The spectrum of eosinophil functions has expanded from fighting helminths to multiple novel roles in malignancy, infection, cancer, and metabolism. In asthma, glucocorticoids, prostaglandins (PG), and cysteinyl-leukotrienes (LT) regulate eosinophil biology through separate signaling pathways. Here we’ve evaluated the complex interplay between Dexa, PGE2, and CysLTs in eosinopoiesis and eosinophil biology in an allergic asthma model. Methodology: We used different inbred mouse strains to probe the interactions between these agents in eosinophil differentiation and maturation in bone marrow culture. Flow cytometry and histological analyses evaluated eosinophil precursor proliferation, maturation, and VLA-4 expression. The in vivo function of eosinophils was assessed by their in vivo migration into allergen-challenged sites. Results: Eosinophil production in IL-5-stimulated bone marrow cultures is enhanced by dexamethasone but suppressed by PGE2, which triggers eosinophil apoptosis via inducible NO synthase (iNOS). Dexamethasone-primed cultures contain mostly immature eosinophils; by contrast, dexamethasone associated with PGE2 leads to the production of mature eosinophils (without inducing eosinophil apoptosis), by a mechanism independent of iNOS. Interaction between dexamethasone and LTD4 in culture produces mostly mature eosinophils expressing VLA-4, capable of migration into the lungs in ovalbumin-sensitized and -challenged mice. Discussion: The combination of dexamethasone and either PGE2 or LTD4 – both mediators of allergic inflammation – supports the maturation of eosinophils (overcoming the maturation blockade observed with dexamethasone alone), which are functional in an in vivo model of asthma.