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Notes: A great variety of provocations of the airway mucosa produce extravasation of plasma from the abundant subepithelial microvessels. A plasma exudate has important actions through its volume, its specific and unspecific binding proteins, its enzyme systems, and its potent peptides (of kinin. complement, coagulation, fibrinolysis and other systems). If allowed to operate on the surface of an intact mucosa the plasma exudale would have important roles in normal airway defence. Recent observations in guinea-pig tracheo-bronchial airways and in human nasal airways suggest that the mucosal exudation of plasma into the airway lumen is a non-injurious fully reversible process. Threshold exudative responses thus resulted in the appearance of an ‘unfiltered’ plasma exudate not only in the lamina propria but also on the surface of an undisrupted mucosa. Even after extensive luminal entry of exudate the epithelial lining was intact, as judged by light, fluorescence and electron microscopy. Hence, the epithelial barrier was reversibly permeable when approached from beneath by the plasma exudate. This was a distinct increase in outward permeability, because even during the exudation of plasma the mucosa remained a barrier to luminal solutes. It is possible that the exudate itself, by a slight compressive action on the basolateral aspect of epithelial cells, creates intercellular pathways for its entry into the lumen. Contrary to current beliefs, we propose that plasma exudation should be considered a first line respiratory defence mechanism operating together with other systems of the mucosal surface.

Notes: A‘nasal pool’(NP) device, a compressible plastic container with an adapted nozzle, was used to perform a continuous 10-min nasal provocation and lavage. This novel technique brings known concentrations of agents into contact with a large and defined area of the nasal mucosal surface for extended periods of time. Simultaneously, the surface exudations/secretions of the same nasal mucosa are effectively sampled by the NP fluid. A concentration-response study of histamine (80, 400 and 2000 μg/ml) was performed in 12 normal subjects on three different occasions. Exudation of plasma albumin into the lavage fluid was measured to quantitate the histamine-induced airway inflammation. The effect of the dwell time on exudation was examined using histamine (400 μg/ml) instilled in the nasal cavity for time periods from 10 sec to 10 min. The time course of histamine-induced plasma exudation response was studied by exposing the mucosa to histamine (400 μg/ml) for 12 min, with the NP renewed every minute. Allergen-provocations were performed in subjects with hay fever and TAME-esterase activity in the returned lavage fluid was determined to indicate the degree of response. Histamine produced a concentration-dependent increase in albumin levels in the NP fluid; 123·3 ± 25·6, 213·8 ± 19·7 and 430·2 ± 32·0 μg/ml (mean ± s.e.m.), respectively. The time-course study demonstrated that plasma exudation into the lumen occurred promptly and that the exudation response reached a maximum after exposure to histamine for 6–10 min. The dwell-time experiments supported this finding. After 10 min the exudation appeared to decline despite the continued presence of histamine. Allergen provocation resulted in a concentration-dependent increase in TAME-esterase activity of the NP-fluids. It is concluded that the NP technique provides new possibilities in studies of pathophysiology and pharmacology of human airway mucosa. With the NP technique controlled concentrations of mediators, drugs, tracers, etc. can be applied for desirable lengths of time on a well-defined mucosal surface area. This particular area is gently and effectively lavaged during the presence of the NP fluid and when the fluid is recovered by decompressing the NP device.

Notes: Background Oral anti-histamine drugs are widely used in the treatment of seasonal allergic rhinitis. Recently, anti-histamines have become available also for topical treatmentObjective The present study, involving healthy subjects, examined the effect of topical azelastine on iuminal entry of α2-macroglobulin and symptoms evoked by repeat histamine challenges during 24 h. The effect was cotnpared to a clinical dose of the oral anti-histamine cetirizine and to placebo treatments.Methods Placebo and azelastine (0.254 mg per nasal cavity) were delivered as two consecutive actuations per nasal cavity using a nasal spray device. Oral placebo and cetirizine (10mg) were given as single doses in a placebo-controlled (double-dummy), double-blind, and cross-over design. Histamine-challenges were given 1 h before treatment, and 1, 6. 9, 12. and 24 h after each treatment. The nasal mucosal surface was lavaged after each challenge. The lavage-fiuid levels of α2-macroglobulin were determined to assess mucosal exudation of bulk plasma, and nasal symptoms were scored.Results Histamine (40–400 μg/mL) produced dose-dependent exudation and symptoms. Compared between each treatment and placebo, azelastine and cetirizine reduced the 40 and/or 400μg/mL histamine-induced mucosal exudation of plasma from 1–12 h after treatment. In addition, cetirizine reduced the 40μg/mL histamine-induced mucosal exudation of plasma 24 h after treatment. Differences between the two treatments were not evident regarding nasal symptoms.Conclusion Histamine challenge-induced mucosal exudation of plasma appears to be a useful method for studies of the duration of action of antihistamines. We conclude that topical azelastine is suited for b.i.d. therapy and that neither the exudative process nor watery secretion may impede the efficacy or the duration of action of tbis nasal drug.

Notes: Passive exposure to cigarette smoke has emerged as a significant risk factor in the development of asthma and allergic airways disease. The pathogenetic mechanisms are not known, but increased absorption across the airway epithelial lining has been suggested as one possible mechanism of this effect of cigarette smoke. This study examines the absorption-permeability of the nasal epithelial lining in cigarette smokers and non-smokers. For comparison, the effect of a detergent, dioctylsodium sulfosuccinate (DS), is also examined. A solution containing 51Cr-EDTA (51-chromium labeled ethylene diamine tetraacetic acid) (mol. wt. 372 Da) was instilled and maintained in the nasal cavity in six smokers and 12 non-smokers for 15min. Urine was collected for 24 h after the instillation. The accumulated amount of excreted 5lCr-EDTA was measured and expressed as millilitre nasal instillate. In six non-smokers the procedure was repeated when DS has been added to the instillate. The median recovered amount of 51Cr-EDTA in smokers 0.07 ml (range 0.04-0.32) was not significantly different from that in non-smokers 0.16 ml (0.01-1.22). The recovered amount of 51Cr-EDTA increased from a median of 0.18 ml (0.01-1.22) to 1.13 ml (0.53-1.80) after addition of the detergent (P= 0.028). We conclude that the nasal airway absorption-permeability is not increased in smokers. Hence, passive exposure to cigarette smoke may not produce an impairment of airway barrier functions.

Notes: Coronavirus-induced common cold and allergen-induced rhinitis are characterized by nasal mucosal exudation of bulk blood plasma. The mucosal exudation process involves ‘flooding’ of the lamina propria with plasma-derived binding proteins and it is possible that subepithelial inflammatory cytokines and mediators may be moved by the exudate to the mucosal surface. In this study, we have analysed cytokine levels in nasal lavage (NAL) fluids from non-allergic subjects inoculated with coronavirus (n= 20) and from subjects with allergic (birch pollen) rhinitis subjected to additional allergen challenge (samples were obtained 35min post challenge) in the laboratory (n= 10). Ten of the 20 inoculated subjects developed common cold and 10 remained healthy. Interferon-γ (IFN-γ), interleukin-1β (IL-1β), granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-4, and IL-6 were analysed in unprocessed NAL fluids using immunoassays. The subjects who developed common cold had increased NAL fluid levels of IFNγ (P 〈 0.05) that correlated well with the symptoms (P 〈 0.001). IFNγ did not increase in subjects with allergic rhinitis. IL-1β levels were similar in NAL fluids obtained from all inoculated subjects. In the subjects with allergic rhinitis NAL fluid levels of both IL-1β and GM-CSF were increased (P 〈 0.05). GM-CSF was not detected in common cold. IL-4 and IL-6 were not detectable in any of the NAL fluids. The present cytokines may not only emanate from superficial mucosal cells. By aiding plasma exudation subepithelial cytokines may potentially also be retrieved on the mucosal surface. Our study provides original in vivo data supporting the notion that a TH-1 profile of cytokines, notably IFNγ, is present in viral infection and further supporting the view that GM-CSF is an important cytokine in allergic airways disease.

Notes: 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.

Notes: Background Rofleponide palmitate is an esterified glucocorticosteroid pro-drug with a promising pre-clinical profile designed to deliver topical airway treatment for allergic rhinitis and asthma in a novel manner. Thus, the rofleponide palmitate pro-drug is designed to provide topical exposure of the mucosa to the inactive lipophilic drug, which would be locally metabolized to the more hydrophilic and readily cleared drug rofleponide.Objective To examine whether rofleponide palmitate affects nasal symptoms and peak inspiratory flow (PIF) in a pollen-season model of allergic rhinitis and to compare any such effects with those of another glucocorticosteroid (i.e., budesonide).Methods During the pollen-free season, 40 patients with strictly seasonal allergic rhinitis received topical nasal spray treatment with an aqueous solution of rofleponide palmitate 400 μg and an aqueous solution of budesonide 128 μg once daily for 10 days in a double-blind, placebo-controlled, and crossover study. After 3 days of drug treatment, individualized allergen challenges were given once daily for 7 days while the treatment continued. The washout periods between each of the challenge series were 2 weeks. Nasal symptoms and PIF were recorded in the morning and evening, as well as 10 and 20 min after each allergen challenge. The mean recordings obtained during the last 3 days of the allergen-challenge series, when symptoms were established and when the treatment had lasted for 8–10 days, were used in the analysis.Results Both active treatments reduced nasal symptoms and improved nasal PIF compared with placebo (P〈0.01–0.001). There was no overall difference in efficacy between rofleponide palmitate 400 μg and budesonide 128 μg.Conclusions Topical treatment with aqueous solutions of rofleponide palmitate attenuates nasal symptoms and improves nasal PIF in allergic rhinitis. The overall efficacy of 400 μg of rofleponide palmitate is similar to that of 128 μg of budesonide in the pollen-season model used in this study.

Notes: Background Genetic engineering of the major birch pollen allergen (Bet v 1) has led to the generation of recombinant Bet v 1 derivatives with markedly reduced IgE-binding capacity, but with retained T cell activating ability.Objective To compare the mucosal reactivity to rBet v 1 derivatives with rBet v 1 wild-type as basis for new therapeutic strategies for birch pollen allergy based on mucosal tolerance induction.Methods Outside the pollen season, 10 patients with birch pollen allergic rhinitis and mild asthma underwent four nasal challenge-sessions in a randomized, double-blind, and cross-over design, employing increasing doses of rBet v 1 fragment mix, rBet v 1 trimer, rBet v 1 wild-type and diluent (albumin). Nasal lavage fluids (NAL) were collected before the challenge-series as well as 10 min, 4 and 24 h thereafter. Nasal lavage fluid levels of tryptase as well as EPO and ECP were measured as indices of mast cell and eosinophil activity, respectively.Results All 10 patients tolerated the highest accumulated dose, 8.124 µg, when challenged with rBet v 1 trimer, eight with rBet v 1 fragments compared to one when challenged with rBet v 1 wild-type. No late phase reactions were observed. The change in tryptase levels (pre-challenge vs. 10 min) was significantly lower after challenges with rBet v 1 trimer and rBet v 1 fragments than with rBet v 1 wild-type. The change in EPO/ECP concentration pre-challenge versus 4 h post-challenge was lower for rBet v 1 trimer and the change was significantly lower when pre-challenge versus 24 h post-challenge to rBet v 1 fragments and rBet v 1 wild-type was examined.Conclusion The derivatives induced significantly fewer symptoms and lower mast cell and eosinophil activation than rBet v 1 wild-type upon application to the nasal mucosa. They could in the future be candidates for immunotherapy based on mucosal tolerance induction.

Notes: 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 (α2-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 α2-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.