Li Qiming, Jin Yunpeng, Ye Xiaoqi, Wang Wei, Deng Gang, Zhang Xiaojian
The Department of Cardiology, The Fourth Affiliated Hospital of Zhejiang University School of Medicine, N1 Shangcheng Road, Yiwu, 322000, Zhejiang, China.
Nursing Department, The Fourth Affiliated Hospital of Zhejiang University School of Medicine, Yiwu, 322000, Zhejiang, China.
Nanoscale Res Lett. 2021 Jul 2;16(1):111. doi: 10.1186/s11671-021-03559-2.
Myocarditis is a disease characterized by localized or diffuse inflammation of the myocardium without efficient treatment. This study explored the regulatory mechanism of microRNA-133 (miR-133) secreted from bone marrow mesenchymal stem cell-derived exosome (BMSC-Exo) on myocardial fibrosis and epithelial-mesenchymal transition (EMT) in viral myocarditis (VMC) rats through regulating mastermind-like 1 (MAML1). BMSCs in rats were isolated and cultured to identify their immune phenotype and osteogenic and adipogenic ability, and BMSC-Exo were extracted and identified. Exosomes were obtained through ultracentrifugation, which were identified by transmission electron microscope and western blot analysis. The rats were injected with Coxsackie B3 virus for preparation of VMC model, and cardiomyocytes were isolated, cultured and grouped in the same way as animal experiments (NC, Ad-miR-133a, Adas-miR-133a). In vivo and in vitro experiments were conducted to figure out the roles of exosomal miR-133a and MAML1 in inflammation, apoptosis, EMT, fibrosis, and cell viability. The targeting relationship between miR-133a and MAML1 was verified by dual luciferase reporter gene assay. BMSC-Exo raised miR-133a expression in VMC rats and effectively improved the VMC rat cardiac function and myocardial fibrosis, increased cardiomyocyte viability and inhibited the EMT process. Elevated miR-133a in exosomes strengthened the improvements. Silenced miR-133a effectively reversed the effects of BMSC-Exo on VMC rats. miR-133a targeted MAML1. Inhibition of MAML1 improved cardiac function and myocardial fibrosis in VMC rats and could reverse the effect of miR-133a-silenced exosomes on VMC rats. Our study suggests that elevated exosomal miR-133a suppresses myocardial fibrosis and EMT in rats with VMC via down-regulating MAML1, thereby inhibiting the progression of myocarditis.
心肌炎是一种以心肌局限性或弥漫性炎症为特征且缺乏有效治疗方法的疾病。本研究通过调控主调控样蛋白1（MAML1），探索骨髓间充质干细胞来源的外泌体（BMSC-Exo）分泌的微小RNA-133（miR-133）对病毒性心肌炎（VMC）大鼠心肌纤维化和上皮-间质转化（EMT）的调控机制。分离并培养大鼠的骨髓间充质干细胞以鉴定其免疫表型以及成骨和成脂能力，然后提取并鉴定BMSC-Exo。通过超速离心获得外泌体，并用透射电子显微镜和蛋白质免疫印迹分析进行鉴定。给大鼠注射柯萨奇B3病毒以制备VMC模型，分离、培养心肌细胞并按照与动物实验相同的方式进行分组（NC、Ad-miR-133a、Adas-miR-133a）。进行体内和体外实验以明确外泌体miR-133a和MAML1在炎症、凋亡、EMT、纤维化及细胞活力方面的作用。通过双荧光素酶报告基因检测验证miR-133a与MAML1之间的靶向关系。BMSC-Exo提高了VMC大鼠中miR-133a的表达，并有效改善了VMC大鼠的心脏功能和心肌纤维化，增加了心肌细胞活力并抑制了EMT过程。外泌体中升高的miR-133a增强了这些改善作用。沉默miR-133a有效地逆转了BMSC-Exo对VMC大鼠的作用。miR-133a靶向MAML1。抑制MAML1可改善VMC大鼠的心脏功能和心肌纤维化，并可逆转miR-133a沉默外泌体对VMC大鼠的作用。我们的研究表明，外泌体miR-133a升高通过下调MAML1抑制VMC大鼠的心肌纤维化和EMT，从而抑制心肌炎的进展。
