Fig. 3.

Effect of punicalagin on autophagy. (A–B) The LC3-I and LC3-II levels in HeLa or SiHa cells treated with 140 or 90 μM punicalagin for 2, 4, and 6 h (A) or the indicated concentrations of punicalagin for 6 h (B) were determined using western blotting. (C) The LC3-I and LC3-II levels in HeLa or SiHa cells pretreated with 100 μM chloroquine for 1 h, followed by treatment with 140 or 90 μM punicalagin for 6 h were examined using western blotting. (D) HeLa cells were infected with AdPlus-mCherry-GFP-LC3B adenovirus at the multiplicity of infection of 200 for 24 h, followed by pretreatment with 100 nM bafilomycin A1 for 1 h and treatment with 140 μM punicalagin for 24 h. The fluorescent images were captured using a confocal fluorescence microscope. The red and yellow LC3-II puncta were quantitatively analyzed in 100 cells per group and the results were reported as mean ± standard deviation. (E) HeLa cells treated with 140 μM punicalagin for 2 h were examined under a transmission electron microscope. The representative autophagosomes and autolysosomes are indicated by red and black arrows, respectively. ** P<0.01. All experiments were repeated three times.