TUDCA inhibits ceramide-induced ER stress. (A) ACC-M and ACC-2 cells were seeded into 60 mm culture dishes and the next day cells were pretreated with 5 mM 4-PBA or 1 mg/ml TUDCA. After 3 h incubation, 100 μM ceramide was added for further 12 h incubation in full medium. Total RNA was isolated for detection of XBP1 mRNA splicing using RT-PCR. The experiment was repeated three times and the representative result is shown. (B) Cells were treated as in A. After 5 mM 4-PBA or 1 mg/ml TUDCA treatment for 3 h, 100 μM ceramide was added for further 3 h incubation. Whole cell lysates were collected to analyze phosphorylated eIF2α expression. The experiment was repeated several times and the representative result is shown.