Surfactant protein levels in bronchoalveolar lavage after segmental allergen challenge in patients with asthma

BACKGROUND: Allergic asthma is associated with airway inflammation and dysfunction of pulmonary surfactant. Because surfactant proteins (SP) account for immunomodulatory functions as well as biophysical functions, we hypothesized that the allergic response in asthma might be accompanied by a dysregulation of SPs. METHODS: We measured levels of SP-A, SP-B, SP-C and SP-D by enzyme-linked immunosorbent assay in bronchoalveolar lavage (BAL) fluid of 23 asthma patients and 10 healthy control subjects under well-controlled conditions before and 24 h after segmental allergen provocation. These data were related to surfactant function, Th(2) cytokine levels in BAL fluid and to the degree of eosinophilic inflammation. RESULTS: In patients with asthma, allergen challenge increased BAL levels of SP-B, SP-C and SP-D while SP-A was decreased. For SP-B and SP-D, a moderate increase was also observed after saline challenge. In contrast, no alterations were observed in healthy control subjects. Levels of SP-B and SP-C in asthmatics correlated with the ratio of small to large surfactant aggregates (SA/LA ratio) and correlated negatively with BAL surface activity. Furthermore, increased SP-C but not SP-B levels after allergen challenge correlated with eosinophil numbers, interleukin (IL)-5, and IL-13 in BAL while increased SP-D levels only correlated with eosinophil numbers. CONCLUSIONS: This study demonstrates significant alterations of all SPs in BAL fluid after allergen challenge of which SP-C was most closely related to surfactant dysfunction and the degree of the allergic inflammation.     

Segmental allergen challenge in patients with atopic asthma leads to increased IL-9 expression in bronchoalveolar lavage fluid lymphocytes

BACKGROUND: IL-9 is a T(H)2 cell-derived cytokine that might be involved in the pathophysiology of allergic diseases. Little is known about its expression and release during the allergic response in the human lung. OBJECTIVE: The expression of IL-9 was measured in 10 atopic subjects with mild asthma and 5 nonatopic healthy control subjects at baseline and 24 hours after segmental sham and allergen challenge. METHODS: IL-9 protein was measured in bronchoalveolar lavage (BAL) fluid by means of ELISA and detected within the BAL cells by means of immunocytochemistry. Furthermore, IL9 mRNA expression of BAL cells was detected by means of real-time PCR. RESULTS: Although only low or undetectable amounts of IL9 mRNA and IL-9 protein were present in nonatopic control subjects and atopic asthmatic patients at baseline, there was an increase after segmental allergen challenge in the atopic subjects. Lymphocytes were identified as major cellular sources of IL-9 production by means of immunocytochemistry. Furthermore, IL-9 protein and IL9 mRNA expression correlated with eosinophil numbers in BAL fluid. CONCLUSIONS: These findings demonstrate that IL-9 is specifically upregulated after local allergen challenge in the lungs of atopic asthmatic patients. Lymphocytes are the major cellular source of IL-9. The increased expression and its correlation with eosinophil numbers suggest a potential role for IL-9 in the late phase of the allergic response.     

Heat-labile neutrophil chemotactic activity in subjects with asthma after allergen inhalation: relation to the late asthmatic reaction and effects of asthma medication

Neutrophil chemotactic activity (NCA) in serum is raised in subjects with asthma after inhalation of an allergen. Two kinds of NCA have been demonstrated, heat-stable and heat-labile. In addition, inhibitory activity is generated after inhalation challenge. In the present study we have investigated the relationship of heat-labile NCA to the development of the late asthmatic reaction (LAR) in 13 subjects with asthma after allergen challenge and the effects of asthma medication on the formation of this activity. Heat-labile NCA peaked 120 to 240 minutes after challenge and demonstrated at this time significant (p less than 0.001) quantitative relationships to the ensuing LAR. The inhalant corticosteroid, budesonide, significantly inhibited (p less than 0.001) the generation of heat-labile NCA and the development of LAR both after a single dose and after 4 weeks of pretreatment. Single-dose disodium cromoglycate pretreatment, initially, slightly enhanced (p less than 0.05) heat-labile NCA but, after 120 minutes, slightly inhibited (p less than 0.05) the activity. Disodium cromoglycate also slightly abrogated the development of LAR. Single-dose pretreatment with the beta 2-agonist, terbutaline, inhibited generation of heat-labile NCA (p less than 0.001) but was without effect on LAR. It is concluded that the generation of heat-labile NCA is related to the development of the LAR and may be of importance for the attraction of inflammatory cells to the lung in the development of the inflammatory reaction probably responsible for LAR. However, the pharmacologic control of heat-labile NCA indicates that the process is multifactorial and not solely dependent on the generation of NCAs detected in serum.     

Frequency and intensity of late asthmatic reactions after bronchial allergen challenge in asthma and rhinitis

The occurrence of late asthmatic reactions after bronchial allergen challenge was studied in 50 house dust mite allergic patients subdivided in three groups: one group had asthma without nasal symptoms, another group had rhinitis without pulmonary symptoms and a third group had a combination of both asthma and rhinitis. Late asthmatic reactions were present in 80% of asthmatic patients and in 18.7% of rhinitis patients. The degree of non-specific bronchial reactivity to histamine (provocative dose 15 or PD15 histamine) and the degree of immediate reactivity to allergen (PD15 house dust mite) did not differ significantly between patients with and without late asthmatic reactions. These findings suggest that an important difference between asthma and rhinitis is the lack of late asthmatic reactions in rhinitis patients, whereas the degree of immediate bronchial reactivity to the allergen is similar in asthma and rhinitis.     

Late bronchial response and increase in methacholine hyperresponsiveness after exercise and distilled water challenge in atopic subjects with asthma with dual asthmatic response to allergen inhalation

We investigated the occurrence of late asthmatic response and increased methacholine responsiveness after exercise and ultrasonically nebulized distilled water (UNDW) inhalation in 12 subjects with asthma with dual asthmatic response and increased responsiveness after allergen challenge. On 3 separate days, allergen, exercise, and UNDW challenges were performed 2 hours after methacholine. FEV1 was measured for 8 hours to detect any delayed change in airway caliber. If there were a further significant reduction in FEV1 after the recovery from the immediate bronchoconstriction, methacholine challenge was performed again when FEV1 had returned to baseline. Reproducibility of any observed late response to exercise and UNDW was also investigated by repeating these challenges on 2 subsequent days. After allergen inhalation only nine subjects had an early asthmatic response, whereas all the tested subjects demonstrated a late reaction and increased methacholine responsiveness. Ten subjects had an immediate response to exercise, and this was followed by a late response in only four patients. Nine subjects demonstrated early response to UNDW inhalation, and five subjects also had a late reaction. These late responses were associated with an increase in methacholine responsiveness in a subset of the tested subjects. Late-phase reactions to exercise and UNDW were not reproducible.     

Tryptase in nasal lavage fluid after local allergen challenge: Relationship to histamine levels and TAME-esterase activity

The activation of mast cells is generally considered to be an important trigger mechanism in the immediate allergic response. This study focused on the determination of three markers of mast cell activation after an allergen challenge. Nasal allergen challenges were performed in 25 subjects with seasonal allergic rhinitis using three allergen doses increasing in 10-fold steps in a standardised nasal lavage model for the subsequent recovery of the markers of mast cell activation. The levels of histamine and tryptase in the nasal lavage fluid were determined using radioimmunoassays, while the TAME-esterase activity was determined using a radiochemical technique. The nasal symptoms obtained on challenge were assessed using a scoring technique. The allergen challenge resulted in significant increases in the levels of all three markers, tryptase, histamine and TAME-esterase. In the individual measurements after the challenges there was a highly significant correlation between the TAME-esterase levels and the tryptase levels (r = 0.71; P less than 0.001), while the generation of histamine and tryptase was not significantly correlated. When comparing the cumulative generation of the three markers, significant correlations were found between all three. Allergen challenges in six non-allergic controls using the same technique did not result in any increase in tryptase levels. The findings suggest that the determination of tryptase in nasal lavage fluid may be a valuable indicator of mast cell activation in the upper airways.     

Increases in eotaxin‐positive cells in induced sputum from atopic asthmatic subjects after inhalational allergen challenge

BACKGROUND: Eosinophils are believed to be critical proinflammatory cells in airway mucosal damage in asthma. Eotaxin is a C-C chemokine with selective activity for eosinophils and basophils. Previous studies have shown increased expression of eotaxin in the airways of asthmatics at baseline. We aimed to investigate eotaxin expression during the late-phase reaction to allergen inhalation in atopic asthmatics. METHODS: Sputum induction was performed before and 24 h after inhalational allergen challenge in atopic asthmatics, and eotaxin protein was detected immunocytochemically. RESULTS: Thirteen patients with a mean decrease in forced expiratory volume in 1 s of 28% (+/-1.5) during the early asthmatic reaction, and 39% (+/-4.7) during the late asthmatic reaction produced sufficient sputum for study. The percentage of eosinophils in sputum was increased 24 h after allergen challenge (P<0.004), and eosinophil percentages in sputum after challenge correlated with the magnitude of the late-phase reaction (r=0.56, P=0.05). The percentage of eotaxin-positive cells increased from 12.6% (range 2-43.8) to 24.3% (8.1-47.1, P<0.005). Allergen-induced increases in eotaxin-positive cells correlated with increases in eosinophils (r=0.63, P<0.01). CONCLUSIONS: These findings suggest that eotaxin may contribute to allergen-induced recruitment of eosinophils to the airway in asthmatic subjects.     

Cellular and protein changes in bronchial lavage fluid after late asthmatic reaction in patients with red cedar asthma

To investigate the sequence of cellular and protein changes after a late asthmatic reaction (LAR), bronchial lavage was carried out in 44 patients with red cedar asthma at different time intervals after bronchial challenge with plicatic acid. The results were compared to five patients with red cedar asthma who became asymptomatic after removal from exposure to red cedar for more than 2 months and 31 healthy subjects without asthma. The LAR was found to be associated with an increase in eosinophils in the lavage fluid, an increase in sloughing of bronchial epithelial cells, and an increase in degenerated cells consisting mainly of degenerated epithelial cells and alveolar macrophages. There was an increase in vascular permeability as reflected by an increase in albumin in the lavage fluid. Although there was a slight but significant increase in neutrophils 48 hours after bronchial challenge, neutrophil infiltration was not a prominent feature earlier. The potential role of loss of epithelial cells to account for an increase in nonspecific bronchial hyperresponsiveness after an LAR was discussed.     

Blood markers of early and late airway responses to allergen in asthmatic subjects. Relationship with functional findings

We evaluated the relationship between blood markers of mast-cell (plasma histamine and serum level of heat-stable neutrophil chemotactic activity [NCA]) and eosinophil (serum eosinophil cationic protein [ECP]) activation during early airway response (EAR) and late airway response (LAR) to allergen inhalation in 24 asthmatic subjects. After EAR, 14 subjects showed significant LAR (FEV₁ fall: 25%), while 10 subjects showed equivocal LAR (FEV₁ fall: 15–20%). A significant increase from baseline value was observed in plasma histamine and in serum NCA during both EAR and LAR, while serum ECP significantly increased only during LAR. The sensitivity of different markers to detect significant FEV₁ fall during EAR and LAR was low, except for NCA. Changes in blood mediators were similar in both groups with significant and equivocal LAR. There was a significant relationship between the increase in NCA during EAR and the severity of LAR. Stepwise regression between changes in different blood markers showed a significant relationship between histamine increase during EAR and ECP increase during LAR. Thus, serum NCA is a more sensitive marker of EAR and LAR than plasma histamine and serum ECP, and its increase during EAR seems predictive of the severity of the subsequent LAR.     

Repeated exposure to low-dose diesel exhaust after allergen challenge exaggerates asthmatic responses in mice

BACKGROUND: In conjunction with allergens, diesel exhaust particles act as an adjuvant to enhance IgE responses, inducing expression of cytokines/chemokines and adhesion molecules, and increasing airway hyper-responsiveness (AHR). As most studies were designed to expose animals to diesel exhaust throughout the periods of both sensitization and allergen challenge, it remains unclear whether diesel exhaust (DE) exposure exaggerates airway responses in asthmatic animals. OBJECTIVE: To study effects of exposure to low-dose DE on AHR and allergic airway inflammation in asthmatic mice. METHODS: BALB/c mice were sensitized by intraperitoneal injection of ovalbumin and challenged by intranasal administration with ovalbumin. They were exposed to low-dose DE for 7 h/day, 5 days/week, for up to 12 weeks. AHR to methacholine was evaluated by whole-body plethysmography as well as bronchoalveolar lavage cell analysis and cytokine gene expression in lungs. RESULTS: Repeated exposure of asthmatic mice to low-dose DE resulted in increased AHR and gene expression of several pro-asthmatic cytokines/chemokines, but these effects rapidly subsided with continued exposure to DE. CONCLUSION: Repeated exposure to low-dose DE after ovalbumin challenge exaggerates allergic responses in mice, but effects are not prolonged with continuous DE exposure.     

Basic fibroblast growth factor in asthma: measurement in bronchoalveolar lavage fluid basally and following allergen challenge

Airway remodeling in asthma refers to a collection of chronic structural changes including subepithelial fibrosis, airway smooth muscle hypertrophy/hyperplasia, and possibly angiogenesis. The mechanisms leading to remodeling are not well defined. One molecule of possible relevance is basic fibroblast growth factor (bFGF), which is a potent mitogen for fibro-blasts, airway smooth muscle cells, and endothelial cells. To test the hypothesis that bFGF expression is increased in asthma, we measured levels of the growth factor in bronchoalveolar lavage (BAL) fluid. Basally, BAL fluid bFGF concentrations were significantly higher in subjects with atopic asthma than in control subjects without asthma (median 0.22 vs 0.06 pg/mL, P = .003). The effect of acute allergen exposure was examined with a segmental bronchoprovocation model in a separate group of subjects with atopic asthma. Ten minutes after segmental bronchoprovocation there was a 5-fold increase in bFGF levels in BAL fluid recovered from allergen-challenged sites compared with control saline-challenged sites (1.52 vs 0.30 pg/mL, P < .002). We conclude that basal levels of BAL fluid bFGF are increased in atopic asthma and that a further increase occurs in response to acute allergen exposure. These findings lend support to the hypothesis that bFGF is implicated in airway remodeling in asthma.     

Effect of ciclesonide on allergen challenge in subjects with bronchial asthma

BACKGROUND: The aim of this clinical trial was to investigate whether repeated inhalation of the new inhaled steroid ciclesonide reduces the early-phase (EAR) and late-phase (LAR) reactions after allergen challenge in patients with mild allergic asthma. Also, this study provides further data on safety and tolerance of ciclesonide. METHODS: The study was designed as a double-blind placebo-controlled randomized crossover trial. Following a baseline period, patients were randomized to either of two treatment sequences (ciclesonide/placebo, placebo/ciclesonide) each of which lasted for one week and were separated by 3-5 weeks from the alternate treatment sequence. Patients received 800 micro g ciclesonide twice daily by means of a Cyclohaler. At the end of each treatment patients were subjected to an allergen challenge. RESULTS: Thirteen asthmatic patients (mean FEV1 of 91% predicted) who experienced an EAR and LAR after allergen challenge participated in the study. The time-average FEV1 decreases 0-2 h (2-12 h) after allergen challenge as measure of the EAR (LAR) were significantly reduced (P < 0.05, one-sided) from 0.426 L to 0.233 L (EAR) and from 0.443 L to 0.213 L (LAR), respectively. Thus, the study results suggest that ciclesonide significantly lowered the extent of EAR and LAR compared to placebo. Ciclesonide was well tolerated and no drug-related adverse events were reported. Cortisol excretion in 24-h urine showed no significant difference between ciclesonide and placebo. CONCLUSIONS: The study supports the efficacy and safety of ciclesonide.     

Factors influencing the occurrence of a late reaction to allergen challenge in atopic asthmatics

Thirty extrinsic asthmatics were challenged by inhalation with Dermatophagoides pteronyssinus extract. In twenty-four an immediate reaction was observed and in sixteen this was followed by a late reaction. Those with late reactions tended to have more severe asthma but did not report greater sensitivity to housedust mite. The occurrence of a late reaction was not related to the degree of airways obstruction before challenge or to the intensity of the immediate reaction. Patients in whom the early reaction was induced by a low dose of inhaled antigen were those most likely to develop a late response. Results of histamine challenge testing suggested that this greater sensitivity of the airway might in part be due to greater non-specific bronchial reactivity.     

Effects of prednisone on the cellular responses and release of cytokines and mediators after segmental allergen challenge of asthmatic subjects.

BACKGROUND: Systemic glucocorticoids are a major therapy for the management of allergic inflammation and asthma; however, information about their effects in vivo are limited. OBJECTIVE: This study was performed to examine the effects of prednisone on inflammatory mediators, cytokines, and cellular responses in the model of segmental allergen challenge (SAC) of allergic asthmatic subjects. METHODS: The effects of a 3-day pretreatment with oral prednisone (30 mg twice daily) on the physiologic and inflammatory responses to SAC were studied in 10 allergic asthmatic subjects in a double-blind, placebo-controlled, crossover protocol. RESULTS: Prednisone improved baseline FEV(1) by 10% and modestly inhibited the SAC-induced fall in FEV(1) at 30 minutes and at 6 to 8 hours. Five minutes after challenge, levels of histamine, PGD(2), 9alpha,11beta-PGF(2), and thromboxane B(2) increased in bronchoalveolar lavage fluid (median increase, 5- to 14-fold); prednisone did not inhibit these responses. Prednisone inhibited (median decrease, 66%-97%) the total influx of inflammatory cells, specifically eosinophils, basophils, and some subsets of T lymphocytes (CD4, CD45RA, and CD45RO cells) assessed 19 hours after SAC, but it did not inhibit the influx of neutrophils. Increases in soluble E-selectin, kinins, and albumin were also inhibited by the glucocorticoid (median decrease, 36%-74%). Prednisone treatment inhibited the appearance of mRNA, protein, or both for T(H)2 cytokines (IL-4 and IL-5), as well as for IL-2 and transforming growth factor alpha, but did not inhibit increases of immunoreactive GM-CSF in bronchoalveolar lavage fluid. CONCLUSION: These studies indicate that prednisone suppresses multiple components of allergic airway inflammation, including cell recruitment, adhesion molecule expression or release, airway permeability, and production of cytokines potentially involved in airway immunity or remodeling.     

Bronchial allergen challenge in subjects with low levels of allergic sensitization to indoor allergens

BACKGROUND: Low skin reactivity to common inhalant allergens is frequently found in asymptomatic individuals as well as in patients with respiratory complaints. However, most studies on bronchial allergen challenge concern patients with high levels of allergic sensitization. The present study was directed to bronchial reactions after allergen challenge in subjects with low skin reactivity to Dermatophagoides pteronyssinus or cat dander. METHODS: Titrated intracutaneous skin tests, skin prick tests, specific IgE assays, histamine release on washed leukocytes, and bronchial histamine and allergen-challenge tests were performed in 20 subjects with an intracutaneous skin test threshold for cat dander (Felis domesticus) or D. pteronyssinus above 0.1 BU/ml (mean wheal diameter in skin prick test with 10000 BU/ml: 4.4mm). Ten of the 20 patients had specific IgE below the detection limit in at least one of the three IgE assays which were done. Fifteen patients had a specific IgE level below 2 kU/I in all three tests. As a positive control group, the same parameters were studied in seven moderately sensitized patients with an intracutaneous skin test threshold below 0.1 BU/ml (mean wheal diameter with 10000 BU/ml: 7.2mm). RESULTS: The 20 subjects with low levels of allergic sensitization had an early decrease in FEV1 of 8.6% (P<0.01) and a mean late decrease of 6.3% (P<0.05). There was a trend for decrease in PC20 histamine 24h after allergen challenge (-0.4 doubling doses, P=0.09). CONCLUSIONS: In this group of subjects with low levels of allergic sensitization, a statistically significant early and late decrease in FEV1 was found. However, the decrease in lung function was small and unnoticed by most patients. The increase in nonspecific bronchial hyperresponsiveness after bronchial allergen challenge did not reach statistical significance in the study group. The results indicate that allergen exposure in patients with low levels of allergic sensitization may lead to airways changes in the absence of acute symptoms.