Figure 3

GSK2578215A induces mitophagy. (a–g) SH-SY5Y cells were co-transfected with pDsRed2-mito and GFP-LC3 plasmids and 24 h later treated with GSK2578215A. Confocal microscopy pictures showing the mitochondrial morphology (a and d) and the localization of GFP-LC3 vesicles (b and e). In (f), arrows indicate the colocalization of both proteins. (g) Bar diagram representing the distance between mitochondria and autophagosomes in treated and untreated control cells at the times indicated. (h) The inhibition of mitochondrial fission by mdivi-1 blocks GSK2578215A-induced autophagy in SH-SY5Y cells. At 24 h after transfection with GFP-LC3, SH-SY5Y cells were treated with 1 nM GSK2578215A in the presence or absence of 10 μM mdivi-1, and 12 h later, the percentage of autophagic cells was determined. Nuclei were stained with Hoechst. Data shown in the bar graph are the mean±S.E.M. of at least three independent experiments, each performed in triplicate. Statistical significance was determined by a two-tailed Student’s t-test: *P<0.05 and ***P<0.001. Scale bar, 10 μm