The total cellular and mitochondrial ROS contents in frozen heart sections were evaluated by DHE staining (10 μM, S0063, Beyotime, China) and staining with the fluorescent probe MitoSOX (100 mM, M36008, Thermo Fisher Scientific, USA), respectively, as previously described [21 (link)]. Images were captured with a confocal laser scanning microscope (Nikon, Japan) and analysed with ImageJ software (NIH, USA).
Total cellular ROS in NCMs were detected by a ROS/Superoxide Detection Assay Kit (Ab139476, Abcam, USA) as previously described via a microplate reader [22 (link)]. The mitochondrial ROS in NCMs were detected by staining with the fluorescent probe MitoSOX (100 mM, M36008, Thermo Fisher Scientific, USA) following the manufacturers’ protocols via flow cytometry analysis with a BD FACS Aria II flow cytometer.
The Total Glutathione Peroxidase (GPx) Assay Kit with NADPH (S0058, Beyotime, China) and Lipid Peroxidation Malondialdehyde (MDA) Assay Kit (Ab118970, Abcam, USA) were used according to the manufacturer’s instructions to detect cardiomyocyte GPx activity and MDA levels and further assess oxidative stress levels.
Total cellular ROS in NCMs were detected by a ROS/Superoxide Detection Assay Kit (Ab139476, Abcam, USA) as previously described via a microplate reader [22 (link)]. The mitochondrial ROS in NCMs were detected by staining with the fluorescent probe MitoSOX (100 mM, M36008, Thermo Fisher Scientific, USA) following the manufacturers’ protocols via flow cytometry analysis with a BD FACS Aria II flow cytometer.
The Total Glutathione Peroxidase (GPx) Assay Kit with NADPH (S0058, Beyotime, China) and Lipid Peroxidation Malondialdehyde (MDA) Assay Kit (Ab118970, Abcam, USA) were used according to the manufacturer’s instructions to detect cardiomyocyte GPx activity and MDA levels and further assess oxidative stress levels.