The MMP was measured using an MMP assay kit (Solarbio, Beijing, China). Cells were stained with the unique fluorescence probe JC-1 at 37 °C for 20 min, and then were washed twice with phosphate-buffered saline. The fluorescence intensity of the cells was observed using a flow cytometer (BD FACSCalibur, NJ, USA) and a confocal laser scanning microscope (Olympus, Japan). Then, the average fluorescence intensity of green monomers and red aggregates was determined, and the ratio was calculated.
MitoTracker™ was used to detect changes in mitochondrial dynamics. For mitochondrial labeling, the cell samples were incubated with a 100 nM solution of MitoTracker™ Green FM (Thermo Fisher Scientific, MA, USA) for 30 min at 37 °C, 5% CO2. Images were obtained using a Nikon confocal microscope system and camera (Nikon Instruments, NY, USA). The fluorescent dye and length of mitochondria were measured using Image J software. We selected 10 random fluorescence fields from each group.
To track mPTP opening, we treated HCAECs with tetramethylrhodamine ethyl ester, based on our previous study. The fluorescent signal of tetramethylrhodamine ethyl ester was determined using a Nikon confocal microscope system and camera.
MitoTracker™ was used to detect changes in mitochondrial dynamics. For mitochondrial labeling, the cell samples were incubated with a 100 nM solution of MitoTracker™ Green FM (Thermo Fisher Scientific, MA, USA) for 30 min at 37 °C, 5% CO2. Images were obtained using a Nikon confocal microscope system and camera (Nikon Instruments, NY, USA). The fluorescent dye and length of mitochondria were measured using Image J software. We selected 10 random fluorescence fields from each group.
To track mPTP opening, we treated HCAECs with tetramethylrhodamine ethyl ester, based on our previous study. The fluorescent signal of tetramethylrhodamine ethyl ester was determined using a Nikon confocal microscope system and camera.