Rationale: Mesenchymal stem cells (MSCs) play important roles in tissue repair and regeneration. However, the molecular mechanisms underlying MSCs activation remain largely unknown, thus hindering their clinical translation. Exosomes are small vesicles that act as intercellular messengers, and their potential for stem cell activation in pathological conditions has not been fully characterized yet. Here, we aim to investigate whether serum exosomes are involved in the remote activation of MSCs after myocardial infarction (MI).

Methods: We established MI mouse model by ligating the left anterior descending branch of the coronary artery. Afterwards, serum exosomes were isolated from control (Con Exo) and MI mice (MI Exo) by differential centrifugation. Exosomes were characterized through transmission electron microscopy and nanoparticle tracking analysis. The cell proliferation rate was evaluated by CCK-8 and EdU incorporation assays. Exosomal miRNA and protein levels were assessed using qRT-PCR and western blotting, respectively. VEGF levels in the supernatant and serum were quantified by ELISA. Matrigel plug and tube formation assays were used to evaluate angiogenesis. To explore miR-1956 roles, overexpression and knock-down experiments were performed using mimic and inhibitor, respectively. Finally, miR-1956 target genes were confirmed using the luciferase reporter assay.

Results: Both types of exosomes exhibited typical characteristics and could be internalized by adipose-derived MSCs (ADMSCs). MI Exo enhanced ADMSCs proliferation through the activation of ERK1/2. Gain- and loss-of-function studies allowed the validation of miR-1956 (enriched in MI Exo) as the functional messenger that stimulates ADMSCs-mediated angiogenesis and paracrine VEGF signaling, by downregulating Notch-1. Finally, we found that the ischemic myocardium and kidney may be the main sources that release serum exosomes after MI.

Conclusions: Cardio-renal exosomes deliver miR-1956 and activate paracrine proangiogenic VEGF signaling in ADMSCs after MI; this process also involves Notch-1, which functions as the core mediator.

基本原理：间充质干细胞（MSC）在组织修复和再生中起重要作用。但是，MSCs激活的分子机制仍是未知之数，因此阻碍了它们的临床翻译。外泌体是充当细胞间信使的小囊泡，其在病理条件下激活干细胞的潜力尚未得到充分表征。在这里，我们的目的是调查心肌梗死（MI）后血清外泌体是否参与MSC的远程激活。

方法：我们结扎冠状动脉左前降支建立了MI小鼠模型。之后，通过差速离心从对照（Con Exo）和MI小鼠（MI Exo）分离血清外泌体。外泌体通过透射电子显微镜和纳米粒子跟踪分析进行了表征。通过CCK-8和EdU掺入测定法评估细胞增殖速率。分别使用qRT-PCR和Western blotting评估外泌体miRNA和蛋白质水平。通过ELISA定量上清液和血清中的VEGF水平。基质胶塞和管形成试验用于评估血管生成。为了探索miR-1956的作用，分别使用模拟物和抑制剂进行了过表达和敲低实验。最后，使用荧光素酶报告基因测定法确认了miR-1956靶基因。

结果：两种类型的外泌体均表现出典型特征，并且可以被脂肪来源的MSC（ADMSC）内在化。MI Exo通过激活ERK1 / 2增强了ADMSC的增殖。功能获得和功能丧失研究允许通过下调Notch-1来验证miR-1956（富含MI Exo）作为刺激ADMSCs介导的血管生成和旁分泌VEGF信号传导的功能信使。最后，我们发现缺血性心肌和肾脏可能是MI后释放血清外泌体的主要来源。

结论：心肌梗死后ADMSCs中肾肾外泌体传递miR-1956并激活旁分泌促血管生成的VEGF信号传导。此过程还涉及Notch-1，它是核心介体。