Fig. 4. BITC-induced LC3B conjugation in AGS cells was not dependent on class III PI3K. (A) For the single treatment, AGS cells were treated with 2.5, 5, 10, or 15 μM BITC for 24 h. For the combined treatment, the cells were pre-treated with 0.5 µM wortmannin for 2 h followed by 10 µM BITC for 24 h. Then, the whole cell lysate was prepared, and the class III PI3K protein level was measured by western blotting. β-actin was used as a loading control. (B) The cytotoxicity of wortmannin was detected by MTT assay after treating the cells with different concentrations of wortmannin for 24 and 48 h. (C) AGS cells were pre-treated with 0.25 or 0.5 µM wortmannin for 2 h followed by 10 or 15 µM BITC for 24 h. After cell lysis, the change in LC3B II level was detected by western blotting. β-actin was used as a loading control. Data were calculated from three independent experiments and expressed as the mean ± SEM. **p<0.01 and ***p<0.001 versus the control.
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