Extracellular matrix (ECM) plays a critical role in the provision of the necessary microenvironment for the proper regeneration of the cardiac tissue. However, specific mechanisms that lead to ECM-mediated cardiac regeneration are not well understood. To elucidate the potential mechanisms, we investigated ultra-structures of the cardiac ECM using electron microscopy. Intriguingly, we observed large quantities of micro-vesicles from decellularized right atria. RNA and protein analyses revealed that these contained exosomal proteins and microRNAs (miRNAs), which we referred to herein as ECM-derived extracellular vesicles (ECM-EVs). One particular miRNA from ECM-EVs, miR-199a-3p, promoted cell growth of isolated neonatal cardiomyocytes and sinus nodal cells by repressing homeodomain-only protein (HOPX) expression and increasing GATA-binding 4 (Gata4) acetylation. To determine the mechanisms, we knocked down Gata4 and showed that miR-199a-3p actions required Gata4 for cell proliferation in isolated neonatal cardiomyocytes and sinus nodal cells. To further explore the role of this miRNA, we isolated neonatal cardiac cells and recellularized into atrial ECM, referred here has engineered atria. Remarkably, miR-199a-3p mediated the enrichment of cardiomyocyte and sinus nodal cell population, and enhanced electrocardiographic signal activity of sinus nodal cells in the engineered atria. Importantly, antisense of miRNA (antagomir) against miR-199a-3p was capable of abolishing these actions of miR-199a-3p in the engineered atria. We further showed in Ang II-infused animal model of sinus nodal dysfunction that miR-199-3p-treated cardiac cells remarkably ameliorated and restored the electrical activity as shown by normalization of the ECG, in contrast to untreated cells, which did not show electrical recovery. In conclusion, these results provide clear evidence of the critical role of ECM, in not only providing a scaffold for cardiac tissue growth, but also in promoting atrial electrical function through ECM-derived miR-199a-3p.

中文翻译：

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细胞外基质衍生的细胞外囊泡可促进工程性心房心肌细胞的生长和电活动

细胞外基质（ECM）在提供必要的微环境以使心脏组织正常再生中起着至关重要的作用。但是，导致ECM介导的心脏再生的具体机制还没有被很好地理解。为了阐明潜在的机制，我们使用电子显微镜研究了心脏ECM的超微结构。有趣的是，我们观察到了来自脱细胞的右心房的大量微囊泡。RNA和蛋白质分析显示，它们包含外泌体蛋白质和microRNA（miRNA），我们在本文中将其称为ECM衍生的细胞外囊泡（ECM-EV）。来自ECM-EV的一种特定的miRNA，miR-199a-3p，通过抑制仅同源异域蛋白（HOPX）表达并增加GATA结合4（Gata4）乙酰化作用，促进分离的新生儿心肌细胞和窦房结细胞的细胞生长。为了确定机制，我们敲低了Gata4并显示miR-199a-3p的作用需要Gata4在分离的新生儿心肌细胞和窦房结细胞中进行细胞增殖。为了进一步探索这种miRNA的作用，我们分离出了新生儿心脏细胞并重新细胞化为心房ECM，此处称为心房改造。值得注意的是，miR-199a-3p介导了工程化心房中心肌细胞和窦房结细胞群的富集，并增强了窦房结细胞的心电图信号活性。重要的是，针对miR-199a-3p的miRNA（antagomir）反义能够消除工程化心房中miR-199a-3p的这些作用。我们还进一步发现，在灌注了Ang II的窦房结功能异常的动物模型中，与未处理的未显示电信号的细胞相比，经miR-199-3p处理的心肌细胞显着改善并恢复了电活动，如心电图正常化所显示的那样。恢复。总之，这些结果清楚地证明了ECM的关键作用，不仅为心脏组织的生长提供了支架，而且还通过ECM衍生的miR-199a-3p促进了心房的电功能。