Figure 6

Butyrate limited replication of cytosolicS. Infantis by inhibiting autophagy. A Western blot analysis of Caco-2 cells treated with siRNA-targeting ATG5 (mean ± SEM, n = 3), ***P < 0.001. B Western blot analysis for the expression levels of ZO-1 and LC3 proteins. The right panel shows the quantitation analysis by ImageJ software. (mean ± SEM, n = 3), **P < 0.01, ***P < 0.001. C Detection of the distribution and expression of ZO-1 (Red) of Caco-2 cells by immunofluorescence staining. Blue: DAPI, Scale bar, 10 µm. D Cell death was measured using Calcein-AM/PI assay. E Effect of siATG5 on intracellular S. Infantis was assessed by the gentamicin protection assay. F The number of infected cells containing 1 to 10, 10 to 20, 20 to 50, or more than 50 bacteria per cell at 8 hpi was measured using a fluorescence microscope. At least 100 cells were counted for each group and the percentage of cells with the different numbers of bacteria is presented (mean ± SEM, n = 3), *P < 0.05, **P < 0.01. G Caco-2 cells were infected with S. Infantis-pFPV. Cells at 8 hpi were stained with LAMP-1 (Red) to label S. Infantis in SCV, while the cytosolic S. Infantis was green. “Zoom” denotes a magnified image of the white dotted area. Green: S. Infantis. Blue: DAPI. Scale bar, 10 µm.