(pacing HL-1 cells+exosomes from miR-16-5p inhibitor group)and E group(pacing HL-1 cells+exosomes from LPS+GW4869 group).Patch clamp was used to detect action potential duration at 50％and 90％repolarization(APD50 and APD90).Western blotting was uesd to detect expression levels of phosphatidylinositol kinase(PI3K),protein kinase B(AKT),phosphorylated AKT(p-AKT)in cells of 5 groups.Results Relative expression level of miR-16-5p in macrophages of miR-16-5p mimics group was higher than that of miR-16-5p NC group(P＜0.001),while miR-16-5p inhibitor group was lower than that of miR-16-5p NC group(P＜0.05).There were no significant difference of APD50 and APD90 in HL-1 cells between A group and B group(P＞0.05).APD50 and APD90 in HL-1 cells of C group were all shorter than those in A group(P＜0.05).APD50 and APD90 in HL-1 cells of D group and E group were longer than those in A group(P＜0.01).There were no significant difference in expression levels of PI3K and P-Akt/AKT in HL-1 cells between A group and B group(P＞0.05).Expression levels of PI3K and P-Akt/AKT in HL-1 cells of C group were lower than those in A group(P＜0.001).Expression levels of PI3K and P-Akt/AKT in HL-1 cells of D group and E group were higher than those in A group(P＜0.001).Conclusion Ml macrophage-derived exosome miRNA-16-5p can shorten action potential duration of atrial myocytes,possibly associated with the inhibition of the PI3K/AKT pathway.
维普
万方数据