MAB1222A Sigma-AldrichAnti-Tryptase Antibody, Mast Cell, clone G3, Alkaline Phosphatase

BACKGROUND: Lung mast cells are stereotypically divided into connective tissue (MC(TC)) and mucosal (MC(T)) mast cells. This study tests the hypothesis that each of these subtypes can be divided further into site-specific populations created by the microenvironment within each anatomical lung compartment. METHODS: Surgical resections and bronchial and transbronchial biopsies from non-smoking individuals were obtained to study mast cells under non-inflamed conditions. Morphometric and molecular characteristics of mast cell populations were investigated in multiple lung structures by immunohistochemistry and electron microscopy. RESULTS: MC(T) and MC(TC) coexisted in all compartments, with MC(T) being the prevailing type in bronchi, bronchioles and the alveolar parenchyma and MC(TC) being more abundant in pulmonary vessels and the pleura. Each of the MC(TC) and MC(T) phenotypes could be further differentiated into site-specific populations. MC(TC) were significantly larger in pulmonary vessels than in small airway walls, while the reverse was observed for MC(T). Within each MC(TC) and MC(T) population there were also distinct site-specific expression patterns of the IgE receptor, interleukin-9 receptor, renin, histidine decarboxylase, vascular endothelial growth factor, fibroblast growth factor, 5-lipoxygenase and leukotriene C4 synthase (eg, bronchial MC(T) consistently expressed more histidine decarboxylase than alveolar MC(T)). Renin content was high in vascular MC(TC) but markedly lower in MC(TC) in other compartments. For both MC(TC) and MC(T), the IgE receptor was highly expressed in conducting airways but virtually absent in alveolar parenchyma. CONCLUSIONS: These findings demonstrate novel site-specific subpopulations of lung MC(TC) and MC(T) at baseline conditions. This observation may have important implications in the future exploration of mast cells in a number of pulmonary diseases.

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.

Lesional (n = 15) and non-lesional (n = 10) skin of subjects with mastocytosis was analysed for the distribution and concentration of trypase positive, chymase negative mast cells (MCT) and tryptase positive, chymase positive mast cells (MCTC) cells and compared to normal skin (n = 23) and non-lesional skin of subjects with unexplained anaphylaxis or flushing episodes (n = 6). Skin biopsies were fixed in Carnoy's fluid and subjected to double immunohistochemical staining with biotinylated mouse monoclonal anti-chymase antibody followed by alkaline phosphatase-conjugated mouse monoclonal anti-tryptase antibody. MCTC cells were the only type of mast cells seen in all specimens analysed and in each case were more numerous in superficial compared to deep regions of dermis. The concentration (mean +/- s.d.) of mast cells in the superficial dermis of mastocytosis lesions (40 985 +/- 21 772 mast cells/mm3) was significantly increased over that in corresponding areas of non-lesional skin from subjects with mastocytosis (7178 +/- 3607 mast cells/mm3), skin from subjects with idiopathic anaphylaxis or flushing episodes (6974 +/- 3873 mast cells/mm3) and normal skin (7347 +/- 2973 mast cells/mm3). The exclusive presence of MCTC cells in skin lesions of mastocytosis which are characterized by non-malignant hyperplasia of mast cells suggests involvement of local tissue factors in mast cell recruitment and differentiation.