A B S T R A C T Specific IgE anti-ragweed antibodies (IgEAR) were measured over two years in two groups of highly sensitive patients treated (immunized) with either ragweed extract or placebo and in a third group of placebo-treated, relatively insensitive patients. TheIgEAR on the patients' basophils were assessed by ragweed antigen E (AgE)-induced histamine release; blocking (IgG) antibodies were measured by their ability to inhibit AgE induced histamine release. These data were evaluated against the clinical severity of ragweed hay fever in each patient.

Immunization of BALB/c mice with alum-adsorbed OVA, followed by three bronchoprovocations with aerosolized OVA, resulted in the development of airway hyperresponsiveness (AHR) and allergic inflammation in the lung accompanied by severe infiltration of eosinophils into airways. In this murine asthma model, administration of monoclonal anti-IL-5 Ab before each Ag challenge markedly inhibited airway eosinophilia, but the treatment did not affect the development of AHR. Immunization and aerosol challenges with OVA following the same protocol failed to induce AHR in the mast cell-deficient W/Wv mice, but induced AHR in their congenic littermates, i.e., WBB6F1 (+/+) mice. No significant difference was found between the W/Wv mice and +/+ mice with respect to the IgE and IgG1 anti-OVA Ab responses and to the airway eosinophilia after Ag provocations. It was also found that reconstitution of W/Wv mice with bone marrow-derived mast cells cultured from normal littermates restored the capacity of developing Ag-induced AHR, indicating that lack of mast cells was responsible for the failure of W/Wv mice to develop Ag-induced AHR under the experimental conditions. However, the OVA-immunized W/Wv mice developed AHR by increasing the frequency and Ag dose of bronchoprovocations. The results suggested that AHR could be developed by two distinct cellular mechanisms. One would go through mast cell activation and the other is IgE/mast cell independent but an eosinophil/IL-5-dependent mechanism.

Both human and mouse c-kit ligand induced differentiation of human mast cells in a long-term culture ofthe mononuclear cells of umbilical cord blood. Growth factor activity for human mast cells present in conditioned medium of BALB/3T3 fibroblasts was due to mouse c-kit ligand. Recombinant c-kit ligand induced differentiation and proliferation of mast cell progenitors in early stages of culture. However, apparent selective growth of mast cells by c-kit ligand in cord blood cell cultures is mainly due to the effect of the cytokine to selectively maintain survival of immature mast cells. Electron microscopic analysis indicated that human mast cells developed by c-kit ligand were similar to human mast cells in the lung and gut mucosa, while those developed in coculture of cord blood cells with Swiss albino/3T3 fibroblasts were similar to skin mast cells. This conclusion was supported by the fact that the majority of mast cells developed by c-kit ligand contained only tryptase in their granules, whereas those developed in the cocultures contained both tryptase and chymase. It was also found that mast cells developed by c-kit ligand were immature even after culture for 14 weeks. Nevertheless, these cells express FceRI, and could be sensitized with human IgE for anti-IgE-induced release of histamine, prostaglandin D2, and leukotriene C4.Several murine cytokines, such as interleukin (IL)-3, IL4, IL-9, and IL-10 promote differentiation and proliferation of mouse mast cells (1-4). In contrast, reproducible growth of human mature mast cells had not been achieved until 1989, when we succeeded in developing morphologically and functionally mature human mast cells in a long-term coculture of mononuclear cells of cord blood with Swiss albino/3T3 fibroblasts (5). Subsequent studies revealed that the culture supernatant of 3T3 fibroblasts contained growth factors that promote differentiation of human mast cells (6). Human mast cell growth-promoting activity could be enriched by fractionation of the culture supernatant with ammonium sulfate precipitation and by ion-exchange chromatography. The molecular size of the factor was estimated by gel filtration to be between 70 and 100 kDa (7).While our studies were in progress, a cytokine, c-kit ligand, was characterized and molecular cloning of the cytokine has been accomplished by several groups of investigators (8)(9)(10)(11)(12). The present experiments show that both human and murine c-kit ligand induce differentiation of cord blood cells to human mast cells in culture and provide evidence that human mast cell growth-promoting activity in culture supernatants of 3T3 fibroblasts is associated with murine c-kit ligand. MATERIALS AND METHODSCell Cutures and Microscopic Examination. Mononuclear cells were obtained from heparinized umbilical cord blood (5) and suspended in RPMI 1640 medium (GIBCO) supplemented with 10%o fetal bovine serum (FBS; GIBCO), 50 ,M 2-mercaptoethanol, 2 mM L-glutamine, 100 units of penicillin per ml, 50 ,ug of streptomycin per ml, and 25 ,ug of gentami...

Effects of recombinant human interleukins on hematopoiesis were explored by using suspension cultures of mononuclear cells of human umbilical-cord blood and bone marrow. The results showed that interleukin 5 induced the selective differentiation and proliferation of eosinophils. After 3 weeks in culture with interleukin 5, essentially all nonadherent cells in both bone marrow and cord blood cell cultures became eosinophilic myelocytes. Culture of the same cells with interleukin 4 resulted in the selective growth of OKT3' lymphocytes. However, OKT3' cells did not develop if the bone marrow cells were depleted of OKT3'/OKT11+ cells prior to the culture, indicating that interleukin 4 induced the proliferation of a subpopulation of resting T cells present in cord blood and bone marrow cell preparations. In suspension cultures of bone marrow cells and cord blood cells grown in the presence of interleukin 3, basophilic, eosinophilic, and neutrophilic myelocytes and macrophages developed within 2 weeks. By 3 weeks, however, the majority of nonadherent cells became eosinophilic myelocytes. In contrast to mouse bone marrow cell cultures, neither interleukin 3 nor a combination of interleukins 3 and 4 induced the differentiation of mast cells in human bone marrow or cord blood cell cultures.