Fig. 5

G-Rh2 represses starvation-induced autophagic flux. a HeLa and C-33A cells were treated with DMSO or G-Rh2 or autophagy regulators in the presence or absence of serum and were harvested at 24 h. Whole cell lysates were prepared and analyzed for p62 and LC3, tubulin served as the loading control. b HeLa and C-33A cells were transiently transfected with mRFP- GFP-LC3 and cells were treated with DMSO or G-Rh2 in the presence or absence of serum and harvested for 24 h. At the end of treatment, cells were observed for the change of both green and red fluorescence using a confocal microscope. c Acidic vesicular organelles were examined by incubating HeLa and C-33A cells with acridine orange followed by fluorescence microscopy. d After HeLa and C-33A cells were treated with DMSO or G-Rh2, the expression of CTSB was determined by western blotting