Activation of TRPML1 protects PA-induced mitochondrial damage. (A) Effects of ML-SA5 treatment on PA-induced mitochondrial fragmentation. Mitochondria were fluorescently labeled with MitoTracker. (B) Quantitation of the effects of ML-SA5 treatment as shown in A. Average and individual results are shown. The relative mitochondrial length was calculated by normalizing the mitochondrial length under each condition with that in the control group. (C) Effects of ML-SA5 treatment on PA-induced mitochondrial fragmentation. Mitochondria were immunolabeled with an anti-Tomm20 antibody. Images in the left and right panels were taken with confocal and SIM microscopes, respectively. (D) The effects of ML-SA5 co-treatment (10 μM) on PA (100 μM, 6 h)-induced changes on mitochondria; membrane potential assessed with JC-1 labeling. (E) Summary of ML-SA5 effects on PA-induced mitochondrial fragmentation, as shown in panel C (SIM images) and quantified in terms of average mitochondrial length. For each experimental condition, ≥50 randomly selected cells from at least three independent repeats were analyzed. Mitochondrial length was measured and normalized to that of the control group. (F) Quantitation of ML-SA5 treatment effects on mitochondria membrane potential assessed by JC-1 labeling. Normalized mitochondrial membrane potentials were calculated by normalizing the ratio of red versus green signals to that in each control group. (G) Effects of ML-SA5 co-treatment (10 μM) or ML-SA8 (3 μM) on PA (100 μM, 6 h)-induced changes in mitochondrial membrane potential assessed with a TMRE assay. (H) Quantitation of the results as shown in G. Scale bar = 10 μm. Data are presented as means ± SEMs; ***P < 0.001.