Total blood eosinophils, serum eosinophil cationic protein and eosinophil protein X in childhood asthma: relation to disease status and therapy

Blood eosinophils, and serum levels of the eosinophil proteins, eosinophil cationic protein (ECP) and eosinophil protein X (EPX) were measured in childhood asthma. Seventeen patients mean age 11.9 years who were symptomatic with asthma, were enrolled in a study examining the eosinophil counts and eosinophil proteins at the onset of study and after treatment in relation to changes in their baseline forced expiratory volume at 1 second (FEV1) and % predicted FEV1. The patients with symptomatic asthma were compared with 17 patients mean age 12.0 years with asymptomatic asthma maintained on daily inhaled steroid and 13 patients, mean age 12.0 years, without asthma but with urticaria who served as non-asthma controls. Patients with symptomatic asthma did not have significantly higher initial eosinophil counts compared with those with asymptomatic asthma (0.43 x 10(9)/l vs 0.26 x 10(9)/l, P = 0.09) but had higher serum ECP levels (28.9 micrograms/l vs 18.5 micrograms/l). Both asthma patient groups had significantly higher serum ECP levels (P < 0.01) than the controls (9.8 micrograms/l). After therapy consisting of increased dose of inhaled steroids and/or oral steroids, patients in the symptomatic asthma group demonstrated a significant rise in FEV1 (1.67 l/sec at Visit 1 vs 2.08 l/sec at Visit 2, P < 0.001). A similar rise was seen for % predicted FEV1.(ABSTRACT TRUNCATED AT 250 WORDS)     

Serum eosinophil cationic protein in infants during first wheezing episode

We measured serum ECP levels in infants during first wheezing episode. Serum ECP in these infants are significantly higher than in control infants, although much higher in children with asthma. Serum ECP in these infants with high serum IgE and/or positive RAST score are higher than in infants with normal serum IgE and negative RAST score. In children with bronchial asthma serum ECP is correlated with peripheral eosinophil counts, but in infants during first wheezing episode serum ECP is often elevated not associated with increased peripheral eosinophil counts. These suggest that activated eosinophils could be responsible for bronchoconstriction in wheezing patients with atopic diathesis even in very early phase and that these eosinophilic inflammations could contribute to formation of increased airway reactivity and bronchial asthma.     

Induced sputum eosinophil cationic protein (ECP) measurement in asthma and chronic obstructive airway disease (COAD)

BACKGROUND: Induced sputum is a useful way to monitor airway inflammation in asthma, but cell counts are time-consuming and labour intensive. OBJECTIVE: The aim of this study was to evaluate a novel processing method using eosinophil cationic protein (ECP) as a biochemical marker of sputum eosinophil number and activation in subjects with asthma and other airway diseases. METHODS: Sputum was dispersed with dithiothreitol and centrifuged to yield cell free supernatant and a cell pellet. The pellet was treated with a cellular lysis buffer to release cell-associated ECP. ECP was measured in sputum supernatant and in the lysed cell pellet and was compared with sputum eosinophil counts in 31 adults with asthma, chronic obstructive airway disease (COAD), bronchiectasis and healthy controls. The ratio of supernatant to pellet ECP was evaluated as an index of eosinophil degranulation. The effect of sputum processing reagents and storage time on ECP measurement was also evaluated. RESULTS: ECP measured in the cell pellet lysate correlated closely with sputum absolute eosinophil counts across a range of subject groups (r = 0.72, P = 0.004). Sputum eosinophil counts were less well correlated with supernatant ECP levels (r = 0.54, P < 0.05). Incubation with dithiothreitol or lysis buffer did not influence ECP measurement and sputum ECP levels were stable over a 6-9 month period. Sputum supernatant and pellet lysate ECP concentrations were increased in stable asthma, asthma exacerbations and COAD/bronchiectasis (P < 0.05). The ratio of supernatant to pellet ECP was used as an index of eosinophil degranulation and found to be elevated in asthma exacerbations, COAD and bronchiectasis, but not in stable asthma. CONCLUSION: The measurement of ECP in the sputum cell pellet provides a reliable and efficient estimate of sputum eosinophil counts which can potentially be used in clinical trials and epidemiological surveys. The ECP ratio may be a useful marker of eosinophil activation, and was increased in asthma exacerbation and COAD. The increased ECP in COAD reflects a non-selective accumulation of eosinophils in this condition.     

Effect of a bed covering system in children with asthma and house dust mite hypersensitivity

Allergen avoidance is regarded as an important approach to management of atopic asthma. The effect of Intervent bed covering systems on house dust mite (HDM) allergen exposure, asthma symptoms and markers of inflammation was investigated in 31 HDM sensitive asthmatic children. Dust concentrations of Dermatophagoides pteronyssinus allergen 1 (Der p 1) were monitored before and after covering the mattress, duvet and pillow with active and placebo covers for 3 months, in a single-blind, cross-over trial. Twice daily peak expiratory flow rate (PEFR), daily symptom scores and treatment schedule were recorded. Bronchial hyperresponsiveness was monitored by histamine challenge (provocative concentration of histamine causing a 20% fall in forced expiratory volume in one second (PC20)), and inflammation by measuring eosinophil cationic protein (ECP), eosinophil protein X (EPX), eosinophil peroxidase (EPO), and soluble interleukin-2 receptor (sIL-2R) in serum. There was a significant reduction in Der p 1 when the mattress, duvet and pillow were covered with the active bedding. There was no significant improvement in symptoms of asthma, PEFR, bronchodilator usage of PC20. Also, ECP, EPX, sIL-2R concentrations did not change for either treatment. EPO concentrations were significantly lower in the active compared to the placebo period. The active bed covers reduced retrievable Dermatophagoides pteronyssinus allergen 1 (Der p 1) from the bedding, with short term clinical benefit.     

Serum levels of eosinophil cationic protein and eosinophil protein x in pollen atopic patients with stable asthma and its relation with bronchial hyperresponsiveness

Eosinophils are important effector cells in allergic inflammation described in allergic rhinitis (AR) and allergic bronchial asthma (BA). During the pollen season serum levels of eosinophil cationic protein (ECP) and eosinophil X protein/eosinophil-derived neurotoxin (EPX/EDN) are increased in BA. The aim of the present study was to evaluate the serum levels of ECP and EPC in pollen atopic patients with AR and BA during the winter. 92 patients were studied. They were divided into three groups: I 29 patients with AR, II 51 patients with BA and III 12 healthy subjects. Allergic rhinitis and bronchial asthma were diagnosed by routine clinical tests: clinical history, skin tests, total IgE and specific IgE. In addition ECP and EPX were determined in serum. All patients were asymptomatic, stable and without medical treatment. Methacholine challenge test (MCT) was performed in all patients. MCT were positive in 4 patients of group I and 45 patients of group II. ECP levels (ug/l) were: 21 (I), 24 (II) and 7 (III). EPX levels (ug/l) were 35 (I), 45 (II) and 21 (III). Statistical differences (p < 0.01) were observed both in ECP and EPX levels in patients with MCT positive in relation to patients with MCT negative, and in allergic patients (I and II) in comparison with the healthy subjects (III) (p < 0.01). ECP and EPX serum levels are increased in patients with a positive MCT in the winter, out of the pollen season, when patients are asymptomatic, stable and without treatment. This fact suggests that eosinophils play an important role in the pathogenesis of bronchial asthma.     

Treatment of recurrent corneal erosion using phototherapeutic keratectomy with the excimer laser

Tachyphylaxis to methacholine has been reported in nonasthmatic subjects. In a recent study on the prevalence of airway hyperresponsiveness (AHR) and atopy, we performed duplicate methacholine inhalation tests at a 60-min interval, in subjects with symptomatic asthma (n = 33), asymptomatic AHR (AAHR) (n = 72) and in a group of normal subjects (n = 130); 135/235 subjects were atopic. All subjects had a respiratory questionnaire, allergy skin prick tests, blood eosinophil counts and determination of total serum IgE level. In asthmatic subjects, PC20 just failed to be significantly higher on a second methacholine challenge (p = 0.09); when they were stratified according to severity of AHR and use of inhaled corticosteroids, we observed a significant increase in PC20 on the second test in asthmatic subjects with mild AHR not using corticosteroids (p < 0.01). In normal controls, PC20 methacholine was slightly increased on rechallenge (p < 0.01) as it was in those with AAHR (p < 0.01). There was no relationship between the magnitude of the change in PC20 and age, sex, baseline airway responsiveness, percent fall in FEV1 on the first challenge, atopic score, blood eosinophil counts and serum IgE levels. In conclusion, tachyphylaxis to methacholine is observed in normal or mild asthmatic subjects not using inhaled corticosteroids and in subjects with AAHR; however, in most subjects this change is of a small magnitude.     

Suppressive effect of corticosteroids on the gene expression of interleukin-5 and eosinophil activation in asthmatics

Although cortico steroids are effective anti-inflammatory agents in ameliorating asthma symptoms and bronchial hperreactivity, their mechanism of action is unknown. Interleukin (IL)-5 is known to play a key role in regulating eosinophil proliferation and activation. Therefore, we examined the changes of IL-5 mRNA expressions in PBMC semi-quantitatively with RT-PCR as well as serum ECP levels and MCH-PC20 values in asthmatics before and after being treated with corticosteroids. The results revealed that there were significant decrease in the level of IL-5 mRNA and serum ECP concentration after therapy (P < 0.05) and there was remarkable improvement in the values of MCH-PC20 and FEV1% (P < 0.05). It was also found that the changes of serum ECP levels or MCH-PC20 values were accompanied by a reduction in IL-5 mRNA expression (r = 0.5426 or 0.4857, P < 0.05). These results suggested that the therapeutic effects of corticosteroids in asthma may result from modulation of IL-5 gene expression with consequent inhibition of eosinophil activation.     

Long-lasting sensitization to food during the first two years precedes allergic airway disease. The MAS Study Group, Germany

Measurement of eosinophil percentages and ECP concentration in induced sputum may be useful in the diagnosis and assessment of the variability of airway inflammation in bronchial asthma (BA). To evaluate the usefulness of sputum eosinophil counts and ECP concentrations in the diagnosis of BA, we measured these parameters in 68 patients with respiratory complaints. In addition, we followed-up 14 BA patients with variable airflow limitation for 45.4 +/- 10.4 days. The BA group (n = 41) showed a higher percentage of sputum eosinophilia (24.5 +/- 7.6 vs. 2.2 +/- 2.9%, p < 0.001) and a higher level of sputum ECP (198.2 vs. 90.6 micrograms/L, p < 0.05) than those in the nonasthmatic group (NBA, n = 27). The sensitivity and specificity of sputum eosinophilia (> or = 5%) for the diagnosis of BA were 85.4% and 92.6%, respectively, which were better than the sensitivity (68.3%) and specificity (55.5%) of the increased level of sputum ECP (> or = 100 micrograms/L). Patients with moderate-to-severe persistent BA had a higher percentage of sputum eosinophil (n = 23, 34.6 +/- 10.6%) than those of mild persistent BA (n = 18, 10.7 +/- 5.2%, p < 0.01), but we could not find significant difference in ECP levels between mild persistent and moderate-to-severe persistent asthma. The percentages of sputum eosinophilia showed a moderate correlation with ECP (r = 0.4358, p < 0.01) and with the peak expiratory flow rate (PFR, r = -0.4746, p < 0.01) but sputum ECP did not correlate with PFR. In 14 BA patients who were followed, there was a relationship between changes of PFR and the percentage of sputum eosinophil (r = -0.7238, p < 0.01), but the change of PFR did not correlate with the change of sputum ECP levels. These results suggest that the sputum eosinophil count and sputum ECP level could be helpful in the diagnosis of BA, but that sputum ECP is not satisfactory for the assessment of variability of airway eosinophilic inflammation during the initial anti-inflammatory management of BA.     

Induction of leaves directly from leaves in the maize mutant Lax midrib1-O

BACKGROUND: Asthma exacerbations are closely associated with respiratory virus infections. However, the pathophysiological consequences of such infections in asthma are largely unclear. OBJECTIVE: To examine the effect of rhinovirus 16 (RV16) infection on airway hypersensitivity to histamine, and on interleukin-8 (IL-8) in nasal lavage. METHODS: Twenty-seven non-smoking atopic, mildly asthmatic subjects participated in a placebo-controlled, parallel study. A dose of 0.5-2.9 x 10(4) TCID50 RV16 or placebo was nasally administered. Cold symptoms were recorded by questionnaire throughout the study. Histamine challenges were performed at entry, and on days 4 and 11 after inoculation. Nasal lavages were obtained at entry, and on days 2 and 9. The response to histamine was measured by PC20 (changes expressed as doubling doses: DD) IL-8 levels were obtained by ELISA, and were expressed in ng/ml. RESULTS: RV infection was confirmed by culture of nasal lavage and/or by antibody titre rise in each of the RV16-treated subjects. Among the 19 RV 16-treated subjects, eight developed severe cold symptoms. Baseline FEV1, did not change significantly during the study in either treatment group (P = 0.99). However, in the RV16-treated subjects there was a decrease in PC20 at day 4, which was most pronounced in those with a severe cold (mean change +/- SEM: -1.14 +/- 0.28 DD, P = 0.01). In addition, IL-8 levels increased in the RV16 group at days 2 and 9 (P < 0.001). The increase in nasal IL-8 at day 2 correlated significantly with the change in PC20 at day 4 (r = -0.48, P = 0.04). CONCLUSION: We conclude that the severity of cold, as induced by experimental RV16 infection, is a determinant of the increase in airway hypersensitivity to histamine in patients with asthma. Our results suggest that this may be mediated by an inflammatory mechanism, involving the release of chemokines such as IL-8.     

Relaxin activates the L-arginine-nitric oxide pathway in vascular smooth muscle cells in culture

The effects of usual or low doses of inhaled corticosteroids on airway mucosal inflammation have not yet been examined. We therefore, compared the effects of inhaled beclomethasone dipropionate (BDP) 336 microg x day(-1) on asthma control outcomes and markers of airway inflammation. Twenty-four adult subjects with mild and moderate asthma were randomized to receive either BDP or placebo for four weeks; then subjects entered a single blind four week placebo run-in period. We found that the BDP group had significantly greater improvements in forced expiratory volume in one second (FEV1), morning peak flow, and rescue salbutamol use than the placebo-treated group. The improvement in FEV1 largely reversed one week after treatment was stopped. The decrease in the median percentage of eosinophils in induced sputum in the BDP group from 3.8% to 3.4% was not significant, but because eosinophils increased from 8.4% to 12.7% in the placebo group, there was a significant difference between treatment groups (p=0.03). There was no significant difference between groups during treatment in the levels of eosinophil cationic protein (ECP), tryptase mucin-like glycoprotein, or fibrinogen in induced sputum. The change in FEV1 in the BDP group did not correlate significantly with the change in eosinophil percentage or ECP levels. We concluded that four weeks of treatment with inhaled beclomethasone dipropionate 336 microg x day(-1) was associated with significant improvements in peak flow, forced expiratory volume in one second, and rescue salbutamol use in asthmatic subjects but was not associated with large reductions in markers of eosinophilic inflammation, bronchovascular permeability, or mucus hypersecretion.     

Chemotherapy as a part of each treatment fraction in a twice-a-day hyperfractionated schedule: a new chemoradiotherapy approach for advanced head and neck cancer

In allergic asthma, there is convincing evidence that changes in eosinophil and lymphocyte state of activation in blood may reflect disease activity. We evaluated whether simple blood eosinophil or lymphocyte counts in atopic children with asthma could reflect the degree of allergic sensitization. Seventy-six asthmatic children, sensitized to house dust mites (HDM), in stable conditions at the time of the study, and 53 sex- and age-matched controls (CTR) were studied. As compared to CTR, allergic patients showed higher eosinophil numbers and percentages (p < 0.001) but similar lymphocyte numbers and proportions (p > 0.1). Both in CTR and in allergic patients, eosinophil counts did not correlate with lymphocyte counts (p > 0.05; each comparison) but positive correlations were observed between eosinophil numbers and percentages and paper radio immunosorbent test (PRIST) levels or radio-allergo sorbent test (RAST) classes (p < 0.001; each comparison). When allergic asthmatic individuals were subdivided according to their age into two subgroups (Gr), no differences were found in eosinophil and lymphocyte counts and in PRIST levels and RAST values between Gr1 (< or =5 years old [preschool children]) and Gr2 (>5 years old [school children]) (p > 0.05; each comparison). Interestingly, although positive correlations between eosinophil counts and PRIST levels were found in both subgroups (p < 0.05; each comparison), only in Gr2 did eosinophil counts correlate positively with RAST classes (p < 0.001). No correlations between lymphocyte counts and PRIST levels or RAST classes were demonstrated (p > 0.05; each comparison). These data suggest that although blood eosinophilia was similar in preschool and in allergic asthmatic school children sensitized to HDM, only in the oldest children did blood eosinophil counts appear to be related to the degree of HDM-specific sensitization.     

Double-blind, placebo-controlled evaluation of sublingual immunotherapy with standardized olive pollen extract in pediatric patients with allergic rhinoconjunctivitis and mild asthma due to olive pollen sensitization

For evaluation of the efficacy and the safety of specific sublingual immunotherapy with high allergen dose, 66 children with seasonal asthma, rhinitis, and conjunctivitis due to sensitization to olive pollen were enrolled in a double-blind, randomized, placebo-controlled study between October 1994 and October 1996 in Greece. Thirty-four patients were randomly allocated to the active group, and 32 received placebo. Immunotherapy consisted of olive-allergen extracts (Stallergènes SA) administered sublingually pre- and coseasonally from January to July for 2 consecutive years. Serial concentrations from 1 to 300 IR. were used up to the maintenance dose of 20 drops of 300 IR daily. The cumulative dose for each patient was 300 times higher than in parenteral immunotherapy, and the cumulative dose of the major allergen Ole e 1 was 8.1 mg/2 years. The patients were assessed by clinical parameters (symptom and medication scores from patients' daily diaries) and immunologic measurements (specific IgE, IgG4, eosinophil cationic protein [ECP]) were performed. The actively treated patients had a significantly lower score for dyspnea (P<0.04 during the first season; P<0.03 during the second season). At the pollinic peak during the second year, a lower score of conjunctivitis was recorded (P<0.05) in the actively treated patients. The analysis of intragroup evolution showed that the total score of rhinitis increased significantly during the pollinic peak in the group under placebo, whereas there was no symptomatic peak for the same period in the group under active treatment. However, the difference between the groups was not significant. The medication score did not differ significantly between the groups. Oral steroids were the only variables with a P value near the significance level (P=0.06) in favor of the actively treated group. A significant decrease in skin reactivity was recorded in the active group after 2 years of treatment. No significant variation in specific IgE and IgG4 was detected. A significantly lower level of serum ECP was observed at the pollinic peak in the actively treated patients during the first pollen season (P=0.01), but this was not confirmed the second year when the ECP levels doubled in both groups without correlation to the clinical findings. Tolerance was excellent with only a few minor side-effects reported. In conclusion, high-dose specific sublingual immunotherapy appears to be safe and effective in improving mild seasonal asthma and conjunctivitis linked to olive-pollen sensitization.     

Effects of ketotifen on symptoms and on bronchial mucosa in patients with atopic asthma

Ketotifen is marketed throughout the world as an antiallergy drug, but whether it affects infiltration of inflammatory cells into airway mucosa is not known. We studied the effects of ketotifen on symptoms, pulmonary function, and airway inflammation in 25 patients with atopic asthma. Patients took ketotifen (1 mg twice daily) or a matching placebo for 8 weeks in a double-blind, parallel-group study. Data recorded on diary cards were used for 2 weeks before treatment began, and they were used for the last 2 weeks of treatment to study asthma symptoms, use of beta 2-agonists, and peak expiratory flow (PEF). Pulmonary function tests, bronchial responsiveness to methacholine, and fiberoptic bronchoscopy were performed before and after treatment. Biopsy specimens were obtained by bronchoscopy. Specimens were stained immunohistochemically with monoclonal antibodies against stored eosinophil cationic protein (EG1), the secreted form of eosinophil cationic protein (EG2), mast-cell tryptase (AA1), neutrophil elastase (NP57), CD3, CD4, CD8, and CD25. The numbers of positively stained cells in the lamina propria were counted. Compared with the placebo, the ketotifen-treated group exhibited significant improvement of asthma symptoms (P < 0.05) and bronchial responsiveness (P < 0.05). This was accompanied by a reduction of EG2+ eosinophils (P < 0.05), CD3+ T cells (P < 0.001), CD4+ T cells (P < 0.01), and CD25+ activated T cells (P < 0.01) in the bronchial mucosa. These results suggested that the beneficial effects of ketotifen in bronchial asthma may result from consequent inhibition of activated eosinophils and T-cell recruitment into the airway. Moreover, ketotifen may relieve allergic inflammation in bronchial asthma.     

Eosinophil cationic protein in the asthmatic infant: correlation with the clinic and pulmonary function

PATIENTS AND METHODS: Serum eosinophil cationic protein (ECP) was measured in 99 chronic asthmatic patients (51 males and 48 females) with a mean age of 10.59 years and correlated with the number of eosinophils, lung function, symptoms in the last 6 months and clinical scoring (that reflecting the clinical situation during the last 15 days). RESULTS: Serum ECP showed a significant correlation with the total number of eosinophils (p < 0.001, R = 0.44), clinical scoring (p < 0.05, R = 0.26), number of inhaled beta 2-agonist doses needed in the last 15 days (p < 0.05, R = 0.26), forced expiratory volume during 1 second (FEV1; p < 0.01, R = -0.27), forced vital capacity (FVC; p < 0.05, R = -0.23), maximal mid-expiratory flow (FEF25-27; p < 0.001, R = -0.37). However, there was no significant correlation between the total number of eosinophils and the clinical situation of the children or the FEV1, but we found a significant correlation with the FEF25-27. Patients with ECP < 20 had better results on lung function tests than patients with ECP > 20 (FEV1: 108.89 +/- 17.7 vs 100.5 +/- 22 (p < 0.05), FEF25-27: 93.81 +/- 24.4 vs 75.21 +/- 24.5 (p < 0.001). CONCLUSIONS: The findings of this study suggest that the ECP level is a good marker of the situation of asthma in childhood. The levels of ECP will probably be able to help us to evaluate the degree of bronchial inflammation that neither the clinical state nor the lung function define completely.     

Biliary excretion in persons with low blood glutathione levels

To examine their possible predictive value for the development of asthma, the serum concentration of eosinophil cationic protein (ECP) and the total eosinophil count were measured at admission in 25 children aged 1-17 months hospitalized for their first episode of bronchiolitis. After an average of three years the parents of 23 index patients answered a questionnaire to determine development of asthma. Eight children were defined as having asthma at follow-up based on at least three episodes of wheezing. The remaining 15 children had experienced only one or two episodes of wheezing, and all of these children had been wheeze free for the last year. The serum concentrations of ECP were similar in children who subsequently developed asthma (8.0 microg/l; 3.6 to 14.2 (median; quartiles)) and in those who did not (12 microg/l; 4.5 to 16.8). Moreover, the total eosinophil counts were similar in asthmatic (0.10 x 10(9)/l; 0.04 to 0.20) and non-asthmatic patients (0.09 x 10(9)/l; 0.02 to 0.13). In conclusion, our study suggest that neither the serum concentration of ECP nor the total eosinophil count can predict the development of asthma when measured in children admitted for their first episode of bronchiolitis, but larger studies need to be carried out to confirm these results.     

Eosinophilic inflammation in cough variant asthma

Eosinophils are considered to play a central pathogenetic role in asthma. We previously reported that sputum eosinophilia was observed in patients with cough variant asthma (CVA), as well as in "classic" asthma with wheezing. This study was undertaken to further investigate the involvement of eosinophils in CVA. The serum eosinophil cationic protein (ECP) level, the percentage of eosinophils in bronchoalveolar lavage (BAL) fluid, and the number of eosinophils in bronchial biopsy specimen were examined in 14 patients with CVA, 21 with classic asthma, and in seven healthy controls. For the two asthmatic groups, the clinical severity was classified with scores of 1-3. Pulmonary function and bronchial responsiveness were not significantly different between the patients with classic asthma and those with CVA. BAL, tissue eosinophil and serum ECP were all significantly increased in both classic asthma and CVA when compared with the controls but were not different between classic asthma and CVA. In both groups of asthmatics, the clinical severity significantly correlated with serum ECP and tissue eosinophils. In conclusion, eosinophilic inflammation is involved in cough variant asthma as well as in classic asthma. Anti-inflammatory treatment may be essential in patients with CVA, as in those with classic asthma.     

The effect of theophylline on blood and sputum eosinophils and ECP in patients with bronchial asthma

It was recently reported that theophylline has an anti-inflammatory and bronchodilating effect on bronchial asthma. Accordingly, to examine the anti-inflammatory effect of theophylline on asthma, especially its effect on eosinophil activation, a sustained-release theophylline preparation (Theolong) was administered (daily dose: 400 mg) to 18 patients with mild to moderate bronchial asthma. This was done in order to study the preparation's effects on lung function, blood and sputum eosinophils and ECP four weeks pre- and post-administration. Lung function was determined by spirometry and sputum by induced sputum. Blood and sputum ECP levels were determined using an ECP RIA kit. In lung function, there were no differences in vital capacity (VC) or in forced expiratory volume 1 s (FEV 1.0) pre- and post-administration. There were also no differences in the number of blood and sputum eosinophils, but serum and sputum ECP levels decreased. Theophylline is thus expected to exert an inhibitory effect on eosinophil activation and it is suggested as an effective therapeutic drug for bronchial asthma.     

Peak expiratory flow rate (PEF) and serum eosinophil cationic protein (ECP) changes in nocturnal asthmatics

We have investigated the serum ECP and peak expiratory flow rate in 20 patients with nocturnal asthma. Changes of PEF were measured in every 2 hours around the whole day, and the blood samples were obtained at 4:00 and 16:00 to measure the serum ECP level and the peripheral Eo numbers. In addition, 10 asthmatics as well as normal subjects received methacholine challenge at 4:00 and 16:00. It was found that the PEF reached the lowest point at 4:00 and obviously less than that at 16:00 (187.50 +/- 120.31 L/min vs 313.00 +/- 108.14 L/min, P < 0.05), that the airway reactivity at 4:00 was significantly higher than at 16:00 (P < 0.05) and the difference of MCH-PC20 between the two time points was 0.34 +/- 0.31 mg/ml, that the serum ECP level at 4:00 was obviously higher than that at 16:00 (11.14 +/- 7.40 micrograms/ml vs 5.49 +/- 4.12 micrograms/ml, P < 0.05). The change rate of PEF markedly related to the change of serum ECP between the two time points (r = 0.61, P < 0.05). The findings suggested that the activation of Eo and its release of ECP might be effect of the circadian-rhythmic change of pulmonary functions in nocturnal asthma.     

Ultrasonic nebulization of hypertonic solution: a new method for obtaining specimens from nasal mucosa for morphologic and biochemical analysis in allergic rhinitis

Various techniques are used to collect specimens from the nasal mucosa for morphologic and biochemical analysis. The purpose of this study was to devise a method that overcomes some of the disadvantages (e.g., invasive procedure, samples not suitable for cytologic and biochemical analysis, lack of standardization, and poor reproducibility) of these techniques. The new method requires subjects, with neck extended, to inhale an ultrasonic nebulization of a hypertonic (3% NaCl) solution (UNHS) for 5 min. They then blow their nose into a Petri dish, one nostril at a time with the other one blocked. The secretions are dispersed with 0.1% dithiothreitol in phosphate buffer solution for 20 min. Total cell count (TCC) is evaluated, and the cellular suspension is divided into two aliquots: one is centrifuged and the supernatants are collected for eosinophil cationic protein (ECP) measurements; the other is cytocentrifuged and the slides, stained with Diff-Quik, are used for differential cell count. The results obtained with the UNHS and nasal lavage (NL) methods were compared. Eleven nonatopic healthy subjects and 19 allergic rhinitic patients were studied. Total cell count (x10(5)) was significantly higher with UNHS than with NL (13.0+/-12.3 vs 1.9+/-1.6; P<0.01) The differential cell count was similar with the two procedures. ECP levels (microg/l) were higher with UNHS than with NL (39.1+/-38.2 vs 16.7+/-41.2; P<0.01). For evaluation of reproducibility, four healthy and six rhinitic subjects underwent UNHS on two occasions within 5 days, and the results of two samples (sample 1 vs sample 2) were analyzed. Reproducibility was good as to TCC, differential cell count, and ECP.     

The relationship of eosinophil viability enhancing activity in sputum and clinical symptoms in asthma

We studied the correlation of eosinophil viability enhancing activity (EVEA) in sputum from asthmatic children and clinical symptoms. Sputum from asthmatic children and equal volume of saline were mixed with a Vortex mixer and centrifuged at 40000 G. Clear supernatants were obtained and filtered with 0.22 micron membrane filter. Periphral blood eosinophils purified by Percoll density gradient centrifugation and CD16 negative selection/ immunomagnetic beads technique were incubated with sputum extract for 4 days. Eosinophil viability was examined by staining the cells with fluorescen diacetate and propidium iodide and expressed as eosinophil viability enhancing activity (EVEA). Eosinophil cationic protein (ECP) concentrations in sputum were also measured by a radioimmunoassay. Correlation between EVEA in sputum and pulmonary functions, attack score, treatment score, or sputum ECP was determined by Spearman correlation test. Sputum EVEA was correlated 1) inversely with pulmonary functions on sampling, 2) with attack score of 2 weeks period before sampling, 3) with treatment score of 1 month to 1 week before and 1 to 2 weeks after sampling, and 4) with sputum ECP in individual cases. Sputum EVEA may serve as a suitable parameter for monitoring airway inflammation in asthma.