Department of Cardiac Surgery, Medical University of Innsbruck, Anichstrasse 35, 6020 Innsbruck, Austria.
Austrian Cluster for Tissue Regeneration, Vienna, Austria.
Cardiovasc Res. 2020 May 1;116(6):1226-1236. doi: 10.1093/cvr/cvz209.
As many current approaches for heart regeneration exert unfavourable side effects, the induction of endogenous repair mechanisms in ischaemic heart disease is of particular interest. Recently, exosomes carrying angiogenic miRNAs have been described to improve heart function. However, it remains challenging to stimulate specific release of reparative exosomes in ischaemic myocardium. In the present study, we sought to test the hypothesis that the physical stimulus of shock wave therapy (SWT) causes the release of exosomes. We aimed to substantiate the pro-angiogenic impact of the released factors, to identify the nature of their cargo, and to test their efficacy in vivo supporting regeneration and recovery after myocardial ischaemia.
Mechanical stimulation of ischaemic muscle via SWT caused extracellular vesicle (EV) release from endothelial cells both in vitro and in vivo. Characterization of EVs via electron microscopy, nanoparticle tracking analysis and flow cytometry revealed specific exosome morphology and size with the presence of exosome markers CD9, CD81, and CD63. Exosomes exhibited angiogenic properties activating protein kinase b (Akt) and extracellular-signal regulated kinase (ERK) resulting in enhanced endothelial tube formation and proliferation. A miRNA array and transcriptome analysis via next-generation sequencing were performed to specify exosome content. miR-19a-3p was identified as responsible cargo, antimir-19a-3p antagonized angiogenic exosome effects. Exosomes and target miRNA were injected intramyocardially in mice after left anterior descending artery ligation. Exosomes resulted in improved vascularization, decreased myocardial fibrosis, and increased left ventricular ejection fraction as shown by transthoracic echocardiography.
The mechanical stimulus of SWT causes release of angiogenic exosomes. miR-19a-3p is the vesicular cargo responsible for the observed effects. Released exosomes induce angiogenesis, decrease myocardial fibrosis, and improve left ventricular function after myocardial ischaemia. Exosome release via SWT could develop an innovative approach for the regeneration of ischaemic myocardium.
由于目前许多心脏再生方法都有不利的副作用，因此，诱导缺血性心脏病的内源性修复机制特别有趣。最近，携带血管生成 miRNA 的外泌体已被描述为可改善心脏功能。然而，刺激缺血心肌中特定的修复性外泌体释放仍然具有挑战性。在本研究中，我们试图检验冲击波治疗（SWT）的物理刺激会导致外泌体释放的假设。我们旨在证实释放因子的促血管生成作用，确定其货物的性质，并测试其在体内支持心肌缺血后再生和恢复的功效。
通过 SWT 对缺血肌肉进行机械刺激，在体外和体内均可引起内皮细胞释放细胞外囊泡（EV）。通过电子显微镜、纳米颗粒跟踪分析和流式细胞术对 EV 进行特征描述，显示出具有外泌体特征的形态和大小，并且存在外泌体标志物 CD9、CD81 和 CD63。外泌体具有激活蛋白激酶 b（Akt）和细胞外信号调节激酶（ERK）的血管生成特性，从而导致增强的内皮管形成和增殖。通过下一代测序进行 miRNA 阵列和转录组分析以确定外泌体的内容。鉴定出 miR-19a-3p 为负责的货物，抗 miR-19a-3p 拮抗血管生成外泌体的作用。在左前降支结扎后，将外泌体和靶 miRNA 经心肌内注射到小鼠体内。外泌体可改善血管化，减少心肌纤维化，并提高经胸超声心动图显示的左心室射血分数。
SWT 的机械刺激会导致血管生成外泌体的释放。miR-19a-3p 是负责观察到的作用的囊泡货物。释放的外泌体可诱导血管生成，减少心肌纤维化，并改善心肌缺血后的左心室功能。通过 SWT 释放外泌体可能为缺血性心肌的再生开发一种创新方法。
