Fig. 8

Pinocembrin-induced mitochondrial autophagy is mediated by the JNK/ERK/BNIP3/LC3 pathway. a Whole-cell proteins from BV-2 cells were harvested immediately after different treatments. Immunoprecipitation was performed by an anti-LC3 antibody and was immunoblotted for BNIP3 and LC3. b Representative western blotting images of ubiquitin and P62 of BV2 cell mitochondrial fraction. c Microglia were double-stained with BNIP3 and TOM20 (MitoTracker). Representative micrographs for BNIP3 (red), TOM20 (green), DAPI (blue), and merged pictures (yellow) from each group were captured by a confocal microscope. Bar = 50 μm. d The immunocytochemistry of BV2 microglial cells was conducted with anti-BNIP3 and anti-LC3 antibodies. Confocal microscopy revealed that the SP600125 and FR 108204 groups had few positive staining of autophagosome marker LC3 compared with the pinocembrin-only group, as well as less co-localizing with BNIP3. Bar = 50 μm. Data are representative of at least 3 independent experiments and presented as mean ± SEM. ^†p < 0.05 vs. the NA group; ^#p < 0.05 vs. the IH group; *p < 0.05 vs. the IH + PIN group; **p < 0.01 vs. the IH + PIN group