AIM To explore the role of the histidine triad nucleotide-binding 2 (HINT2) protein in heart failure. METHODS 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. RESULTS 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. CONCLUSION HINT2 overexpression protects cardiac function in adult mice after myocardial infarction by maintaining mitochondrial NAD homeostasis.

中文翻译：

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组氨酸三联体核苷酸结合蛋白2的过表达保护成年小鼠急性心肌梗塞后的心脏功能。

目的探讨组氨酸三联体核苷酸结合蛋白2（HINT2）在心力衰竭中的作用。方法采用新生小鼠心室肌细胞（NMVM）和心肌梗死诱发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9型（AAV9）载体用于调节HINT2表达。通过定量mRNA和蛋白质水平来确定HINT2的表达。使用CCK-8试剂盒和TUNEL染色分析细胞存活。线粒体功能由线粒体膜电位和耗氧率决定。心肌梗死后24小时注射AAV9-HINT2，然后在4周后进行经胸超声心动图和组织学分析。正电子发射断层摄影术，计算机断层摄影术（PET / CT）和靶向代谢组学分析被用于探索体内代谢状态。使用比色试剂盒测量NAD水平。使用AUTODOCK4进行计算机模拟的刚体分子对接。使用生物层干涉术进行分子结合动力学测定。结果在缺氧的NMVM中HINT2被下调。ADV-HINT2诱导的HINT2过表达提高了缺氧后NMVM的存活率。缺氧条件下ADV-HINT2组的线粒体功能得以保留。体内实验表明，HINT2的过表达保留了心脏功能和代谢状态。HINT2过表达恢复了线粒体NAD水平；这取决于烟酰胺单核苷酸（NMN）。使用计算机模拟的分子对接分析和生物层干涉法，我们观察到HINT2可能与NMN结合并缔合。结论HINT2的过表达通过维持线粒体NAD稳态来保护心肌梗死后成年小鼠的心脏功能。