Figure 3

B82 activates ER stress in H460 cells and CHOP knockdown inhibits B82-induced H460 cell apoptosis. A and B. B82 induced CHOP expression in H460 (A) and HepG2 (B) cells. Cells were treated with B82 at the indicated concentrations for 12 h. Western blot results was calculated and represented as the percent of control (n = 3, **p < 0.01, vs. vehicle group). C. B82 could not induce CHOP expression in BEAS-2B cells. Cells were treated with B82 at the indicated concentrations for 12 h. The protein level of CHOP in total proteins was detected by western blot. D-E. B82 induced mRNA expression of GRP78 (D), ATF-4 (E), XBP-1(F), and CHOP (G) in H460 cells. Gene mRNA expression were examined by RT-qPCR after 6- or 12-h treatment of B82. H-K. Cells were treated with B82 at indicated concentrations for 24 h. The caspase-3 activity in H460 (H) and HepG2 (I) cells was assayed. The total protein in H460 (J) and HepG2 (K) cells was extracted and cleavaged caspase-3 was examined by Western blot. L-N. H460 cells were transfected with CHOP siRNA virus. Forty-eight hours after transfection, the CHOP- and EGFP-expressing cells were counted using fluorescent microscopy (L). Cells were treated with B82 (20 μM) for 12 h and then CHOP expression was determined by Western blotting (M). Cells were treated with B82 at indicated concentrations for 48 h, and the cell survival was determined using MTT assay, and the inhibitory rates were calculated as percent of DMSO-treated cells (N). * p < 0.05, ** p < 0.01, vs. vehicle control.