Figure 5
Figure 5. The presence of uPAR on the surface of either macrophages or neutrophils is required for enhanced phagocytosis of viable neutrophils by macrophages. (A) uPAR−/− macrophages do not demonstrate enhanced phagocytosis when combined with viable uPAR−/− neutrophils. A total of 106 viable WT or uPAR−/− neutrophils were added to uPAR−/− macrophages and phagocytosis assays were performed. ***P < .001, compared with phagocytosis with viable WT neutrophils. (B) Incubation of uPAR−/− neutrophils or macrophages, but not both, with suPAR restores the ability of uPAR−/− macrophages to phagocytose viable uPAR−/− neutrophils. Viable uPAR−/− neutrophils alone (neutrophil), uPAR−/− macrophages alone (macrophage), or both (Both) were preincubated with 1 μg/mL suPAR for 30 minutes. The cells were then resuspended in RPMI plus 5% FBS and added to WT macrophages for phagocytosis assays. Phagocytosis of viable uPAR−/− neutrophils preincubated with 1 μg/mL BSA by uPAR−/− macrophages preincubated BSA were used as a control. **P < .01, ***P < .001, compared with the control group treated with BSA. (C) WT and uPAR−/− macrophages demonstrate similar ability in ingesting carboxylate-modified beads. Carboxylate-modified beads (2 μm) were incubated WT or uPAR−/− macrophages for 1 hour. The phagocytic index was calculated as the percentage of macrophages that ingested beads. (D) suPAR did not enhance the uptake of beads by uPAR−/− macrophages. uPAR−/− macrophages were preincubated without or with 1 μg/mL suPAR, and phagocytosis performed as in panel C.

The presence of uPAR on the surface of either macrophages or neutrophils is required for enhanced phagocytosis of viable neutrophils by macrophages. (A) uPAR−/− macrophages do not demonstrate enhanced phagocytosis when combined with viable uPAR−/− neutrophils. A total of 106 viable WT or uPAR−/− neutrophils were added to uPAR−/− macrophages and phagocytosis assays were performed. ***P < .001, compared with phagocytosis with viable WT neutrophils. (B) Incubation of uPAR−/− neutrophils or macrophages, but not both, with suPAR restores the ability of uPAR−/− macrophages to phagocytose viable uPAR−/− neutrophils. Viable uPAR−/− neutrophils alone (neutrophil), uPAR−/− macrophages alone (macrophage), or both (Both) were preincubated with 1 μg/mL suPAR for 30 minutes. The cells were then resuspended in RPMI plus 5% FBS and added to WT macrophages for phagocytosis assays. Phagocytosis of viable uPAR−/− neutrophils preincubated with 1 μg/mL BSA by uPAR−/− macrophages preincubated BSA were used as a control. **P < .01, ***P < .001, compared with the control group treated with BSA. (C) WT and uPAR−/− macrophages demonstrate similar ability in ingesting carboxylate-modified beads. Carboxylate-modified beads (2 μm) were incubated WT or uPAR−/− macrophages for 1 hour. The phagocytic index was calculated as the percentage of macrophages that ingested beads. (D) suPAR did not enhance the uptake of beads by uPAR−/− macrophages. uPAR−/− macrophages were preincubated without or with 1 μg/mL suPAR, and phagocytosis performed as in panel C.

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