Figure 6

Coating with AIM enhanced debris engulfment by F4/80 positive, M1- and M2a-like macrophages. (a) Peritoneal cells derived from zymosan model AIM ^−/− mice. Phagocytic assay using cells collected from the peritoneum of zymosan model mice on day 7. The proportion of engulfment of FVD780-positive cell debris within DAPI⁻Mac-1⁺F4/80⁺Gr-1⁻ cells and DAPI⁻Mac-1⁺F4/80⁻Gr-1⁺ cells was assessed by flow cytometry. Precise methods are described in the Methods section. F4/80 positive macrophages had a greater phagocytic ability than Gr-1 positive neutrophils at 10 and 30 min. F4/80 positive macrophages had a significantly higher engulfing ability for rAIM coated dead cell debris than for non-coated dead cell debris. n = 3 dishes for each group. Confocal microscopic images of engulfment by F4/80 positive macrophages (red) of labeled dead cell debris (green) with and without a coating of rAIM. Phagocytosis by F4/80 positive macrophages was enhanced by coating the surface of the debris with rAIM. Yellow colors (merge) indicates that the dead cell debris engulfed by the macrophages (red). The arrows indicate phagocytosis. Supplementary Videos 1 and 2 provide more precise images of phagocytosis by M1- or M2a-like macrophages. Scale bars, 25 μm. (b,c) M1- and M2a-like macrophages modified from bone marrow cells. (b) M2a-like macrophages from AIM ^+/+ mice phagocytosed dead cell debris to a greater extent than M1-like macrophages at 10 and 30 min after incubation. At 10 min, M2a-like macrophages from AIM ^−/− mice phagocytosed dead cell debris to a greater extent than M1-like macrophages. n = 3 dishes for each group. (c) Phagocytosis by both M2a- and M1-like macrophages from AIM ^−/− mice at 30 min was enhanced by coating the surface of the dead-cell debris with rAIM. Control: Dead cell debris was not incubated, rAIM: Dead cell debris was coated with rAIM. n = 3 dishes for each group.