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FcepsilonRI receptors play an important role in allergen-induced mediator release and antigen presentation by mast cells, basophils, and monocyte/macrophages in atopic disorders. The expression of FcepsilonRI by tissue eosinophils in atopic asthma after allergen challenge has not been established. For this reason we attempted to identify mRNA and protein product + FcepsilonRIalpha eosinophils in cytospins made from bronchoalveolar lavage (BAL) from atopic asthmatics (n = 9) and nonatopic normal subjects (n = 4) 24 h after segmental challenge with allergen or diluent. Messenger RNA for FcepsilonRIalpha was determined using in situ hybridization and FcepsilonRIalpha protein expression by immunocytochemistry using a mouse monoclonal antibody 22E7. Colocalization of FcepsilonRIalpha receptors to eosinophils was performed using chromotrope 2R. When compared with a control challenge, segmental challenge with Dermatophagoides pteronyssinus induced significant BAL eosinophilia (p = 0.007). The total number of BAL FcepsilonRIalpha mRNA and protein-positive cells also increased in asthmatics, median values 2 (0.7-7.2) and 11.5 (0.6-65.0) x 10(6) cells (p = 0.02) and 0 (0-0.3 x 10(6)) and 3.1 x 10(6) (0.45 - 162.5 x 10(6)) cells (p = 0.007), respectively, for mRNA and protein. Net increases in FcepsilonRIalpha+ cells correlated with the net increases in BAL eosinophils (r = 0.98, p = 0.0001 for mRNA and r = 0.72, p = 0.02 for protein). Colocalization studies with chromotrope 2R revealed that only 4% of FcepsilonRIalpha+ cells were eosinophils after control challenge and, in contrast, 85 to 95% of FcepsilonRIalpha+ cells were eosinophils after allergen. There were no differences in the numbers of FcepsilonRIalpha+ cells or eosinophils in normal control subjects. Our results demonstrated that local endobronchial allergen provocation in atopic asthmatics results in increased synthesis and expression of FcepsilonRIalpha predominantly on BAL eosinophils.

We have investigated the phenotype of interleukin-5 (IL-5) mRNA+ cells in the nasal mucosa of subjects with allergic rhinitis. Serial cryostat sections were cut from paraformaldehyde-fixed snap-frozen nasal biopsies from six patients, before and 24 h after local allergen provocation with grass pollen. Immunocytochemistry (APAAP) was followed by in situ hybridization on the same sections. For immunocytochemistry, antibodies against CD3, tryptase, and major basic protein (MBP) were used to identify T cells, mast cells, and eosinophils, respectively. Hybridization studies were performed using a Digoxigenin-labeled IL-5 riboprobe. Nitroblue tetrazolium (NBT) and X-phosphate-5-bromo-4-chloro-3- indoly phosphate (BCIP) served as chromogens to detect hybridized IL-5 mRNA signals. The majority of IL-5 mRNA+ cells were CD3+ (83.2%), whereas the remainder were either tryptase+ (11.3%) or MBP+ (5.4%). In contrast, only a few IL-5 mRNA+ cells were observed in nasal biopsies before challenge, all of which were co-localized to CD3+ cells. These results indicate that CD3+ cells are the principal cellular source of IL-5 transcripts in the nasal mucosa 24 h after allergen-induced late-phase nasal responses.