GM-CSF and TNF-α promote neutrophil survival through a PI3-kinase–dependent mechanism, as assessed by cell morphology.

Peripheral blood 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 the time periods indicated. Cells were harvested at 6 and 20 hours, and cytospins were fixed and stained as detailed in the “Materials and methods” section. (A-D) Representative photomicrographs (original magnification, × 400) of neutrophils incubated with or without GM-CSF or LY294002 for 20 hours. (E-F) TNF-α caused an early (6-hour) increase and a late (20-hour) inhibition of neutrophil apoptosis, whereas GM-CSF caused a significant survival effect at both time points. LY294002 had no effect on the constitutive rate of neutrophil apoptosis or the early proapoptotic effect of TNF-α, but it inhibited the survival effect of GM-CSF and TNF-α at 20 hours. Data represent percentage apoptosis from 8 independent experiments, each performed in triplicate, together with mean ± SEM values. *P < .05 and **P < 0.005 compared with control values, unless otherwise indicated.