Department of Cardiovascular Surgery Intensive Care Unit, The Second Affiliated Hospital of Hainan Medical College, Haikou City, Hainan Province, China.
Department of Critical Care Medicine, The Second Affiliated Hospital of Hainan Medical College, Haikou City, Hainan Province, China.
Cardiovasc Drugs Ther. 2023 Oct;37(5):891-904. doi: 10.1007/s10557-022-07339-7. Epub 2022 May 11.
Mesenchymal stem cell (MSC)-derived exosomes play significant roles in ameliorating cardiac damage after myocardial ischemia-reperfusion (I/R) injury. Long non-coding RNA alpha-2-macroglobulin antisense RNA 1 (Lnc A2M-AS1) was found that might protect against myocardial I/R. However, whether Lnc A2M-AS1 delivery via MSC-derived exosomes could also regulate myocardial I/R injury remains unknown.
Exosomes were isolated by ultracentrifugation, and qualified by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and Western blot. Hypoxia/reoxygenation (H/R) treatment in human cardiomyocytes was used to mimic the process of myocardial I/R in vitro. The viability and apoptosis of cardiomyocytes were detected using cell counting kit-8, flow cytometry, and Western blot assays. The contents of lactate dehydrogenase (LDH), malondialdehyde (MDA), and superoxide dismutase (SOD) were evaluated using corresponding commercial kits. The quantitative real-time polymerase chain reaction and Western blot were used to determine the expression levels of Lnc A2M-AS1, microRNA (miR)-556-5p, and X-linked inhibitor of apoptosis protein (XIAP). The binding interaction between miR-556-5p and Lnc A2M-AS1 or XIAP was confirmed by the dual-luciferase reporter, RIP and pull-down assays.
Exosomes isolated from hMSCs (hMSCs-exo) attenuated H/R-induced apoptosis and oxidative stress in cardiomyocytes. Lnc A2M-AS1 was lowly expressed in AMI patients and H/R-induced cardiomyocytes. Besides, Lnc A2M-AS1 was detectable in hMSCs-exo, exosomes derived from Lnc A2M-AS1-transfected hMSCs weakened H/R-induced apoptosis and oxidative stress, and enhanced the protective action of hMSCs-exo on H/R-induced cardiomyocytes. Further mechanism analysis showed that Lnc A2M-AS1 acted as a sponge for miR-556-5p to increase XIAP expression level. Importantly, miR-556-5p overexpression or XIAP knockdown reversed the action of exosomal Lnc A2M-AS1 on H/R-induced cardiomyocytes.
Lnc A2M-AS1 delivery via MSC-derived exosomes ameliorated H/R-induced cardiomyocyte apoptosis and oxidative stress via regulating miR-556-5p/XIAP, opening a new window into the pathogenesis of myocardial I/R injury.
间充质干细胞（MSC）衍生的外泌体在改善心肌缺血再灌注（I/R）损伤后心脏损伤方面发挥重要作用。长链非编码 RNA α-2-巨球蛋白反义 RNA 1（Lnc A2M-AS1）被发现可能对心肌 I/R 具有保护作用。然而，MSC 衍生的外泌体递送 Lnc A2M-AS1 是否也可以调节心肌 I/R 损伤仍然未知。
通过超速离心分离外泌体，并通过透射电子显微镜（TEM）、纳米颗粒跟踪分析（NTA）和 Western blot 进行鉴定。体外使用缺氧/复氧（H/R）处理模拟心肌 I/R 过程。使用细胞计数试剂盒-8、流式细胞术和 Western blot 测定检测心肌细胞的活力和凋亡。使用相应的商业试剂盒评估乳酸脱氢酶（LDH）、丙二醛（MDA）和超氧化物歧化酶（SOD）的含量。使用定量实时聚合酶链反应和 Western blot 测定确定 Lnc A2M-AS1、微小 RNA（miR）-556-5p 和 X 连锁凋亡抑制蛋白（XIAP）的表达水平。通过双荧光素酶报告、RIP 和下拉实验证实 miR-556-5p 与 Lnc A2M-AS1 或 XIAP 之间的结合相互作用。
从 hMSCs（hMSCs-exo）分离的外泌体减轻了 H/R 诱导的心肌细胞凋亡和氧化应激。Lnc A2M-AS1 在 AMI 患者和 H/R 诱导的心肌细胞中表达水平较低。此外，Lnc A2M-AS1 可在 hMSCs-exo 中检测到，转染 Lnc A2M-AS1 的 hMSCs 衍生的外泌体减弱了 H/R 诱导的凋亡和氧化应激，并增强了 hMSCs-exo 对 H/R 诱导的心肌细胞的保护作用。进一步的机制分析表明，Lnc A2M-AS1 作为 miR-556-5p 的海绵以增加 XIAP 表达水平。重要的是，miR-556-5p 的过表达或 XIAP 的敲低逆转了外泌体 Lnc A2M-AS1 对 H/R 诱导的心肌细胞的作用。
通过 MSC 衍生的外泌体递送 Lnc A2M-AS1 通过调节 miR-556-5p/XIAP 改善 H/R 诱导的心肌细胞凋亡和氧化应激，为心肌 I/R 损伤的发病机制开辟了新的窗口。
