目的 阐明通脉养心丸通过调节过氧化物酶体增殖物激活受体γ共激活因子-1α（peroxisome proliferator-activated receptor gamma coactivator-1α，PGC-1α）介导的线粒体功能从而发挥心肌缺血/再灌注损伤（myocardial ischemia-reperfusion injury，MI/RI）保护作用的机制。方法 构建缺氧复氧（hypoxia-reoxygenation，H/R）心肌细胞模型，给予通脉养心丸干预后，采用DCFH-DA和MitoSOX^TM Red荧光探针检测细胞内活性氧（reactive oxygen species，ROS）水平；采用增强型三磷酸腺苷（adenosine triphosphate，ATP）检测试剂盒检测ATP水平；JC-1染色观察线粒体膜电位变化；采用Seahorse XF细胞线粒体压力测试分析线粒体呼吸功能；Mitotracker与透射电镜观察线粒体的形态与结构变化；采用Western blotting与qRT-PCR检测线粒体融合分裂[线粒体融合素1（mitofusin 1，Mfn1）、线粒体融合素2（mitofusin 2，Mfn2）、线粒体分裂蛋白1（fission 1，Fis1）、动力相关蛋白1（dynamin-related protein 1，Drp1）]、生物合成[核呼吸因子1&（nuclear respiratory factor 1，Nrf1）、线粒体转录因子A（mitochondrial transcription factor A，&TFAM）、线粒体DNA（mitochondrial DNA，mtDNA）拷贝数]及自噬[Beclin1、PTEN诱导激酶1（PTEN-induced putative kinase 1，PINK1）、帕金蛋白（Parkin）、p62]相关蛋白和基因的表达水平。通过PGC-1α siRNA转染实验进一步观察通脉养心丸对PGC-1α沉默表达后H9c2细胞上述功能的影响。结果 与模型组比较，通脉养心丸显著增强线粒体ATP合成（P＜0.01），改善线粒体膜电位（P＜0.01），减少氧化应激（P＜0.01），并促进线粒体自噬（P＜0.05、0.01）。沉默PGC-1α后，通脉养心丸对线粒体功能的保护作用显著削弱（P＜0.05、0.01）。结论 通脉养心丸能够通过PGC-1α调控线粒体的生物合成、动态平衡及自噬，从而有效改善MI/RI引发的线粒体功能紊乱与形态结构变化。揭示了PGC-1α作为线粒体功能调控的核心因子，其在MI/RI中的潜在治疗价值，为MI/RI的治疗提供了新的思路。

Objective To elucidate how Tongmai Yangxin Pill (通脉养心丸, TMYX) confers cardioprotective effects during myocardial ischemia/reperfusion injury (MI/RI) through the regulation of peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α)-mediated mitochondrial function. Methods A hypoxia-reoxygenation (H/R) model in cardiomyocytes was established. After intervention with TMYX, intracellular reactive oxygen species (ROS) levels were measured using DCFH-DA and MitoSOX^TM red fluorescent probes. Cellular adenosine triphosphate (ATP) content was quantified using an enhanced ATP assay kit. Changes in mitochondrial membrane potential were evaluated by JC-1 staining. Mitochondrial respiratory function was assessed using Seahorse XF cell mitochondrial stress test. Mitochondrial morphology and ultrastructural alterations were examined using MitoTracker staining and transmission electron microscopy. Western blotting and qRT-PCR were used to determine the expression levels of proteins and genes associated with mitochondrial fusion and fission [mitofusin 1 (Mfn1), mitofusin 2 (Mfn2), fission 1 (Fis1), dynamin-related protein 1 (Drp1)], biogenesis [nuclear respiratory factor 1 (Nrf1), mitochondrial transcription factor A (TFAM), mitochondrial DNA (mtDNA) copy number] and autophagy [Beclin1, PTEN-induced putative kinase 1 (PINK1), Parkin, p62]. To further investigate the effects of TMYX on the above-mentioned functions of H9c2 cells after PGC-1α silencing expression through PGC-1α siRNA transfection experiment. Results Compared with model group, TMYX significantly enhanced mitochondrial ATP synthesis (P < 0.01), improved mitochondrial membrane potential (P < 0.01), reduced oxidative stress (P < 0.01), and promoted mitochondrial autophagy (P < 0.05, 0.01). After silencing PGC-1α, the protective effect of TMYX on mitochondrial function was significantly weakened (P < 0.05, 0.01). Conclusion TMYX could effectively improve mitochondrial dysfunction and morphological changes induced by MI/RI by regulating mitochondrial biosynthesis, dynamic balance and autophagy through PGC-1α. This study reveals PGC-1α as a core factor regulating mitochondrial function and its potential therapeutic value in MI/RI, providing new ideas for the treatment of MI/RI.

R285.5

国家自然科学基金青年科学基金资助项目（82104565）；天津中医药大学组分中药国家重点实验室青苗基金资助项目（QMJJ202403）