Mitochondria are important sites for the production of ATP and the generation of ROS in cells. However, whether acute hypoxia increases ROS generation in cells or affects ATP production remains unclear, and therefore, monitoring the changes in ATP and ROS in living cells in real time is important. In this study, cardiomyocytes were transfected with RoGFP for ROS detection and MitGO-Ateam2 for ATP detection, whereby ROS and ATP production in cardiomyocytes were respectively monitored in real time. Furthermore, the oxygen consumption rate (OCR) of cardiomyocytes was measured. Similar results were produced for adult and neonatal rat cardiomyocytes. Hypoxia (1% O₂) reduced the basal OCR, ATP-linked OCR, and maximal OCR in cardiomyocytes compared with these OCR levels in the cardiomyocytes in the normoxic group (21% O₂). However, ATP-linked OCR, normalized to maximal OCR, was increased during hypoxia, indicating that the electron leakage of complex III exacerbated the increase of ATP-linked oxygen consumption during hypoxia and vice versa. Combined with the result that cardiomyocytes expressing MitGO-Ateam2 showed a significant decrease in ATP production during hypoxia compared with that of normoxic group, acute hypoxia might depress the mitochondrial oxygen utilization efficiency of the cardiomyocytes. Moreover, cardiomyocytes expressing Cyto-RoGFP or IMS-RoGFP showed an increase in ROS generation in the cytosol and the mitochondrial intermembrane space (IMS) during hypoxia. All of these results indicate that acute hypoxia generated more ROS in complex III and increased mitochondrial oxygen consumption, leading to less ATP production. In conclusion, acute hypoxia depresses the mitochondrial oxygen utilization efficiency by decreasing ATP production and increasing oxygen consumption as a result of the enhanced ROS generation at mitochondrial complex III.

线粒体是细胞中产生 ATP 和产生 ROS 的重要场所。然而，急性缺氧是否会增加细胞中 ROS 的产生或影响 ATP 的产生尚不清楚，因此，实时监测活细胞中 ATP 和 ROS 的变化很重要。在这项研究中，用 RoGFP 转染心肌细胞进行 ROS 检测和 MitGO-Ateam2 进行 ATP 检测，从而分别实时监测心肌细胞中的 ROS 和 ATP 产生。此外，测量了心肌细胞的耗氧率（OCR）。对成年和新生大鼠心肌细胞产生了类似的结果。缺氧 (1% O ₂) 与常氧组 (21% O ₂）。然而，标准化为最大 OCR 的 ATP 相关 OCR 在缺氧期间增加，表明复合物 III 的电子泄漏加剧了缺氧期间 ATP 相关耗氧量的增加，反之亦然。结合表达 MitGO-Ateam2 的心肌细胞与常氧组相比，缺氧期间 ATP 产生显着降低的结​​果，急性缺氧可能会降低心肌细胞的线粒体氧利用效率。此外，表达 Cyto-RoGFP 或 IMS-RoGFP 的心肌细胞在缺氧期间显示出胞质溶胶和线粒体膜间隙 (IMS) 中 ROS 的产生增加。所有这些结果表明，急性缺氧会在复合体 III 中产生更多的 ROS，并增加线粒体耗氧量，导致 ATP 产生减少。总之，