Asthma is characterized by an increase in airway smooth muscle mass and a decreased distance between the smooth muscle layer and the epithelium. Furthermore, there is evidence to indicate that airway smooth muscle cells (ASMC) express a wide variety of receptors involved in the immune response. The aims of this study were to examine the expression of CCR3 on ASMC, to compare this expression between asthmatic and nonasthmatic subjects, and to determine the implications of CCR3 expression in the migration of ASMC. We first demonstrated that ASMC constitutively express CCR3 at both mRNA and protein levels. Interestingly, TNF-α increases ASMC surface expression of CCR3 from 33 to 74%. Furthermore, using FACS analysis, we found that ASMC CCR3 is expressed to a greater degree in asthmatic vs control subjects (95 vs 75%). Functionality of the receptor was demonstrated by calcium assay; the addition of CCR3 ligand eotaxin to ASMC resulted in an increase in intracellular calcium production. Interestingly, ASMC was seen to demonstrate a positive chemotactic response to eotaxin. Indeed, ASMC significantly migrated toward 100 ng/ml eotaxin (2.2-fold increase, compared with control). In conclusion, the expression of CCR3 by ASMC is increased in asthmatics, and our data show that a CCR3 ligand such as eotaxin induces migration of ASMC in vitro. These results may suggest that eotaxin could be involved in the increased smooth muscle mass observed in asthmatics through the activation of CCR3.

The effects of remodeling of airway smooth muscle (SM) by hyperplasia on airway SM contractility in vivo are poorly explored. The aim of this study was to investigate the relationship between allergen-induced airway SM hyperplasia and its contractile phenotype. Brown Norway rats were sensitized with ovalbumin (OVA) or saline on day 0 and then either OVA-challenged once on day 14 and killed 24 h later or OVA-challenged 3 times (on days 14, 19, and 24) and killed 2 or 7 days later. Changes in SM mass, expression of total myosin, SM myosin heavy chain fast isoform (SM-B) and myosin light chain kinase (MLCK), tracheal contractions ex vivo, and airway responsiveness to methacholine (MCh) in vivo were assessed. One day after a single OVA challenge, the number of SM cells positive for PCNA was greater than for control animals, whereas the SM mass, contractile phenotype, and tracheal contractility were unchanged. Two days after three challenges, SM mass and PCNA immunoreactive cells were increased (3-and 10-fold, respectively; P Ͻ 0.05), but airway responsiveness to MCh was unaffected. Lower expression in total myosin, SM-B, and MLCK was observed at the mRNA level (P Ͻ 0.05), and total myosin and MLCK expression were lower at the protein level (P Ͻ 0.05) after normalization for SM mass. Normalized tracheal SM force generation was also significantly lower 2 days after repeated challenges (P Ͻ 0.05). Seven days after repeated challenges, features of remodeling were restored toward control levels. Allergen-induced hyperplasia of SM cells was associated with a loss of contractile phenotype, which was offset by the increase in mass. phenotype; animal model; myosin heavy chain isoform STRUCTURAL AIRWAY REMODELING has been widely reported in both human asthma (4, 13) and animal models of experimental asthma (11,23,37). Most tissues of the airway wall are affected by remodeling, and the observed changes include shedding of the epithelium, thickening of the reticular basement membrane, increased mucus gland size and number, goblet cell differentiation, angiogenesis, and augmentation of the airway smooth muscle (SM) mass (4). The changes in airway wall tissues are likely to be of clinical importance since they may control airway caliber, airway elasticity, and responsiveness and may cause symptoms related to mucus overproduction (4). Airway SM remodeling is a reported feature of several animal models of allergic asthma (11,12,22), including the Brown Norway (BN) rat (19,26,31). The increase in SM mass, observed in asthma of varying severity (5, 29, 36), has been proposed to be sufficient to account for altered airway responsiveness (16). The augmentation of airway SM mass has been attributed to hyperplasia through evidence of cell proliferation quantified by bromodeoxyuridine incorporation or PCNA expression in vivo (11,19,28,37). Susceptibility to SM remodeling may be a risk factor for asthma; cultured airway SM cells from asthmatic subjects were shown to have a higher proliferation rate than cells from normal subjects (14), sim...

A pool of six bronchial biopsy specimens can provide tissue of excellent quality in both asthmatic and healthy subjects and, consequently, a valuable sample for morphological analysis of mucosal structures.

Background: Blood eosinophils express CD16 on their surface when stimulated in vitro with platelet-activating factor or IFNγ. Transient expression of CD16 is also observed in vivo following aeroallergen challenge of asthmatic subjects. The present work is aimed at evaluating the possible mechanisms modulating eosinophil expression of CD16 and the biological functions of this receptor. Methods: First, purified blood eosinophils were incubated with IL-1β, IL-2, IL-4, IL-5, IL-9 or IL-16, GM-CSF, IFNγ, eotaxin or 5-oxo-ETE and CD16 expression was measured. Second, the capacity of CD16 to mediate degranulation induced by IgG immune complexes (IC) was evaluated in eosinophils with low and high CD16 expression. Finally, serum allergen-specific IgE and IgG, and total IgE levels were measured at baseline in allergic asthmatics and correlated with changes observed in blood eosinophil CD16 expression (ΔCD16) following allergen challenge. Results: Only IFNγ and IL-2 significantly increased the number of CD16+ eosinophils, respectively, 37 ± 10% (p = 0.0038) and 38 ± 8% (p = 0.0006), compared to control, 7 ± 2%. IgG IC induced degranulation in eosinophils with low and high CD16 expression and monoclonal anti-CD16 and anti-CD32 antibodies inhibited this. IgG IC increased eosinophil CD16 expression (14 ± 6%, p = 0.0008) and this effect was blocked by pretreatment with anti-CD32 antibodies. ΔCD16 following allergen challenge correlated with the specific IgG/total IgE ratio (r² = 0.41, p = 0.036). Conclusion: These data suggest that formation of IgG IC is associated with surface eosinophil CD16 expression in asthma and that CD16 in cooperation with CD32 mediates IC-induced degranulation.

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