The human recombinant histamine-releasing factor (HrHRF) was previously shown to induce histamine release from human basophils from a subset of donors. The ability of HrHRF to directly induce histamine release from only certain basophils was thought to involve interaction between HrHRF and a particular kind of IgE, termed IgE(+), on the surface of these cells. Recent studies disproved the hypothesis that the IgE molecule or its high-affinity receptor, FcepsilonRI, is involved in secretion of histamine and cytokines by basophils stimulated with HrHRF. Rather, data suggest that HrHRF is a cytokine that stimulates basophils by binding to a cell-surface structure other than the IgE molecule. This report describes the effects of HrHRF on another inflammatory cell type: eosinophils from mildly allergic donors. In purified eosinophils primed with granulocyte-macrophage colony-stimulating factor, both tumor necrosis factor alpha (TNF-alpha) and HrHRF induced increased secretion of interleukin (IL) 8. In addition, both HrHRF and IL-5 enhanced secretion of IL-8 stimulated by TNF-alpha. Secretion of IL-8 reached a plateau level in less than 24 hours, was inhibited by cycloheximide, and required the presence of HrHRF throughout the culture period. In some eosinophil preparations, HrHRF induced calcium mobilization that was inhibited by pertussis toxin. Additionally, HrHRF caused secretion of IL-8 from the human eosinophilic cell line, AML14-3D10, which does not possess the alpha chain of FcepsilonRI. These data provide evidence that HrHRF contributes to activation of eosinophils and thus suggest an additional role for HrHRF in the pathophysiologic mechanisms of allergic disease.