Fig. 3

DPD blocks autophagic flux in late phase; Transcriptomics and TEM imaging of DPD action on LN229MG A, B) U87MG and LN229MG cells expressing Ad-mCherry-GFP-LC3B were treated with 0 or 30 µM DPD for 48 h. Images were taken under a confocal microscope (A), scale bar = 10 μm. Ten cells were counted manually, and the average was taken to analyze the ratio of yellow-green spots (B). C, D) U87MG and LN229MG cells expressing GFP-LC3 were treated with 0 or 30 µM DPD or 100 nM BafAl for 48 h, and then stained with LysoTracker Red and imaged under a confocal microscope (C). The green spots in each cell were quantified by manually counting ten cells and taking the average value (D), scale bar = 10 μm. E) p62 protein levels after DPD or/and BafA1 treatment of GBM cells. F) An annotated map of the KEGG pathway differential genes after DPD treatment of LN229MG cells; those shown in red rectangular boxes are associated with cellular autophagy. G) Heatmap of selected genes with significant differences. H) Control mitochondria (black arrowheads) and lysosomes (blue arrowheads) in normal cells; a large number of damaged mitochondria (green arrowheads), mitochondrial autophagosomes (red arrowheads), and autophagic lysosomes (yellow arrowheads) were observed by TEM of drug-treated LN229MG cells after 48 h, scale bar = 1 μm