Methods: In a diluent-controlled allergen (Ag)-challenge cross-over study, ILC2s (lin-FcεRI-CD45+CD127+ST2+CRTH2+), CD4+ T lymphocytes, and levels of intracellular IL-5 and IL-13 expression were enumerated by flow cytometry in steroid-naive mild atopic asthmatics. All subjects (n=7) developed Ag-induced dual bronchoconstriction, airway eosinophilia and increased methacholine airway responsiveness. Bone marrow, blood and sputum samples were collected pre-, 24 and 48h post-challenge and immediately fixed in 1% paraformaldehyde prior to immunofluorescence staining.
Results: Compared to pre-Ag levels, there was a significant increase in sputum ILC2 at 24h (P<0.01) which decreased to baseline levels 48h post-Ag. This was co-incident with a significant decrease in blood and bone marrow ILC2 at 24h post-Ag compared to pre-Ag levels. Activated ILC2 (IL5+ or IL13+) in sputum increased significantly at 24 h post-Ag only. In contrast, although total CD4+ T lymphocytes and IL-13+CD4+ T cells increased at 24h post-Ag (P<0.01), IL-5+CD4+ T cells only showed a trend for an increase at 48h post-Ag. No effect of diluent was observed.
Conclusions: Our findings suggest that post-allergen exposure in mild atopic asthmatics, ILC2 may initiate the development of eosinophilic inflammation while CD4+ T lymphocytes may be involved in the persistence of the asthmatic response.