Fig. 2: The effect of alcohol on the levels of two autophagy markers, LC3B and p62, in Huh7 cells.

a The effect of alcohol on mRNA expression of LC3 and p62. Huh7 cells were incubated with or without alcohol at indicated concentrations for 96 h. The levels of LC3 and p62 mRNA, with normalization to the corresponding GAPDH mRNA level, are expressed as the fold of control (without alcohol treatment, which was defined as 1). b The dose effect of alcohol on protein levels of LC3B and p62. Huh7 cells were treated with or without alcohol at indicated concentrations for 96 h. A representative western blot image shows the protein levels of LC3B-I, LC3B-II, and p62 in alcohol-treated or untreated control cells. The densitometric intensities of western blot bands were quantified by Image J software. The relative ratios LC3B-II/β-actin, LC3B-II/LC3B-I, and p62/β-actin were calculated and shown as the fold of control (without alcohol treatment, which was defined as 1). c The time-course effect of alcohol on protein levels of LC3B and p62. Huh7 cells were treated with or without alcohol (80 mM) for different times (0, 48, 96, and 144 h). A representative western blot image shows the protein levels of LC3B-I, LC3B-II, and p62. The relative ratios of LC3B-II/β-actin, LC3B-II/LC3B-I, and p62/β-actin were calculated and shown as the fold of control (without alcohol treatment, which was defined as 1). d Huh7 cells expressing GFP-LC3 protein were treated with alcohol at indicated concentrations for 96 h. GFP-LC3-positive dots per cell were determined by fluorescence microscope (30 cells were counted per experiment). Scale bar: 50 µm. The data are the mean ± SD of the results of three independent experiments. *p < 0.05, **p < 0.01