微小RNA-423-5p在脂肪间充质干细胞外泌体治疗老龄心力衰竭大鼠的作用及机制

Role of mIR-423-5p in treatment of aged HF rats with adipose mesenchymal stem cells-derived exosomes and its mechanism

目的探讨微小RNA(miRNA,miR)-423-5p在脂肪间充质干细胞(MSC)外泌体治疗老龄心力衰竭(HF)大鼠的作用机制。方法选择18月龄SPF级大鼠38只,随机分为治疗组24只和模型组14只,2组腹腔注射异丙肾上腺素14 d,构造HF模型。治疗组给予心肌内注射脂肪MSC外泌体后饲养2周。用Western blot鉴定其纯度;通过苏木精-伊红染色和Masson染色检测心肌损伤及纤维化程度;通过TargetScan和miRDB数据库对miR-423-5p的靶基因进行预测,并预测信号通路;用实时荧光定量PCR检测miR-423-5p、磷脂酰肌醇3激酶(PI3K)和蛋白激酶B(Akt)基因表达,Western blot检测PI3K和Akt蛋白表达。结果与模型组比较,治疗组miR-423-5p表达量明显降低(P<0.05)。苏木精-伊红和Masson染色显示,治疗组心肌损伤和纤维化均较模型组明显改善。TargetScan和miRDB数据库预测显示,miR-423-5p的靶基因分别是141个和545个,Draw Venn分析显示,2个数据库共同靶基因为52个,占总量的8.2%。预测分析显示,通路富集出4条信号通路,排在首位的为PI3K-Akt信号通路。与模型组比较,治疗组PI3K、Akt基因和蛋白表达显著下降(P<0.05)。结论脂肪MSC外泌体能治疗老龄HF大鼠,其机制可能通过miR-423-5p调节PI3K-Akt信号通路的途径产生作用。

Objective To study the mechanism of miR-423-5p in treatment of aged HF rats with adipose mesenchymal stem cells-derived exosomes.Methods Thirty-eight male SPF rats aged 18 months were randomly divided into treatment group(n=24)and model group(=14).A HF model of rats was established by intraperitoneal injection with isoproterenol for 14 days.The rats in treatment group were fed for 2 weeks after intramyocardial injection with adipose mesenchymal stem cells-derived exosomes.The purity of adipose mesenchymal stem cells-derived exosomes was tested by Western blot.The myocardial damage and fibrosis were assayed with HE staining and Masson staining.The miR-423-5p target genes and their signaling pathways were retrieved from TargetScan database and miRDB database.The expressions of miR-423-5p,PI3K and Akt genes were detected by qPCR and those of PI3K and Akt protein were detected by Western blot.Results The expression levels of miR-423-5p,BNP and ANP were significantly lower in treatment group than in model group(P<0.05).HE staining and Masson staining showed that myocardial damage and fibrosis were improved more significantly in treatment group than in model group.A total of 141 and 545 miR-423-5p target genes were retrieved from TargetScan database and miRDB database respectively.Draw Venn analysis showed that the 52 common target genes covered in the two databases accounted for 8.2%of the total target genes.Four possible signaling pathways were enriched from the pathway with the PI3K-Akt signaling pathway ranked first.The expression levels of PI3K and Akt gene and protein were significantly lower in treatment group than in model group(P<0.05).Conclusion Adipose mesenchymal stem cells-derived exosomes can treat HF in aged rats by regulating the PI3K-Akt signaling pathways through miR-423-5p.