Figure 3. The difference in autophagy flux and cells sensitivity after H₂O₂ treatment between PIG1 and PIG3V cells.

(A) Immunoblots of LC3 in PIG1 and PIG3V cells treated with H₂O₂ (0.5 mM) for different hours. (B) Band intensity normalized to β-actin is expressed as mean ± SD, **p < 0.01. (C) PIG1 and PIG3V cells were transfected with adenovirus expressing mRFP-GFP-LC3. After a 24-hour transfection, cells were treated with or without H₂O₂ for 12 hours. As shown in merged confocal images, the yellow puncta indicate the autophagosome while the red puncta indicate the autolysosome. Scale bars represent 5 μm. (D) The number of autolysosome dots were counted on 40 cells in a minimum of 3 experiments and expressed as mean ± SD, **p < 0.01. (E) PIG1 and PIG3V cells were treated with H₂O₂ for 24 hours, and the level of apoptosis was detected by flow cytometry assay. (F) The level of apoptosis are presented as the mean ± SD, *p < 0.05. (G) PIG1 and PIG3V cells were treated with H₂O₂ for 24 hours, and the MMP was measured by flow cytometry assay. (H) Quantification of MMP are presented as the mean ± SD, *p < 0.05. (I) PIG1 and PIG3V cells were treated with H₂O₂ for 24 hours, then the intracellular ROS level were measured by flow cytometry assay. (J) Quantification of intracellular ROS level are presented as the mean ± SD, *p < 0.05.