Airway inflammation in asthma and perennial allergic rhinitis. Relationship with nonspecific bronchial responsiveness and maximal airway narrowing

BACKGROUND: Eosinophilic airway inflammation is the hallmark of asthma, but it has also been reported in other conditions such as allergic rhinitis. We have tested whether the analysis of cells and chemicals in sputum can distinguish between patients with mild allergic asthma, those with allergic rhinitis, and healthy controls. The relationship between inflammation markers in sputum and nonspecific bronchial hyperresponsiveness to methacholine (BHR) (PD20 and maximal response plateau [MRP] values) was also evaluated. METHODS: We selected 31 mild asthmatics and 15 rhinitis patients sensitized to house-dust mite. As a control group, we studied 10 healthy subjects. Every subject underwent the methacholine bronchial provocation test (M-BPT) and sputum induction. Blood eosinophils and serum ECP levels were measured. Sputum cell differentials were assessed, and eosinophil cationic protein (ECP), tryptase, albumin, and interleukin (IL)-5 levels were measured in the entire sputum supernatant. RESULTS: Blood eosinophils and serum ECP levels were higher in asthma patients and rhinitis than in healthy controls, but no difference between asthma patients and rhinitis patients was found. Asthmatics had higher eosinophil counts and higher ECP and tryptase levels in sputum than rhinitis patients or control subjects. Sputum albumin levels were higher in asthmatics than in controls. Rhinitis patients exhibited higher sputum eosinophils than healthy controls. An association between sputum eosinophil numbers and MPR values (r= -0.57) was detected, and a trend toward correlation between sputum ECP levels and PD20 values (r= -0.47) was found in the rhinitis group, but not in asthmatics. No correlation between blood eosinophilic inflammation and lung functional indices was found. CONCLUSIONS: Induced sputum is an accurate method to study bronchial inflammation, allowing one to distinguish between rhinitis patients and mildly asthmatic patients. The fact that no relationship was detected between sputum inflammation and BHR suggests that other factors, such as airway remodeling, may be at least partly responsible for BHR in asthma.     

Effects of prednisolone on eosinophils, IL-5, eosinophil cationic protein, EG2+ eosinophils, and nitric oxide metabolites in the sputum of patients with exacerbated asthma

Corticosteroids are considered to be one of the most effective medicine for asthma by suppressing airway inflammation. This study was carried out to investigate the effects of prednisolone in the sputum of exacerbated asthmatics. Clinical severity, cell differentials, levels of interleukin (IL)-5, eosinophil cationic protein (ECP), EG2+ eosinophils, and nitric oxide (NO) metabolites were measured. Sputum was examined 2 weeks apart in 13 exacerbated asthmatics before and after prednisolone treatment, and once in 12 stable asthmatics. We used a sandwich ELISA for IL-5, fluoroimmunoassay for ECP, immunohistochemical staining for EG2+ eosinophils, a NO metabolites assay using modified Griess reaction. Exacerbated asthmatics, in comparison with stable asthmatics, had significantly higher proportion of eosinophils, higher level of ECP, higher percentage of EG2+ eosinophils, and NO metabolites. Exacerbated asthmatics after treatment with prednisolone had reduced the proportions of eosinophils, reduced level of IL-5, ECP and percentage of EG2+ eosinophils. FEV1 was correlated with the proportion of eosinophils, ECP, and IL-5 respectively. These findings suggest that prednisolone is considered to be effective medicine for asthma by suppressing eosinophil activation through IL-5.     

Bronchial hyperresponsiveness and airway inflammation in adolescents with asymptomatic childhood asthma

BACKGROUND: About 70% of childhood asthmatics become free of asthma-related symptoms during adolescence. Little is known about bronchial hyperresponsiveness (BHR) and airway inflammation in young adults with "outgrown" childhood asthma. METHODS: We studied 61 nonsmoking medical students (18 intermittent mild asthmatics, 23 students with outgrown childhood asthma but free of asthma-related symptoms for 10 years (asymptomatic asthmatics) and 20 healthy students). BHR and lung function were measured, and induced sputum samples analyzed for eosinophil count, eosinophilic cationic protein (ECP), granulocyte-macrophage colony stimulating factor (GM-CSF), and tumor necrosis factor-alpha (TNF-alpha). RESULTS: BHR was still present in most asymptomatic asthmatics, but it was milder compared with healthy students. Only three subjects with previous asthma had no BHR and no signs of airway inflammation. Percentages of eosinophil, and ECP, TNF-alpha and GM-CSF concentrations in induced sputum of mild asthmatics and asymptomatic asthma groups were higher than in the healthy group. In asymptomatic asthmatics group, the duration of asthma, sputum eosinophil percentage, and the level of TNF-alpha in sputum correlated significantly with BHR. CONCLUSIONS: Only a few subjects with longstanding asymptomatic asthma could be considered as cured; most asymptomatic asthmatics continued to exhibit BHR and signs of airway inflammation. The outcome of childhood asthma and BHR was associated with the degree of airway inflammation and the duration of childhood asthma.     

Airway neutrophil inflammation in nonasthmatic patients with food allergy

BACKGROUND: Patients with food allergy (FA) have been recently shown to develop bronchial hyperresponsiveness (BHR), despite the absence of any concomitant asthmatic manifestation. In order to explain this observation, we sought to examine the presence of a bronchial inflammation in induced sputum of nonasthmatic patients with FA. METHODS: Twelve nonasthmatic patients with FA (urticaria, digestive symptoms, anaphylaxis) were included in the study. Results were compared to these obtained from eight asthmatic patients without food allergy and eight healthy controls. Diagnosis of FA was based on double-blind placebo-controlled challenge. Sputum cells and fluid-phase eosinophil cationic protein (ECP), myeloperoxidase (MPO) and interleukin-8 (IL-8) were measured in induced sputum. BHR was evaluated using methacholine inhalation. RESULTS: Sputum from asthmatics, in comparison with the sputum of healthy subjects and patients with FA contained a higher proportion of eosinophils and higher levels of ECP (< 0.001). In marked contrast, patients with FA exhibited an increased proportion of neutrophils and IL-8 in comparison with asthmatics and controls (P < 0.05 for neutrophils and P < 0.001 for IL-8). There was a significant correlation between sputum neutrophils and IL-8 (r = 0.68, P < 0.001). MPO levels were not different between the groups. There was a trend toward higher levels of IL-8 and ECP in food allergic patients with BHR in comparison with patients with FA without BHR. CONCLUSION: Our results demonstrate that a subclinical neutrophil airway inflammation is present in patients with food allergy free of clinical respiratory symptoms and that IL-8 may be an important mediator of this neutrophilia.     

Involvement of IL-9 in the bronchial phenotype of patients with nasal polyposis

BACKGROUND: Nasal polyposis (NP) is frequently associated with asthma. In this disease, asymptomatic bronchial hyperresponsiveness (BHR) is thought to precede the development of asthma. IL-9 and its receptor have been reported as candidate genes for asthma and to be associated with BHR. OBJECTIVE: The objective of this study was to assess the contribution of 11-9 to the pathogenesis of BHR in NP by comparing the expression of IL-9 and its receptor in bronchial biopsy specimens from three groups of patients with NP: NP without BHR, NP with asymptomatic BHR, and NP with BHR and asthma. METHODS: Bronchial biopsy specimens were examined in terms of cellular infiltration and in terms of expression of IL-9 protein and mRNA as well as of its receptor by using immunohistochemistry and in situ hybridization. RESULTS: Patients with NP with asthma as compared with the two other groups exhibited an increased bronchial infiltration of basophils, eosinophils, and T cells that correlated with the asthma score. The two groups of patients with NP with BHR showed an increased expression in IL-9 protein and mRNA as well as an increase in the expression of IL-9R mRNA at the epithelial level. These modifications were inversely correlated with the airway responsiveness to methacholine, producing a 20% fall in FEV1. There was a close association between IL-9+ cells, IL-5 mRNA expression, and eosinophil infiltration that correlated with each other. CONCLUSIONS: These results suggest an important role for IL-9 in the pathogenesis of BHR and a causal relation between IL-9 and the development of bronchial eosinophilia in asthma.     

Repeated inhalation of low doses of cat allergen that do not induce clinical symptoms increases bronchial hyperresponsiveness and eosinophil cationic protein levels.

The aim of this study was to investigate whether repeated exposure to subclinical doses of cat allergens, not inducing asthma symptoms, could affect eosinophil cationic protein (ECP) levels in bronchoalveolar lavage (BAL) or in peripheral blood, without the appearance of clinical symptoms. Twelve patients with mild asthma, all sensitized to cats and not exposed to cat allergen at home, underwent a series of inhalations of cat allergen or placebo for 8 days over 2 weeks. A methacholine challenge was performed before and after the allergen and saline exposures, and BAL and blood were sampled for ECP measurements and eosinophil counts. No patients experienced asthma symptoms. However, PD20 methacholine (geometric mean) decreased significantly from 263 microg before to 126 microg after inhalation of allergen. Inhalation of saline did not induce any significant change in PD20. The change in log PD20 before and after cat allergen exposure was statistically different from the change in log PD20 before and after saline. Median ECP levels in BAL and serum increased significantly after allergen exposure, from 0.8 to 3.1 microg/l (p<0.02) and from 15.9 to 31.4 microg/l (p<0.05), respectively. No change was observed after saline inhalations. The change in BAL and serum ECP levels was statistically significant compared to that in the control group. The number of eosinophils did not change, however, nor did IL-5 and RANTES levels in BAL and serum. In conclusion, our results show that (1) exposure of asthma patients to repeated low doses of allergen, which did not provoke any clinical symptoms, is capable of inducing a local eosinophil activation associated with an increase in nonspecific bronchial hyperresponsiveness and (2) the increase in serum ECP levels due to eosinophil activation precedes the occurrence of asthma symptoms and may thus be a marker of allergen exposure in allergic asthma.     

Effect of inhaled interleukin-5 on number and activity of eosinophils in circulation from asthmatics

The aim of the present study was to examine the effects of interleukin-5 (IL-5) inhalation on changes in the activity and number of circulating eosinophils, as well as concentrations of serum total IgE, in allergic asthmatics. A randomized double-blind, placebo-controlled study design was employed in which each subject acted as his or her own control. Eight nonsmoking patients with allergic asthma were administered recombinant human IL-5 by nebulization. Total white blood cell counts and differentials, as well as concentrations of ECP and total IgE in serum, were determined before and at 2, 24, and 48 h after inhalation. Our results demonstrated that eosinophil numbers increased from baseline (3.6 +/- 1.1 x 10(5)/ml) to 6.3 +/- 1.2 x 10(5)/ml (P < 0.01) at 24 h and to 5.7 +/- 0.9 x 10(5)/ml (P < 0.01) at 48 h after IL-5 inhalation in asthmatics. Accompanying this significantly increased blood eosinophilia were significantly elevated serum ECP levels. Compared with baseline (6.3 +/- 1.1 ng/ml), ECP levels increased with time following IL-5 inhalation, reaching 17.6 +/- 2.8 ng/ml (P < 0.01) at 24 h and remaining elevated at 48 h (18.1 +/- 2.9 ng/ml, P < 0.01). IL-5 inhalation had no significant effect on levels of serum total IgE, however. These findings provide direct evidence that nebulized IL-5 not only induces a significant blood eosinophilia but also results in the activation of circulating eosinophils. Our data further support the importance of IL-5 in the pathogenesis of bronchial asthma in humans.     

Changes in serum eotaxin and eosinophil cationic protein levels, and eosinophil count during treatment of childhood asthma.

BACKGROUND AND PURPOSE: Increased serum levels of eotaxin are related to the severity of asthma in adults. There are limited data on the effects of oral corticosteroids and inhaled corticosteroid therapy on serum levels of eotaxin and eosinophil cationic protein (ECP) and peripheral blood eosinophil counts (ECs) in pediatric asthma patients. We investigated prospectively the changes in eotaxin and ECP serum levels and peripheral blood ECs after administering oral corticosteroids and then inhaled corticosteroids plus long-acting beta2 agonist treatment in pediatric patients. METHODS: Serum samples of 20 pediatric patients with mild-to-moderate asthma were collected before treatment, after 5-7 days of oral prednisolone treatment, and after 1-2 months of inhaled fluticasone plus salmeterol treatment. Peak expiratory flow was used as the outcome index. RESULTS: Serum eotaxin levels remained the same after oral prednisolone treatment, but decreased after subsequent inhalation treatment compared with the end of oral steroid treatment (64.7 +/- 22.6 vs 85.7 +/- 36.8 pg/mL, p<0.001). The EC and serum ECP levels declined soon after oral steroid treatment, rebounding to initial levels during inhalation treatment. The decrease in ECP level was positively correlated with the decrease in ECs with oral steroid treatment (r(2) = 0.28, p=0.016). There was no correlation between changes in eotaxin levels and peak expiratory flow. CONCLUSIONS: Our data suggest that the serum eotaxin level, not peripheral blood EC or serum ECP level, declines during inhaled fluticasone plus salmeterol treatment and might serve as a surrogate marker of T helper 2 residual activity in pediatric asthma.     

Analysis of induced sputum to examine the effects of inhaled corticosteroid on airway inflammation in children with asthma.

BACKGROUND: Analysis of induced sputum can be performed safely in children with asthma and is useful for both cellular and biochemical markers of inflammation. Glucocorticosteroid inhalation has become the first line therapy for chronic asthma by suppressing airway inflammation, which produces the decrease of bronchial hyperreactivity and reduces the number of eosinophil in bronchial submucosa. OBJECTIVE: To determine the characteristics of the inflammatory cells and their markers in sputum and to examine the pharmacokinetic effects of glucocorticoid within 3 hours after inhalation therapy on FEV1 and sputum inflammatory indices in children with clinically defined chronic asthma. METHODS: Thirty subjects with asthma included 14 current symptomatic asthmatics and 14 normal controls inhaled 4.5% hypertonic saline for 10 minutes by nebulizer. The expectorated sputum were collected from all asthmatics before and 3 hours after corticosteroid inhalation for children with asthma and were reduced by dithiotreitol. Total cell counts and differentials were determined. ECP was measured by CAP system. Interleukin-5, GM-CSF and albumin were measured by double sandwich ELISA. RESULTS: The mean eosinophil percentage and ECP in induced sputum of asthmatics were significantly higher than that of controls. The induced sputum samples obtained after glucocorticoid inhalation showed a significant reduction in mean eosinophil percentage, but FEV1, IL-5, GM-CSF, albumin, and ECP values were not significantly decreased. CONCLUSION: The present results in induced sputum may be interpreted to reflect direct steroid action on airways and lack of effect on bone marrow effectors at 3 hours after glucocorticoid inhalation.     

Oilseed rape flour: another allergen causing occupational asthma among farmers

BACKGROUND: Farmers are exposed to a wide variety of sensitizers. Since occupational asthma (OA) can lead to permanent disability, exposure discontinuation is the preferred treatment. When this is not possible, the identification of the causative allergen may allow an alternative therapy. METHODS: We present three farmers diagnosed with OA as a consequence of handling fodder. We carried out skin tests with common and occupational allergens and with oilseed rape (OSR) extract. Total and specific serum IgE levels were measured. The patients underwent the OSR-bronchial provocation test (OSR-BPT). The day before and 24 h after the OSR-BPT, the methacholine (M)-BPT and induced sputum were performed. Eosinophil percentages and ECP levels were measured in the sputum samples. RESULTS: OSR sensitization (skin tests and specific serum IgE) was detected in all the patients. The OSR-BPT elicited early responses in two subjects. Methacholine sensitivity, sputum eosinophils, and sputum ECP levels increased 24 h after the OSR-BPT in all the patients. CONCLUSIONS: We have demonstrated that inhalation of OSR flour causes bronchoconstriction, induces an eosinophilic inflammatory bronchial response, and increases bronchial hyperresponsiveness in sensitized asthmatics. OSR flour contained in animal fodder should be considered another potential cause of OA among farmers.     

Different airway responsiveness profiles in atopic asthma, nonatopic asthma, and Sjögren's syndrome. BHR Study Group. Bronchial hyperresponsiveness

Background : Different mechanisms may underlie bronchial hyperresponsiveness (BHR) in different diseases. The aim of this study was to investigate the bronchial responsiveness profile produced by three different challenge tests, methacholine, a direct simulus, and two indirect stimuli, adenosine 5′‐monophosphate (AMP) and cold air, in subjects with asthma and patients with Sjögren's syndrome. Methods : The study population comprised 40 adult patients with asthma, 18 subjects with Sjögren's syndrome, and 20 controls. Blood samples were collected before each challenge for measurements of serum eosinophil peroxidase (S‐EPO) and eosinophil cationic protein (S‐ECP). The investigated subjects recorded peak expiratory flow and kept a symptom diary. Results : Atopic subjects with asthma were significantly more hyperresponsive to AMP than nonatopic subjects with asthma (P=0.01) and subjects with Sjögren's syndrome (P=0.02). No difference was seen between atopic and nonatopic subjects with asthma in the case of challenges with methacholine or cold air. In atopic subjects with asthma, a significant correlation was found between challenges with methacholine and AMP (r=0.91, P=0.0001) and methacholine and cold air (r=0.83, P=0.004), but, in nonatopic subjects with asthma, no significant correlation was seen between methacholine and AMP or cold air challenges. In atopic subjects with asthma, the dose‐response slope for AMP was correlated to S‐EPO (r=?0.56; P=0.01) and S‐ECP (r=?0.51, P=0.02), while no correlation between BHR and inflammation markers was found in the two other patient groups. Conclusions : The results of this study suggest that patients with asthma and subjects with Sjögren's syndrome display different bronchial responsiveness profiles for different challenge agents. Atopic subjects with asthma are more hyperresponsive to AMP than nonatopic subjects and patients with Sjögren's syndrome. More than one challenge may be required to detect different aspects of bronchial responsiveness.     

Comparison of allergen-induced changes in bronchial hyperresponsiveness and airway inflammation between mildly allergic asthma patients and allergic rhinitis patients

Bronchial eosinophilic inflammation and bronchial hyperresponsiveness (BHR) are the main features of allergic asthma (AA), but they have also been demonstrated in allergic rhinitis (AR), suggesting a continuity between both diseases. In spite of not fully reproducing natural allergenic exposure, the allergen bronchial provocation test (A-BPT) has provided important knowledge of the pathophysiology of AA. Our aim was to verify the existence of a behavior of AA and AR airways different from the allergen bronchial challenge-induced airway eosinophilic inflammation and BHR changes. We studied a group of 31 mild and short-evolution AA and 15 AR patients, sensitized to Dermatophagoides pteronyssinus. The A-BPT was performed with a partially biologically standardized D. pteronyssinus extract, and known quantities of Der p 1 were inhaled. Peripheral blood (eosinophils and ECP) and induced sputum (percentage cell counts, ECP, albumin, tryptase, and interleukin [IL]-5) were analyzed, before and 24 h after A-BPT. Methacholine BHR, assessed before and 32 h after the A-BPT, was defined by M-PD20 values and, when possible, by maximal response plateau (MRP). The A-BPT was well tolerated by all the patients. AA presented a lower Der p 1 PD20 and a higher occurrence of late-phase responses (LPR). M-PD20 values decreased in AA, but not in AR, patients. MRP values increased in both groups. Eosinophils numbers and ECP levels increased in blood and sputum from both AA and AR, but only the absolute increment of sputum ECP levels was higher in AA than AR patients (P = 0.025). The A-BPT induced no change in sputum albumin, tryptase, or IL-5 values. We conclude as follows: 1) In spite of presenting a lower degree of bronchial sensitivity to allergen, AR patients responded to allergen inhalation with an eosinophilic inflammation enhancement very similar to that observed among AA. 2) MRP levels increased in both AA and AR patients after allergen challenge; however, M-PD20 values significantly changed only in the AA group, suggesting that the components of the airway response to methacholine were controlled by different mechanisms. 3) It is possible that the differences between AR and AA lie only in the quantitative bronchial response to allergen inhalation.     

Airway hyperresponsiveness: a story of mice and men and cytokines

Bronchial hyperresponsiveness (BHR) is an essential part of the definition of asthma. Although our understanding of the allergic inflammatory and immunologic mechanisms of asthma have markedly increased, the mechanism of BHR remains to be elucidated. Increased BHR is associated temporally with exposure to allergens, certain respiratory viruses, pollutants such as ozone, and certain occupational chemicals. An important research use of determining the degree of BHR to direct and indirect challenge is to determine the efficacy of pharmacologic and immunodulatory agents. Beta-adrenergic agents inhibit BHR and certain genetic polymorphisms of the beta-adrenergic receptor are associated with increased BHR. When beta-adrenergic receptors are blocked, sensitivity to allergens is markedly increased in patients with asthma and animal models of asthma. Allergen challenge and clinical asthma are associated with synthesis and release of pro-inflammatory cytokines such as IL-1 and TNF-alpha which have been shown to decrease the response to beta-agonists and increased the reactivity to methacholine and the airways neutrophils and alveolar macrophages. The Th2 cytokine IL-13 is increased in the airways of asthmatics and increases BHR in normal unsensitized animals. The mechanisms of this effect of IL-13 are being intensively investigated. Our group has shown that IL-13 induced BHR persisted for at least 7 days and the soluble receptor IL-13R2alpha protected against their BHR. Other investigators have demonstrated that IL-13 is necessary and sufficient for the induction of BHR and that eosinophilic airway inflammation in the absence of IL-13 fails to induce BHR. These studies indicate that treatment of human asthma with antagonists of IL-13 may be very effective.     

Airway hyperresponsiveness

Bronchial hyperresponsiveness (BHR) is an essential part of the definition of asthma. Although our understanding of the allergic inflammatory and immunologic mechanisms of asthma have markedly increased, the mechanism of BHR remains to be elucidated. Increased BHR is associated temporally with exposure to allergens, certain respiratory viruses, pollutants such as ozone, and certain occupational chemicals. An important research use of determining the degree of BHR to direct and indirect challenge is to determine the efficacy of pharmacologic and immunodulatory agents. Beta-adrenergic agents inhibit BHR and certain genetic polymorphisms of the beta-adrenergic receptor are associated with increased BHR. When β-adrenergic receptors are blocked, sensitivity to allergens is markedly increased in patients with asthma and animal models of asthma. Allergen challenge and clinical asthma are associated with synthesis and release of pro-inflammatory cytokines such as IL-1 and TNF-α which have been shown to decrease the response to β-agonists and increased the reactivity to methacholine and the airways neutrophils and alveolar macrophages. The Th2 cytokine IL-13 is increased in the airways of asthmatics and increases BHR in normal unsensitized animals. The mechanisms of this effect of IL-13 are being intensively investigated. Our group has shown that IL-13 induced BHR persisted for at least 7 days and the soluble receptor IL-13R2α protected against their BHR. Other investigators have demonstrated that IL-13 is necessary and sufficient for the induction of BHR and that eosinophilic airway inflammation in the absence of IL-13 fails to induce BHR. These studies indicate that treatment of human asthma with antagonists of IL-13 may be very effective.     

Effects of allergen inhalation and oral glucocorticoid on serum soluble CTLA-4 in allergic asthmatics

BACKGROUND: The serum soluble cytotoxic T lymphocyte associated antigen-4 (sCTLA-4) concentration is significantly elevated in patients with asthma, and sCTLA-4 concentration correlate with the severity of asthma. The aim of the present study was to investigate effects of allergen inhalation and oral glucocorticoid on concentration of serum sCTLA-4 in patients with allergic asthma. METHODS: Allergen inhalation challenge was conducted in allergic asthmatics with isolated early asthma response and those with dual asthma response. In a randomized, double-blind, placebo-controlled, parallel group fashion, prednisolone or placebo was give orally once a day for 2 weeks. Venous blood samples were collected before and after allergen inhalation or prednisolone administration for obtaining sera. The serum sCTLA-4 concentrations were determined using enzyme-linked immunosorbent assay. RESULTS: The serum sCTLA-4 concentrations in the dual responder group increased from 29.0 (14.5-43.7) microg/l [median (25-75 percentiles)] before allergen inhalation to 44.0 (24.3-61.3) microg/l 24 h after allergen inhalation. In the isolated early responders, there were no significant increase in serum sCTLA-4 concentrations after allergen inhalation compared with baseline levels. There was a significant decrease in serum sCTLA-4 concentrations after 2 weeks of glucocorticoid therapy [22.0 (15.5-31.0) microg/l] compared with baseline values [37.0 (19.5-53.0) microg/l], whereas there was no significant difference in the placebo group. CONCLUSION: This study has demonstrated that serum sCTLA-4 concentrations increased after allergen inhalation in sensitized asthmatic subjects, and that serum sCTLA-4 concentrations were downregulated by prednisolone therapy.     

Eosinophil cationic protein(ECP) and eosinophil protein X (EXP) concentrations in serum and bronchial lavage fluid in asthma. effect of prednisolone treatment.

Background Eoswinophil granule proteins may contribute to hyperresponsiveness in asthma.
Objective To measure eosinophil cationic protein (ECP) and eosinophil protein X (EPX) in sereum and bronchial lavage fluid from 20 asthmatics and 16 control subjects. To asses the effect on these eosinophil proteins of corticosteroid treatment of asthma. To determine ehether serum ECP and EPX measured weekly in a longitudina study for 10 weeks reflected changes in lung function.
Methods Eosinophil granule proteins were measured by radiommunoassy of bronchial wash (BW), bronchoalveolar lavage (BAL) serum.
Results Eosinophils were elevated in BAL (P<0.01) , BW (P<0.01) and blood (P<0.01) from asthmatic compared with control subjects. Eosinophil cationic protein concentration was significantly elevated in BAL (P<0.05) and BW from asthmatics (P<0.01) and EPX was increased in BAL (P<0.05) and BW (P<0.01) . These changes were also reflected in elevated serum ECP (P<0.01) and EPX (P<0.01) concentrations is asthmatic subjects. There was no significant difference between sujects receiving prednisolone and the placebo group, but there was a fall in ECP in BW (P<0.05) and serum (P<0.01) and in EPX in BW (P<0.01) and serum (P<0.01) within the group receiving prednisolone. In the longitudinal study there was only significant difference between ECP values associated with highest and lowest peak expiratory flow rate (PEFR) (P<0.05).
Conclusion These data confirm a role for cosinophil activation in the airway in asthma pathogenesis, and add some support to the hypothesis that corticosteroids may inhibit cosinophil activation in asthma.     

Nitric oxide metabolites in induced sputum: a marker of airway inflammation in asthmatic subjects

BACKGROUND AND OBJECTIVE: The role of nitric oxide (NO) needs to be further clarified in allergic inflammation. This study was designed to investigate the relationships between NO metabolites and eosinophil count, eosinophil cationic protein (ECP), interleukin (IL)-5 in induced sputum from asthmatics. METHODS: Hypertonic saline-induced sputum was obtained in 25 asthmatic subjects, among which 13 patients were examined before and after anti-asthmatic medications including steroid preparations. Ten normal subjects were enrolled as controls. Fresh expectorated sputum separated from saliva was treated with equal volume of dithiothreitol 0.1%, cytospinned for cell count, and the supernatant was collected for biochemical assay. NO metabolites were assayed by using modified Griess reaction. ECP was measured by fluoroimmunoassay, and detected IL-5 by a sandwich ELISA. RESULTS: Asthmatic subjects, compared with controls, had significantly higher concentration of NO metabolites (1035.4 +/- 125.3 vs 557.2 +/- 101.5 micromol/L, P < 0.01), higher percentage of eosinophils (25.6 +/- 4.6 vs 1.7 +/- 0.2%, P < 0.01), and higher levels of ECP (1117.8 +/- 213.9 vs 154.6 +/- 47.4 microg/L, P < 0.01) in the induced sputum. IL-5 was detected more frequently in asthmatic subjects than in control subjects (11/25 [44%] vs 1/10 [10%], P < 0.05). According to asthma severity, moderate to severe asthmatic subjects (n = 18) had higher level of NO metabolites (1143.8 +/- 156.3 vs 575.5 +/- 89.5 micromol/L, P < 0. 01), higher levels of ECP and IL-5 (P < 0.01, respectively) in the induced sputum than in those of mild asthmatic subjects (n = 7). NO metabolites, the percentage of eosinophils, the levels of ECP, and IL-5 were reduced following treatment with anti-asthmatic drugs (P < 0.01, respectively). There were significant positive correlations between NO metabolites and percentage of eosinophils or ECP (r = 0. 34, P < 0.05; r = 0.28, P < 0.05). Negative correlations were noted between FEV1, FEV1/FVC and proportion of eosinophils, ECP, or IL-5 levels. CONCLUSION: These findings confirmed that the level of NO metabolites was increased in the tracheobronchial secretion of asthmatic subjects and was paralleled with severity of asthma. Measurement of NO metabolites in induced sputum may be used for monitoring the degree of airway inflammation in asthmatics.     

Allergen challenge primes for IL-5 mRNA production and abrogates β-adrenergic function in peripheral blood T lymphocytes from asthmatics

BACKGROUND: In previous studies, we have found a dysfunctional adenylyl cyclase (AC) system in patients with asthma after allergen provocation, which resulted in a 40-50% decreased generation of intracellular cAMP. In addition, in activated T helper lymphocyte clones, it has been demonstrated that IFN-gamma (TH1-like cytokine) and IL-5 (TH2-like cytokine) are differentially regulated by the AC system. Therefore, we postulate that an increased IL-5/IFN-gamma ratio as observed in asthmatics might be due to a dysfunctional AC system. OBJECTIVE: To assess whether a dysfunctional AC system as observed in asthmatics after allergen provocation, is responsible for an increased IL-5/IFN-gamma cytokine ratio. METHODS: Peripheral blood T lymphocytes of seven asthma patients were stimulated with anti-CD3 plus anti-CD28 MoAbs in the absence and presence of isoproterenol (ISO) and prostaglandin E2 (PGE2) to activate the AC system. Before, 3 h and 24 h after allergen provocation, IFN-gamma and IL-5 mRNAs were detected by semiquantitative RT-PCR. RESULTS: Before allergen provocation, ISO (10-5 mol/L) significantly downregulated IFN-gamma mRNA (P < 0.03, n = 6), and showed a trend to upregulate IL-5 mRNA (P = 0.138, n = 5). Three and 24 h after allergen provocation, ISO was not longer able to modulate IFN-gamma and IL-5. In contrast with the observations with ISO, PGE2 still dose-dependently inhibited IFN-gamma mRNA, both before, 3 h and 24 h after allergen provocation (n = 7). IL-5 mRNA, but not IFN-gamma mRNA, was significantly upregulated in anti-CD3 plus anti-CD28-activated T cells (P < 0.05, n = 5) 24 h after allergen provocation, compared with before allergen provocation. CONCLUSION: Twenty-four hours after allergen provocation, a significant reduction of beta-adrenergic control on IFN-gamma and IL-5 mRNA expression was observed in peripheral blood T lymphocytes, which coincides with a selective priming of IL-5 mRNA production.     

Bronchial hyper-responsiveness to hypertonic saline and blood eosinophilic markers in 8–13-year-old schoolchildren

BACKGROUND: In adult asthma, bronchial hyper-responsiveness (BHR) to indirect stimuli reflects eosinophilic activation more closely than BHR to stimuli that directly cause smooth muscle contraction. AIM: To assess the relationship between BHR to the indirect stimulus hypertonic saline (HS), blood eosinophil numbers, and serum eosinophilic cationic protein (ECP) in children with and without current wheeze. METHODS: A cross-sectional survey among 8-13-year-old schoolchildren, using the International Study of Asthma and Allergic disease in Childhood questionnaire, bronchial challenge with HS, skin prick tests, serum IgE, blood eosinophil counts and ECP (in a subset). Based upon the presence of current wheeze (WHE) and BHR, we defined three case groups (WHE+BHR+, WHE-BHR+, WHE+BHR-) and the reference group (WHE-BHR-). By regression analyses, each case group was compared with the reference group for differences in atopic sensitization, blood eosinophil counts and serum ECP. RESULTS: Complete data were obtained for 470 children. BHR was present in 103 children (22%), 66 being asymptomatic and 37 symptomatic. Children of all three case groups were more often atopic. Sensitization to indoor allergens particularly occurred in children with BHR, irrespective of symptoms (P < 0.05). Children with WHE+BHR+ had highest values for blood eosinophils and serum ECP (P < 0.05). Children with WHE-BHR+ had less severe responsiveness. In atopic children with WHE-BHR+, serum ECP was higher than in children with WHE-BHR-(P < 0.05). CONCLUSIONS: BHR to HS is associated with blood markers of eosinophilic activation, particularly in atopic children.