骨髓基质细胞来源的外泌体通过 miR-486-5p 靶向 PTEN/PI3K/AKT 信号通路抑制缺血缺氧条件下的心肌细胞凋亡。
Exosomes of bone-marrow stromal cells inhibit cardiomyocyte apoptosis under ischemic and hypoxic conditions via miR-486-5p targeting the PTEN/PI3K/AKT signaling pathway.
机构信息
Department of Cardiology, The First Affiliated Hospital of Soochow University, Suzhou 215006, PR China; Department of Cardiology, Shaoyang Central Hospital, Shaoyang 422000, PR China.
Department of Cardiovascular Surgery, The First Affiliated Hospital of Soochow University, Suzhou 215006, PR China.
出版信息
Thromb Res. 2019 May;177:23-32. doi: 10.1016/j.thromres.2019.02.002. Epub 2019 Feb 2.
BACKGROUND
Myocardial ischemia-reperfusion injury (MIRI) is a major obstacle in the treatment of ischemic heart disease. Recent studies have shown that exosomes-small membrane vesicles secreted by most cell types-could have a protective effect on the ischemic myocardium. In this study we explored the effect of exosomes derived from bone-marrow stromal cells (BMSC-exo) on cardiomyocyte apoptosis and MIRI.
METHODS
Exosomes were purified from culture media using the ExoQuick kit and observed using transmission electron microscopy. Cell viability was assessed by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Cell apoptosis was analyzed by flow cytometry using the Annexin-V/PI stain. The expression levels of microRNA (miRNA), messenger RNA (mRNA) and PTEN/PI3K/AKT-pathway-related proteins were detected by qRT-PCR and western blot, respectively. Myocardial ischemia was simulated by incubating H9C2 cells in a hypoxia/reoxygenation (H/R) conditioned rat MIRI model.
RESULTS
BMSC-exo induced the proliferation of H9C2 cells and rescued H9C2 cells from apoptosis in the H/R model, indicating that BMSC-exo has a protective effect on cardiomyocyte injury caused by H/R. Using transgenic H9C2 cells, we found that miR-486-5p in BMSC-exo suppressed the H/R-triggered apoptosis of H9C2 cells. In addition, BMSC-exo repressed the expression of PTEN in H9C2 cells via miR-486-5p, and subsequently activated the PI3K/AKT pathway in vitro. Moreover, the myocardial injury caused by ischemia/reperfusion was repaired by BMSC-exo which activates the PI3K/AKT pathway via miR-486-5p in vivo.
CONCLUSION
Our results suggested that exosomes from BMSCs have a protective effect on myocardium ischemic injury. MiR-486-5p carried by BMSC-exo plays a pivotal role in the regulatory process by suppressing PTEN expression, activating the PI3K/AKT signaling pathway, and subsequently inhibiting the apoptosis of injured cardiomyocytes.
背景
心肌缺血再灌注损伤（MIRI）是缺血性心脏病治疗的主要障碍。最近的研究表明，外泌体——大多数细胞类型分泌的小膜囊泡——可能对缺血性心肌具有保护作用。在这项研究中，我们探讨了骨髓基质细胞（BMSC）来源的外泌体（BMSC-exo）对心肌细胞凋亡和 MIRI 的影响。
方法
使用 ExoQuick 试剂盒从培养物中纯化外泌体，并通过透射电子显微镜观察。通过 3-(4,5-二甲基-2-噻唑基)-2,5-二苯基-2-H-四唑溴盐（MTT）测定法评估细胞活力。使用 Annexin-V/PI 染色通过流式细胞术分析细胞凋亡。通过 qRT-PCR 和 Western blot 分别检测 microRNA（miRNA）、信使 RNA（mRNA）和 PTEN/PI3K/AKT 通路相关蛋白的表达水平。通过孵育 H9C2 细胞于缺氧/复氧（H/R）条件性大鼠 MIRI 模型中模拟心肌缺血。
结果
BMSC-exo 诱导 H9C2 细胞增殖，并在 H/R 模型中挽救 H9C2 细胞免于凋亡，表明 BMSC-exo 对 H/R 引起的心肌细胞损伤具有保护作用。使用转基因 H9C2 细胞，我们发现 BMSC-exo 中的 miR-486-5p 抑制 H9C2 细胞的 H/R 触发的凋亡。此外，BMSC-exo 通过 miR-486-5p 抑制 H9C2 细胞中 PTEN 的表达，随后在体外激活 PI3K/AKT 通路。此外，BMSC-exo 通过体内 miR-486-5p 激活 PI3K/AKT 通路修复由缺血/再灌注引起的心肌损伤。
结论
我们的结果表明，BMSC 来源的外泌体对心肌缺血性损伤具有保护作用。BMSC-exo 携带的 miR-486-5p 通过抑制 PTEN 表达、激活 PI3K/AKT 信号通路，从而抑制受损心肌细胞的凋亡，在调节过程中发挥关键作用。
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