Phagolysosomes undergo fragmentation instead of exocytosis. (A) IgG-opsonized beads in RAW cells tracked for 24 h. Shown are differential interference contrast (DIC) images. Boxes indicate areas enlarged in (i) and (ii). (B) Number of beads in macrophages over time and normalized to T0. Data are shown as mean ± SEM from three independent experiments with eight cells quantified per experiment. (C, D, and G) Macrophages engulfed filamentous mCherry-Lp for 7 h (C) or mRFP1–E. coli for 2 h (D and G) and then imaged. Dashed lines show cell contours. Main panels in C and D show the red channel in a rainbow scale; red and blue are the highest and lowest intensity levels, respectively. Smaller panels show the red and DIC channels for the first frame. (E and F) Total volume of phagosomes and PDVs per cell over time are shown as mean ± SEM of 3 independent experiments with 10–25 cells analyzed per experiment. Volumes were normalized to T0. (H and I) Quantification of the number of PDVs and intact phagosomes. Data are mean ± SEM of 3 independent experiments with 15 images quantified per time point. One-way ANOVA with Tukey’s test was used to compare each time point against T0 (*, P < 0.05). See corresponding Video 1, Video 2, and Video 3. Scale bars: 10 µm (main panels), 5 µm (insets).
