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Basophils and mast cells play a crucial role in immunological and allergic processes due to the release of inflammatory mediators such as histamine. It has been suggested for a long time that the hist

Recent studies have shown that human basophils, like mast cells, generate interleukin (IL)-4 following immunological activation and may thus participate in late-phase allergic and inflammatory processes. Here, we report the capacity of human basophils to release IL-13 within 24 h following stimulation with anti-IgE. Additionally, in 14 out of 31 experiments, we observed that basophils rapidly release performed IL-4 within 5-10 min, as well as newly generated IL-4, which was released 4 h following stimulation of the cells with anti-IgE. In contrast to the biphasic release of IL-4 from the cells, no preformed IL-13 was detected at earlier times (5-30 min). Preformed IL-4 and IL-4 and IL-13 generated de novo were also released after stimulation of the cells with IL-3; an enhanced production of these cytokines was observed using a combination of IL-3 and anti-IgE. We conclude from these data that, by releasing performed IL-4 and IL-4 and IL-13 generated de novo, human basophils may be centrally involved in the orchestration of allergic inflammation by providing a trigger to IL-4-mediated T helper 2 lymphocyte activation, B cell IgE switching, and increased vascular adhesion molecule expression.

Objective and Design: We report a method for basophil purification from buffy coats, which avoids positive selection of the cells and gives rise to good purity, yield and functional integrity of the cells.¶Subjects: Buffy coat blood (concentrated leukocyte fraction derived from 450 ml venipuncture donations) obtained from healthy blood donors (n = 51).¶Methods: Basophils were enriched by a three-step process starting with Ficoll density centrifugation (1.6 ± 0.1% basophil purity) followed by counter current centrifugal elutriation (17.7 ± 1.4% basophil purity). The final stage involved negative selection using Dynal immunomagnetic beads directed against CD2, CD14, CD16 and CD19 positive cell contaminants. Functional integrity of which was assessed by comparing the anti-IgE or calcium ionophore A23187 induced histamine release from basophils obtained from each enrichment step. Furthermore, basophil morphology was investigated using light and electron microscopy.¶Results: The final mean basophil purity of 67.3 ± 1.4% with a yield of 3.5 ± 0.5 × 106 basophils and a recovery of 21.8 ± 2.4% was achieved. Net histamine release from basophils stimulated with optimal concentrations of anti-human IgE was 39.1 ± 6.5% after Ficoll centrifugation, 41.6 ± 7.7% following elutriation and 35.7 ± 6.8% from the final purified fraction. Additionally, basophils enriched with our method showed intact morphology by electron microscopy and were functionally active to non-immunological stimulation.¶Conclusions: These results compare favourably with previous studies, which have often required the use of positive selection via the Fc RI receptor, which may result in cell degranulation, or cell sorting, which cannot be applied to large cell numbers. Our method provides a reproducible technique for basophil enrichment when large numbers of functionally intact basophils are required.

The elevated interleukin (IL)-4 and IgE production in Schistosoma mansoni infection seems to be induced essentially by the egg stage of the parasite. The underlying mechanism, however, is not known. Since basophils from human peripheral blood can produce IL-4, we asked, whether soluble S. mansoni egg antigens (SEA) would trigger basophils to release IL-4. Basophils from healthy human donors (n = 32) without prior history of schistosomiasis were incubated with SEA in the presence of IL-3. In all donors, IL-4 was produced at different concentrations. The IL-4 production was dependent on the dose of SEA, was correlated with the purity of the basophil preparation, and the IL-4 concentration in the culture supernatant was maximal 5 h after stimulation with SEA. In addition to its IL-4-stimulatory effect, SEA triggered basophils to degranulate, thereby releasing histamine and sulfidoleukotrienes. Stripping of receptor-bound IgE from basophils inhibited both SEA- and anti-IgE-induced, but not ionomycin-induced IL-4 production. Moreover, resensitization of stripped basophils with stripping supernatants or human serum restored SEA-induced IL-4 production. This suggests that IgE is involved in the mechanism of IL-4 induction by SEA. Since IL-4 is induced in basophils from nonexposed donors, basophils may play a role as an early source of IL-4 in S. mansoni infection.