Figure 5.

4-phenylbutyric acid (4-PBA) inhibits SiNP-induced ER stress and autophagy. (a, b) L-02 cells incubated with 25 μg/ml SiNPs were treated with (+) or without (–) 5 mM 4-PBA (ER stress inhibitor) for 12 h before harvesting at 24 h. The L-02 cells that were treated with 300 nM thapsigargin (Tg, a chemical known to rapidly elicit ER stress) for 12 h were used as the positive control. Thereafter, the cells were harvested and the expressions of HSPA5, ATF4, DDIT3, LC3B, and SQSTM1 were analyzed by western blot using the indicated antibodies. Blots are representative of 3 independent experiments. ACTB was used as a sample-loading control. Densitometric HSPA5:ACTB, ATF4:ACTB, DDIT3:ACTB, LC3B-II;ACTB, and SQSTM1;ACTB ratios from at least 3 independent experiments are shown. The value of control without any treatment was set at 1 for each experiment (*, p < 0.05). (c) The LC3B distribution patterns in (a, b) cells were analyzed using indirect immunofluorescence with anti-LC3B antibodies and imaged by LSCM. LC3B (green) and DAPI staining of the nuclei (blue) are shown. Scale bar: 25 μm. The number of punctate LC3B per cell in the untreated control and SiNP-treated cells is shown. Data are expressed as mean ± SD. ··· p · 0.05 versus the control.