Fig. 4

The phagocytosis of the aged mouse peritoneal macrophages was associated with the autophagy. a Representative multi-channel fluorescence microscope images of mouse peritoneal macrophages phagocytosed the E. coli. The cells were incubated with EGFP-expressing E. coli (green) for 1 h, followed by washing with PBS, fixation with 4% paraformaldehyde, and staining for F-actin using phalloidin-Alexa Fluor 633 conjugate working solution (red) and with DAPI (blue). The blue DAPI channel was not shown here separately. White bar = 100 μm. The proportions histogram of the EGFP positive cells counted from the fluorescence microscope images was shown on the right side. b Representative flow cytometry analysis of the Young, Aged, ABT, Rapamycin, and 3-MA treated groups. The peritoneal macrophages were stained with F4/80-PE after co-incubation with EGFP-expressing E. coli. The isotype control group was not shown in the graph. Each group repeated at least for three times. The data in this panel represented the mean ± S.D. * represented P < 0.05 when compared to the Aged group. c-f The proteins Lc3, p62, Trem-2 and Tubulin-α of the peritoneal macrophages from aged mouse were exmained by western blotting after treated by 2.5 μM ABT-263, 100 μM 3-MA, and 1.0 μM Rapamycin for 24 h respectively. The data in the figures d, e, f represented the mean ± S.D. Significant differences between groups were indicated as * P < 0.05 compared to the Ctrl group, n = 4