Human lung mast cells are enriched in the capacity to produce granulocyte-macrophage colony-stimulating factor in response to IgE-dependent stimulation

By using reverse transcription-PCR, in situ hybridization, enzyme-linked immunosorbent assay and immunocytochemistry, we have studied the production of granulocyte-macrophage colony-stimulating factor (GM-CSF) in human lung mast cells induced by cross-linkage of high-affinity Fc epsilon receptors (Fc epsilon RI). We have also confirmed the bioactivity of GM-CSF released from lung mast cells by investigating the effect of the supernatant from lung mast cells activated with anti-IgE on the release of eosinophil cationic protein (ECP) from eosinophils. Mast cells were purified using affinity magnetic selection with monoclonal antibody (mAb) YB5.B8 (93-99 % pure). Purified mast cells were precultured with IgE for 16 h before challenge with 1 mu g/ml anti-IgE with or without stem cell factor (SCF). Eosinophils were purified by immunomagnetic negative selection (> 98 % pure). The activation of mast cells via Fc epsilon RI enhanced the intensity of the GM-CSF signal within 2 h and the cells produced GM-CSF protein 4 h after the activation. In the absence of recombinant human (rh) SCF, anti-IgE induced a median GM-CSF response of 202 (<15 similar to 681) pg/10(6) mast cells/24 h, whereas in the presence of rhSCF the median IgE-dependent GM-CSF release was 356 (152 similar to 1216) pg/10(6) mast cells/24 h. This difference was statistically significant (p = 0.0029, n = 12). In contrast, mast cells produced only a small amount of GM-CSF in the absence of anti-IgE. The mast cell supernatant induced ECP release from eosinophils and the release was significantly inhibited by blocking mAb against GM-CSF. These findings indicate that human mast cells are an important cellular source of GM-CSF and as such may contribute to chronic eosinophil-mediated inflammation.