Circulating eosinophils in asthma, allergic rhinitis, and atopic dermatitis lack morphological signs of degranulation.

BACKGROUND: In allergic diseases, eosinophils in affected tissues release granule proteins with cytotoxic, immunoregulatory, and remodelling-promoting properties. From recent observations, it may be assumed that eosinophils degranulate already in circulating blood. If degranulation occurs in the circulation, this could contribute to widespread systemic effects and provide an important marker of disease. OBJECTIVE: To determine the degranulation status of circulating eosinophils in common allergic diseases. METHODS: Using a novel approach of whole blood fixation and leucocyte preparation, the granule morphology of blood eosinophils from healthy subjects, non-symptomatic patients, symptomatic patients with asthma, asthma and Churg-Strauss syndrome, allergic rhinitis, and atopic dermatitis was evaluated by transmission electron microscopy (TEM) and eosinophil peroxidase (TEM) histochemistry. Plasma and serum levels of eosinophil cationic protein were measured by fluoroenzymeimmunoassay. Selected tissue biopsies were examined by TEM. RESULTS: Regardless of symptoms, circulating eosinophils from allergic patients showed the same granule morphology as cells from healthy subjects. The majority of eosinophil-specific granules had preserved intact electron-density (96%; range: 89-98%), while the remaining granules typically exhibited marginal coarsening or mild lucency of the matrix structure. Abnormalities of the crystalline granule core were rarely detected. Furthermore, granule matrix alterations were not associated with any re-localization of intracellular EPO or increase in plasma eosinophil cationic protein. By contrast, eosinophils in diseased tissues exhibited cytolysis (granule release through membrane rupture) and piecemeal degranulation (loss of granule matrix and core structures). CONCLUSION: In symptomatic eosinophilic diseases, circulating blood eosinophils retain their granule contents until they have reached their target organ.

Lung tissue eosinophils may be cleared through luminal entry rather than apoptosis: effects of steroid treatment.

Spontaneous or steroid-induced eosinophil apoptosis occurring in vitro has not been demonstrated in lung tissues in vivo. This study examines cell apoptosis in rat lungs using the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate (dUTP) nick end labeling (TUNEL) technique and transmission electron microscopy (TEM). After establishing sustained lung edema and eosinophilia by challenge with Sephadex beads intratracheally, budesonide treatment was started intratracheally. Sephadex alone increased the total number of apoptotic cells, which were not efficiently engulfed by macrophages or other cells, in vivo. Yet apoptotic tissue eosinophils were exceedingly rare (1 of 360 TEM-analyzed eosinophils). By contrast, approximately 20% of eosinophils in the airway lumen were apoptotic, and unengulfed. Budesonide promptly inhibited edema but 3 d of steroid treatment were required to reduce the established tissue eosinophilia. Not at any time point did budesonide induce eosinophil apoptosis (0 of 318 TEM-analyzed tissue eosinophils). We conclude that (1) eosinophil apoptosis can occur but is a rare event in vivo in respiratory tract tissues; (2) airway tissue eosinophils, rather than undergoing apoptosis, are eliminated by migration into airway lumen followed by apoptosis and mucociliary clearance; (3) anti-inflammatory steroid treatment may not increase eosinophil apoptosis in vivo nor may it affect the luminal entry of eosinophils; (4) steroids permit elimination of eosinophils into airway lumen and slowly resolve established lung eosinophilia.

Mucosal output of eotaxin in allergic rhinitis and its attenuation by topical glucocorticosteroid treatment.

BACKGROUND: Eotaxin is a chemokine that attracts and activates eosinophils. The present study examines the occurrence of eotaxin in nasal mucosal surface liquids in patients with seasonal allergic rhinitis without allergen exposure and during repeat allergen challenge with and without topical glucocorticosteroid treatment. The number of subepithelial eosinophils and mucosal outputs of bulk plasma (alpha2-macroglobulin) and eosinophil cationic protein (ECP) are also examined. METHODS: Twelve patients underwent daily allergen challenges for 6 days. Separately, 14 patients, who were receiving budesonide and placebo in a parallel group design, also underwent allergen challenge for 6 days. Nasal biopsies were obtained before and 24 h after the allergen challenge series, and lavages were carried out before and 15 min after selected allergen challenges. RESULTS: At baseline nasal lavage fluid levels of eotaxin correlated to levels of alpha2-macroglobulin and ECP. After the first allergen challenge there was a correlation between nasal lavage fluid levels of eotaxin and ECP. Repeat allergen exposure increased the mucosal output of eotaxin (P <0.05) and ECP (P <0.01) as well as eosinophil numbers (P <0.01), but no correlation was found between increased eosinophil numbers and eotaxin. Budesonide reduced eotaxin levels during repeat allergen challenge (P <0.05). CONCLUSIONS: Repeat allergen exposure in allergic rhinitis is associated with increased mucosal output of eotaxin. Topical budesonide attenuates this effect, suggesting the possibility that inhibitory effects on mucosal eotaxin may contribute to anti-eosinophilic actions of topical glucocorticosteroids.

Effects of hydrogen peroxide on the guinea-pig tracheobronchial mucosa in vivo.

Lumenal entry of plasma (mucosal exudation) is a key feature of airway inflammation. In airways challenged with histamine-type mediators and allergen the mucosal exudation response occurs without causing epithelial derangement and without increased airway absorption. In contrast, reactive oxygen metabolites may cause mucosal damage. In this study, involving guinea-pig airways, we have examined effects of H2O2 on airway exudation and absorption in vivo. Vehicle or H2O2 (0.1 and 0.5 M) was superfused onto the tracheobronchial mucosal surface through an oro-tracheal catheter. 125I-albumin, given intravenously, was determined in tracheobronchial tissue and in lavage fluids 10 min after challenge as an index of mucosal exudation of plasma. The tracheobronchial mucosa was also examined by scanning electron microscopy. In separate animals, 99mTc-DTPA was superfused 20 min after vehicle or H2O2 (0.1 and 0.5 M) had been given. A gamma camera determined the disappearance rate of 99mTc-DTPA from the airways as an index of airway absorption. The high dose of H2O2 (0.5 M) produced epithelial damage, increased the absorption of 99mTc-DTPA (P < 0.001), and increased the exudation of plasma (P < 0.001). Notably, it appeared that all extravasated plasma had entered the airway lumen within 10 min. These data demonstrate that H2O2 differs from exudative autacoids such as histamine by causing both epithelial damage and plasma exudation responses. These data also agree with the view that the epithelial lining determines the rate of absorption and is responsible for the valve-like function that allows lumenal entry of extravasated bulk plasma without any increased inward perviousness.

Generation of clusters of free eosinophil granules (Cfegs) in seasonal allergic rhinitis.

Generation of clusters of free eosinophil granules (Cfegs), through lysis of eosinophils, has recently been proposed as a major paradigm for ultimate activation of airway mucosal eosinophils. In the present study involving patients with seasonal allergic rhinitis, we have investigated whether generation of Cfegs in the nasal mucosa is a feature of allergic rhinitis. Nasal mucosal biopsies were obtained before and late in a birch-pollen season, and were subjected to histochemical staining of eosinophil peroxidase. In biopsies obtained before the pollen season, a few, intact eosinophils were observed, and Cfegs were scarce. In biopsies taken during the pollen season, the numbers of eosinophils were increased about 10-fold (P < 0.05) and the Cfegs about 25-fold (P < 0.05). We conclude that generation of Cfegs is a significant feature of seasonal allergic rhinitis and that this process represents the ultimate activation of mucosal eosinophils in this disease.