IL-3 is crucial for IgE-induced mast cell survival. (A) Inhibition of IgE(-Ag)–induced survival by anti–IL-3. The survival assay was performed with 1 μg/mL IgE using WT BMMCs at day 3 of culture in the presence of graded amounts of anti–IL-3, anti–IL-2Rα, or anti–IL-4. (B) Inhibition of IgE(-Ag)–induced survival by anti–IL-3 throughout the culture periods. Survival assays were performed with 1 μg/mL IgE for 4 days in the presence or absence of 1 μg/mL anti–IL-3 (▴) and 10 μg/mL anti–IL-4 Abs (▵). ○ indicates absence of IgE and blocking Abs; ⬡, presence of IgE alone. (C) The levels of IL-3 on stimulation by IgE(-Ag) and IgE(+Ag). WT BMMCs were cultured with 1 μg/mL IgE (IgE; □) or 10 ng/mL DNP-HSA after sensitization with 10 μg/mL IgE at 4°C for 1 hour (IgE + Ag; ▪). At the times indicated, the concentration of IL-3 in the culture supernatants was determined by high-sensitivity ELISA. (D) BMMC survival by exogenous IL-3. WT BMMCs were cultured in the presence of graded amount of recombinant IL-3 for 3 days. (E) Induction of IL-3 mRNA on IgE(-Ag) stimulation. mRNA levels of IL-3 were determined by real-time RT-PCR and expressed as fold-induction over the value of IgE(+Ag) at 1 hour after normalization with the values for glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Data are expressed as means ± SD for triplicate assays. Similar results were obtained from at least 3 independent experiments.