Figure 1

Ethanol-induced lipid accumulation is affected by ethanol metabolism and lysosome function. (A–D) AML12 cells were treated with ethanol for 24 hours in the presence or absence of 4-MP (A,B), or CQ (C,D). Intracellular levels of TG (A,C) and cholesterol (B,D) were then determined. (E) AML12 cells were incubated with ethanol (24 h) with or without CQ, and then stained with 1 µM Bodipy-581/591 for lipid droplets (LD), which were quantified. (F) AML12 cells were cultured on coverslips in 24-well plates, infected with adenoviral GFP-LC3 for 24 h, and then treated with ethanol for 24 hours with or without CQ. The number of GFP-LC3 puncta were quantified. Boxed areas are enlarged in the inserts, showing the GFP-LC3 puncta. (G,H) Cells were cultured with ethanol for 12 hours (G) or 24 hours (H) with or without CQ. Cell lysates were examined by immunoblotting for SQSTM1 and LC3. Densitometry was conducted, normalized to β-actin and expressed as fold of the control level. In all experiments, CQ treatment was only for the last three hours of the culture. Data represent mean ± SEM. ^* P < 0.05, ^** P < 0.01, ^*** P < 0.001.