Human lung mast cell responsiveness to antigen is not governed by surface IgE expression

Anne-Marie Scola, Suzanne Havard, Linda J Kay, Peter T Peachell

University of Sheffield, Sheffield, UK

Mast cells have been implicated in allergic diseases such as asthma. Mast cells express IgE bound to high affinity IgE receptors (FcεRI). Cross-linking of IgE with multivalent antigen initiates the activation of these cells and leads to the generation and/or release of a wide variety of autacoids such as histamine. We have found that the release of histamine from human lung mast cells (HLMC), induced by anti-IgE is very variable ranging from 0 to 70% of the total histamine content. The aim of the study was to investigate the role of surface IgE in the variability of mast cell responsiveness.

Mast cells were obtained by physical and enzymatic disruption of grossly normal tissue from resected human lung. For histamine release, HLMC were challenged with a maximal releasing concentration of anti-human IgE (1:300) for 25 min. Histamine was measured by an automated fluorometric technique. For flow cytometry, mast cells were further purified over a Percoll density gradient. Cells were then stained with anti-c-kit-PE and either anti-IgE-FITC or isotype (goat IgG-FITC) control before fixing with 4% paraformaldehyde for 30 min. The c-kit positive cells were gated and the amount of fluorescence was quantified as Mean Fluorescence Index (MFI) which was calculated as the geometric mean of the anti-IgE-FITC signal subtracted by the geometric mean of the isotype control-FITC signal.

The response of HLMC preparations to anti-IgE is highly variable. Some preparations release a substantial proportion of their histamine content following stimulation whereas other preparations fail to release any, or very little, histamine. Out of a total of 600 mast cell preparations, 78 preparations released less than 5% of their total histamine content in response to a maximal releasing concentration (1:300) of anti-IgE. We investigated whether non-releaser mast cells might respond to IgE-directed stimulation following passive sensitization (20 h, 37 °C) with JW8-IgE, a chimeric IgE specific for the hapten 5-iodo-4-hydroxy-3-nitrophenyl (NIP). The non-releasing HLMC preparations were unable to respond to either antigen (NIP-HSA) or anti-IgE following passive sensitization. By contrast, good and moderate-releasing preparations responded effectively to the antigen. There was an excellent correlation (r2=0.90; P < 0.0001; n=26) between the response of mast cells to anti-IgE and the subsequent extent to which mast cells responded to NIP-HSA following passive sensitization. We considered whether the failure of non-releaser HLMC to respond to IgE-directed stimulation might relate to the expression of surface IgE. Mast cells of enhanced purity were assessed for the presence of surface IgE by flow cytometry using c-kit as a marker for mast cells. The data showed that non-releaser HLMC preparations do express IgE on their surface. There was a wide range in the levels of IgE on the surface of HLMC and we found no significant correlation (r2=0.04; P = 0.53; n=13) between IgE expression and releasability.

In summary, these results suggest that the releasability of mast cells is not governed by the expression of IgE on the cell surface. The variability in responsivesness to antigen may be due to differences within the signalling pathway downstream of the IgE receptor, FcεRI.