GM-CSF prevents the externalization of phosphatidylserine through a PI3-kinase–dependent mechanism.

Freshly isolated human neutrophils were cultured in the presence or absence of GM-CSF (10 ng/mL) or TNF-α (200 U/mL) with or without LY294002 (10 μM) for 20 hours. Cells were harvested, and PS exposure was quantified by flow cytometry as described in the “Materials and methods” section. (A-D) Representative fluorescent histograms of 10 000 events showing the level of AnV binding at 20 hours for (A) control, (B) GM-CSF–, (C) LY294002-, and (D) GM-CSF + LY294002–treated cells. AnV staining is a marker of PS exposure on the cell outer plasma membrane. GM-CSF inhibited AnV staining, indicative of an inhibition of neutrophil apoptosis. The survival effect was abolished by coincubation with LY294002. (E) Quantitation of the percentage AnV high-staining cells at 20 hours following incubation with GM-CSF, TNF-α, or LY294002, or all of them, as indicated. Data represent percentage AnV high cells from 8 independent experiments, each performed in triplicate, together with mean ± SEM values. **P < .005 compared with control, unless otherwise indicated.