Figure 3.

Sustained GM‐CSF signaling required for GM‐CSF‐mediated neutrophil survival. (A–D) Freshly isolated neutrophils were treated with the indicated concentration of GM‐CSF and the percentage of apoptotic cells analyzed after 20 hr culture by flow cytometry. (B) Neutrophils were pretreated with increasing concentrations of CAM‐3001 for 20 min prior to GM‐CSF treatment. (C) Cells were maintained in media containing the indicated concentration of GM‐CSF or the culture media were replaced after 1.5 or 6 hr of treatment with GM‐CSF‐free media for the remainder of the incubation time. Media change alone did not impact apoptosis of control samples without GM‐CSF incubation (data not shown). (D) Human neutrophils were treated with GM‐CSF (1 ng/ml) and this was followed by addition of CAM‐3001 (1 μM) at the indicated time after GM‐CSF treatment began. The bars indicate the response to GM‐CSF and the effect of pretreatment with CAM‐3001 (20 min) prior to treatment with GM‐CSF. Data represents mean ± sem of n = 8 (A), n = 3 (B), n = 5 (C) or n = 3 (D) independent experiments