Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai Institute of Cardiovascular Diseases, Shanghai, China.
Department of Cardiovascular, Affiliated Hospital of Nantong University, Jiangsu, China.
Acta Physiol (Oxf). 2020 Apr;228(4):e13439. doi: 10.1111/apha.13439. Epub 2020 Jan 21.
To explore the role of the histidine triad nucleotide-binding 2 (HINT2) protein in heart failure.
Neonatal mouse ventricle myocytes (NMVMs) and myocardial infarction-induced heart failure mice were used for in vitro or in vivo experiments. Adenovirus (ADV) and adeno-associated virus serum type 9 (AAV9) vectors were used to regulate HINT2 expression. The expression of HINT2 was determined by quantifying the mRNA and protein levels. Cell survival was analysed using the CCK-8 kit and TUNEL staining. Mitochondrial function was determined by the mitochondrial membrane potential and oxygen consumption rates. AAV9-HINT2 was injected 24 h post-myocardial infarction following which transthoracic echocardiography and histological analyses were performed after 4 weeks. Positron emission tomography tomography-computed tomography (PET/CT) and targeted metabolomics analyses were used to explore the metabolic status in vivo. NAD levels were measured using a colorimetric kit. Computer-simulated rigid body molecular docking was performed using AUTODOCK4. Molecule binding kinetics assays were performed using biolayer interferometry.
HINT2 was down-regulated in NMVMs in hypoxia. ADV-HINT2-induced HINT2 overexpression improved NMVM survival after exposure to hypoxia. Mitochondrial function was preserved in the ADV-HINT2 group under hypoxic conditions. In vivo experiments showed that cardiac function and metabolic status was preserved by HINT2 overexpression. HINT2 overexpression restored mitochondrial NAD levels; this was dependent on nicotinamide mononucleotide (NMN). Using computer-simulated molecular docking analysis and biolayer interferometry, we observed that HINT2 potentially binds and associates with NMN.
HINT2 overexpression protects cardiac function in adult mice after myocardial infarction by maintaining mitochondrial NAD homeostasis.
探讨组氨酸三核苷酸结合蛋白 2（HINT2）在心力衰竭中的作用。
使用新生鼠心室肌细胞（NMVMs）和心肌梗死诱导的心力衰竭小鼠进行体外或体内实验。腺病毒（ADV）和腺相关病毒血清型 9（AAV9）载体用于调节 HINT2 的表达。通过定量检测 mRNA 和蛋白水平来确定 HINT2 的表达。使用 CCK-8 试剂盒和 TUNEL 染色分析细胞存活率。通过线粒体膜电位和耗氧量来确定线粒体功能。心肌梗死后 24 h 注射 AAV9-HINT2，4 周后进行经胸超声心动图和组织学分析。正电子发射断层扫描计算机断层扫描（PET/CT）和靶向代谢组学分析用于体内代谢状态的研究。使用比色法试剂盒测量 NAD 水平。使用 AUTODOCK4 进行计算机模拟刚性体分子对接。使用生物层干涉法进行分子结合动力学测定。
缺氧时 NMVMs 中 HINT2 下调。ADV-HINT2 诱导的 HINT2 过表达可改善 NMVM 在缺氧暴露后的存活率。在缺氧条件下，ADV-HINT2 组保留了线粒体功能。体内实验表明，过表达 HINT2 可保存心脏功能和代谢状态。HINT2 过表达恢复了线粒体 NAD 水平；这依赖于烟酰胺单核苷酸（NMN）。通过计算机模拟分子对接分析和生物层干涉法，我们观察到 HINT2 可能与 NMN 结合和相互作用。
过表达 HINT2 通过维持线粒体 NAD 稳态来保护心肌梗死后成年小鼠的心脏功能。
