Figure 2

Inhibition of autophagic flux was revealed in Pb-exposed rPT cells. (a and b) Cells were transiently transfected with GFP-LC3 plasmid, then treated with 0.5 μM Pb for 12 h in the presence or absence of CQ (50 μM, for the last three hours), or 3-MA (2 mM, for the entire course). Representative confocal images are present in a and the number of GFP-LC3 dots per cell (b) was quantified (mean±S.E.M., n=4). (c and d) Cells were treated with 0.5 μM Pb for 12 h in the presence or absence of 50 μM CQ (for the last three hours), or 2 mM 3-MA (for the entire course); then harvested and analyzed for protein levels of LC3 (c) or p62 (d) using western blotting. Upper panels: representative western blot images; lower panels: quantitative western blot analysis (mean±S.E.M., n=4). (e) Cells were transiently transfected with GFP-RFP-LC3 plasmid, treated with 0.5 μM Pb for 12 h or incubated with EBSS medium for 2 h, or treated with 50 μM CQ for 3 h. Puncta showing both green and red fluorescence (indicated as yellow), or showing only the red fluorescence are shown in the representative confocal images of four groups. ns, not significant, **P<0.01