Fig. 2

SAL promotes mitophagy in somata and neurites. A Images of mito-mKeima expressing SH-SY5Y cells treated with SAL or PBS (CTL). 440 nm excitation (green) labels the healthy mitochondria, whereas 589 nm excitation (red) represents the mitochondria delivered to lysosomes for mitophagy. B Quantification of mitophagy index (ratio of fluorescence intensity of λEx = 589 nm against λEx = 440 nm) in A. Data from at least 7 random images in each condition from 3 independent experiments. C Immunoblotting shows the levels of indicated proteins extracted from cytosol (left panels) and mitochondria (right panels) of SH-SY5Y cells treated with or without SAL. GAPDH and VDAC as cytosolic (Cyto) and mitochondrial (Mito) fraction in C. D Quantification of band intensity in C. Data from 3 independent experiments. E IF images of cortical neurons transduced with AAV particles expressing mitoGFP, with indicated treatments. The puncta are labeled with autophagic marker LC3 (red) and mitochondrial GFP (green) and shown in upper panels. Images in white rectangles are amplified and shown in lower panels. F Quantitation shows the number of puncta co-labeled with LC3 and MitoGFP. G, H As described in E and F, except images of neurites are shown and quantified. Error bars indicates mean ± SD from at least 20 cells/neurites from 3 independent experiments (n ≥ 20). *P < 0.05, **P < 0.01, ***P < 0.001; one-way ANOVA test. Scale bar, 10 μm