Fig. 5: Effect of AMPK inhibitor on S. Typhimurium-induced autophagy.

HeLa cells pretreated with CC (1 μM) for 30 min were left uninfected or infected with S. Typhimurium (10 MOI). After incubation for 2 h, the cell lysates were prepared and analyzed for the levels of p62 and LC3 and for the phosphorylation of AMPK^T172, ULK1^S555, ULK1^S317, and ACC^S79 (a) by western blot with the indicated antibodies or the antibodies against total proteins after stripping. Relative protein and phosphorylation levels were analyzed by quantification of the density of the protein bands with NIH Image-J software and presented as bar graphs. *p < 0.05, **p < 0.01, compared to uninfected control; ^#p < 0.05, ^##p < 0.01, compared to S. Typhimurium infection. b, c CC inhibits S. Typhimurium-induced formation of autolysosomes. HeLa cells stably expressing the GFP-RFP-LC3 gene were left uninfected or infected with S. Typhimurium (10 MOI) for 2 h in the absence or presence of CC (1 μM). The cells were then fixed and stained with DAPI. Autophagosomes represented by the orange puncta and autolysosomes represented by the red puncta were visualized under a confocal microscope (b) and statistically analyzed (c). Bar length: 20 μm. *p < 0.05, **p < 0.01, compared to uninfected control; ^#p < 0.05, ^##p < 0.01, compared to S. Typhimurium infection