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Asthma patients show evidence of a procoagulant state in their airways, accompanied by an impaired function of the anticoagulant protein C system. We aimed to study the effect of recombinant human activated protein C (rhAPC) in allergic asthma patients.We conducted a randomised, double-blind, placebo-controlled, proof-of-concept study in house dust mite (HDM) allergic asthma patients. Patients were randomised to receive intravenous rhAPC (24 µg·kg−1·h−1; n=12) or placebo (n=12) for 11 h. 4 h after the start of infusion, a first bronchoscopy was performed to challenge one lung segment with saline (control) and a contralateral segment with a combination of HDM extract and lipopolysaccharide (HDM+LPS), thereby mimicking environmental house dust exposure. A second bronchoscopy was conducted 8 h after intrabronchial challenge to obtain bronchoalveolar lavage fluid (BALF).rhAPC did not influence HDM+LPS induced procoagulant changes in the lung. In contrast, rhAPC reduced BALF leukocyte counts by 43% relative to placebo, caused by an inhibitory effect on neutrophil influx (64% reduction), while leaving eosinophil influx unaltered. rhAPC also reduced neutrophil degranulation products in the airways.Intravenous rhAPC attenuates HDM+LPS-induced neutrophil migration and protein release in allergic asthma patients by an effect that does not rely on coagulation inhibition.

Background: Mast cells are implicated in allergic and innate immune responses in asthma, although their role in models using an allergen relevant for human disease is incompletely understood. House dust mite (HDM) allergy is common in asthma patients. Our aim was to investigate the role of mast cells in HDM-induced allergic lung inflammation. Methods: Wild-type (Wt) and mast cell-deficient Kitw-sh mice on a C57BL/6 background were repetitively exposed to HDM via the airways. Results: HDM challenge resulted in a rise in tryptase activity in bronchoalveolar lavage fluid (BALF) of Wt mice, indicative of mast cell activation. Kitw-sh mice showed a strongly attenuated HDM- induced recruitment of eosinophils in BALF and lung tissue, accompanied by reduced pulmonary levels of the eosinophil chemoattractant eotaxin. Remarkably, Kitw-sh mice demonstrated an unaltered capacity to develop lung pathology and increased mucus production in response to HDM. The increased plasma IgE in response to HDM in Wt mice was absent in Kitw-sh mice. Conclusion: These data contrast with previous reports on the role of mast cells in models using ovalbumin as allergen in that C57BL/6 Kitw-sh mice display a selective impairment of eosinophil recruitment without differences in other features of allergic inflammation.

The complex biology of asthma compels the use of more relevant human allergens, such as house dust mite (HDM), to improve the translation of animal models into human asthma. LPS exposure is associated with aggravations of asthma, but the mechanisms remain unclear. Here, we studied the effects of increasing LPS doses on HDM-evoked allergic lung inflammation. To this end, mice were intranasally sensitized and challenged with HDM with or without increasing doses of LPS (0.001-10 μg). LPS dose-dependently inhibited HDM-induced eosinophil recruitment into the lungs and mucus production in the airways. LPS attenuated the production of Th2 cytokines (IL-4, IL-5, IL-10, and IL-13) in HDM-challenged lungs, while enhancing the HDM-induced release of IL-17, IL-33, IFN-γ, and TNF-α. The shift toward a Th1 inflammatory response was further illustrated by predominant neutrophilic lung inflammation after LPS administration at higher doses. LPS did not influence HDM-induced plasma IgE concentrations. Although LPS did not significantly affect the activation of coagulation or complement in HDM-challenged lungs, it reduced HDM-initiated endothelial cell activation. This study is the first to provide insights into the effects of LPS in an allergic lung inflammation model making use of a clinically relevant allergen without a systemic adjuvant, revealing that LPS dose-dependently inhibits HDM-induced pulmonary Th2 responses.

Intravenous administration of activated protein C (APC) inhibits coagulation and inflammation in the lungs of humans and animals. Investigations in rodents demonstrated that direct intrapulmonary delivery of APC also exerts anticoagulant and anti-inflammatory effects. The effect of intrabronchial administration of recombinant human (rh)APC on lipopolysaccharide (LPS)-induced haemostatic and inflammatory alterations in the bronchoalveolar space of humans was studied. Eight subjects received rhAPC via intrabronchial instillation by bronchoscope, while in a contralateral subsegment subjects received saline; all subjects were challenged bilaterally with LPS in the same lung subsegments. Four additional subjects received rhAPC (75 μg), with saline as a control in the contralateral subsegment, while they were bilaterally “challenged” with saline. After 6 h a bronchoalveolar lavage was performed and coagulation and inflammatory parameters were measured. rhAPC enhanced LPS-induced coagulation activation in the bronchoalveolar space, when compared with the control side. In addition, rhAPC amplified LPS-induced pro-inflammatory responses, as indicated by higher concentrations of cytokines and chemokines. rhAPC alone did not have procoagulant or pro-inflammatory effects. Locally administered rhAPC has unexpected procoagulant and pro-inflammatory effects in LPS-challenged lung subsegments. These data argue against a role for intrapulmonary delivery of rhAPC as a treatment strategy for lung inflammatory disorders in humans.

Lipopolysaccharide (LPS) is ubiquitous in the environment. Inhalation of LPS has been implicated in the pathogenesis and/or severity of several lung diseases, including pneumonia, chronic obstructive pulmonary disease and asthma. Alveolar macrophages are the main resident leukocytes exposed to inhaled antigens. To obtain insight into which innate immune pathways become activated within human alveolar macrophages upon exposure to LPS in vivo, we conducted a study in eight healthy humans, in which we instilled sterile saline into a lung segment by bronchoscope, followed by instillation of LPS into the contralateral lung. Six hours later, a bilateral bronchoalveolar lavage was performed and whole-genome transcriptional profiling was done on purified alveolar macrophages, comparing cells exposed to saline or LPS from the same individuals. LPS induced differential expression of 2,932 genes in alveolar macrophages; 1,520 genes were upregulated, whereas 1,440 genes were downregulated. A total of 26 biological functions were overrepresented in LPS-exposed macrophages; 44 canonical pathways affected by LPS were identified, among which the genes associated with the role of pattern recognition receptors in recognition of bacteria and viruses represented the top pathway. Other pathways included cellular immune response, signaling by tumor necrosis factor (receptor) family members, cytokine signaling and glucocorticoid receptor signaling. These results reveal for the first time a large number of functional pathways influenced by the biologically relevant challenge provided by LPS administered into the airways. These data can assist in identifying novel targets for therapeutic intervention in pulmonary diseases associated with LPS exposure, including pneumonia, asthma and chronic obstructive pulmonary disease.

The airways are continuously exposed to respiratory pathogens, which may result in bacterial pneumonia, one of the most common infectious diseases and the leading cause of sepsis. Considering that recurrent exposure to microbial products can lead to tolerance of immune cells, and that this might contribute to the susceptibility to nosocomial infection, we investigated the effect of in vivo lipopolysaccharide (LPS) instillation on the responsiveness of alveolar macrophages. In eight healthy humans, sterile saline was instilled into a lung segment by bronchoscope, followed by instillation of LPS into the contralateral lung; 6 hours later, a bilateral bronchoalveolar lavage was performed, and purified alveolar macrophages were ex vivo stimulated with LPS or lipoteichoic acid (LTA), triggering Toll-like receptor (TLR)-4 and -2, respectively. In vivo LPS-exposed alveolar macrophages were primed, as reflected by increased ex vivo LPS- and LTA-induced IL-1 beta and IL-6 gene expression and production compared with in vivo saline-exposed alveolar macrophages. LPS instillation did not influence the surface expression of TLR4 or TLR2. Furthermore, LPS instillation did not impact on the expression of a number of extracellular and intracellular regulators of TLR signaling. However, p38 mitogen-activated protein kinase remained phosphorylated in alveolar macrophages upon LPS instillation. The current data demonstrate that LPS instillation in the human lung primes alveolar macrophages for further stimulation with either LPS or LTA, possibly by sustained p38 mitogen-activated protein kinase activation.

OBJECTIVE: Alveolar fibrin deposition is a hallmark of pneumonia. It has been proposed that natural inhibitors of coagulation, including activated protein C, antithrombin, and tissue factor pathway inhibitor, exert lung-protective effects via anticoagulant and possibly anti-inflammatory pathways. We investigated the role of these natural anticoagulants in Streptococcus pneumoniae pneumonia. DESIGN: A controlled in vivo laboratory study. SETTING: Research laboratory of a university hospital. SUBJECTS: Total of 98 male Sprague-Dawley rats. INTERVENTIONS: Rats were challenged intratracheally with S. pneumoniae (serotype 3, 10(6) colony forming units), inducing pneumonia. Rats were randomized to intravenous treatment with normal saline, activated protein C, antithrombin, tissue factor pathway inhibitor, heparin, or tissue-type plasminogen activator. MEASUREMENTS AND MAIN RESULTS: Rats infected with S. pneumoniae had increased thrombin-antithrombin complexes in bronchoalveolar lavage fluid, with decreased levels of antithrombin activity and fibrin degradation products. Administration of activated protein C, antithrombin, and tissue factor pathway inhibitor significantly limited these procoagulant changes. Furthermore, antithrombin treatment resulted in less bacterial outgrowth of S. pneumoniae and less histopathologic damage in lungs. CONCLUSIONS: Anticoagulant treatment attenuates pulmonary coagulopathy during S. pneumoniae pneumonia. Antithrombin seems to exert significant lung-protective effects in pneumococcal pneumonia in rats.

Background: Family members of laboratory animal workers are at risk of developing allergy to laboratory animals. Little is known about the spreading of laboratory animal allergens outside the animal facilities. Objective: To assess the presence of laboratory animal allergens in dust collected from mattresses of laboratory animal workers and unexposed controls. Methods: Mouse and rat urinary proteins were measured in samples of mattress dust collected by laboratory animal workers and unexposed controls. In addition, rat and mouse allergens were determined in extracts of hair-covering caps, used during laboratory animal work, to estimate spreading of allergen through dust captured on hair. Allergen concentrations on hair caps were compared with exposure measured by personal airborne dust sampling. Results: Levels of rat urinary allergens (RUA) and mouse urinary allergens (MUA) and mouse urinary protein (MUP) 8, a specific pheromone-binding mouse allergen, were significantly higher in mattress samples of laboratory animal workers than in those of controls. Hair-covering caps used in animal facilities harboured large amounts of RUA and MUA, which correlated significantly with exposure measured by the personal sampling technique in the animal facility. Conclusions: Occupational laboratory animal allergens are detectable in mattress dust of laboratory animal workers. Transfer of allergens via uncovered hair of animal workers is likely contributing to this phenomenon. This study stresses the importance of using hair caps to prevent spreading of occupational allergens.

Background: Recombinant allergens are preferred over natural allergen extracts in measuring antibodies. We tested the use of recombinant variants of the major mouse allergen Mus m 1 in detection of mouse-specific antibodies in sera of laboratory animal workers and children. Methods: Six recombinant major urinary proteins (MUPs) were produced and antibody-binding capacity was compared to natural Mus m 1 and to mouse urine extract. In a specific subset, cross-reactivity of MUP with Mus m 1 and between the different recombinant MUPs was determined. Results: For IgE antibodies, MUP8 showed high cross-reactivity with Mus m 1. MUP8-specific IgE was found in 55% of the mouse urine IgE-positive sera. Specific IgG and IgG4 antibodies against natural Mus m 1 correlated strongly with antibodies against recombinant MUP8 and were cross-reactive. IgG4 levels against MUP8 and mouse urine extract correlated, but detection of mouse urine-specific IgG4 in the absence of MUP-specific IgG4 was not uncommon. Cross-reactivity of IgG antibodies between MUP8 and Mus m 1 as well as between the different MUPs was high and inhibition varied between 54 and 99%. Conclusion: The mouse allergen Mus m 1 can be replaced in antibody testing by recombinant MUP8. Other MUPs, except MUP4, are interchangeable with MUP8. However, mouse urine extract showed better detection of both mouse-specific IgE and IgG4 levels. Other components in the mouse urine, like mouse albumin and other yet unidentified components, also induce IgE and IgG(4) antibodies.

Objective The tissue factor (TF)-factor VIIa (FVIIa) complex not only is essential for activation of blood coagulation but also affect the inflammatory response during sepsis. The objective of this study was to determine the role of TF-FVIIa in pneumonia caused by Streptococcus pneumoniae, the most important causative organism in community-acquired pneumonia and a major cause of sepsis. Design A controlled, in vivo laboratory study. Setting Research laboratory of a health sciences university. Patients and subjects Patients with unilateral community-acquired pneumonia and female BALB/c mice. Interventions Bilateral bronchoalveolar lavage was performed in patients with community-acquired pneumonia. In mice, pneumonia was induced by intranasal inoculation with S. pneumoniae with or without concurrent inhibition of TF-FVIIa by subcutaneous injections of recombinant nematode anticoagulant protein (rNAPc2). Measurements and main results Patients with unilateral community-acquired pneumonia demonstrated elevated concentrations of FVIIa, soluble TF, and thrombin-antithrombin complexes in bronchoalveolar lavage fluid obtained from the infected site compared with the uninfected site. Mice with S. pneumoniae pneumonia displayed increased TF expression and fibrin deposits in lungs together with elevated thrombin-antithrombin complex levels in bronchoalveolar lavage fluid; inhibition of TF-FVIIa by rNAPc2 attenuated the procoagulant response in the lung but did not affect host defense, as reflected by an unaltered outgrowth of pneumococci and an unchanged survival. Conclusions These data suggest that TF-FVIIa activity contributes to activation of coagulation in the lung during pneumococcal pneumonia but does not play an important role in the antibacterial host defense in this murine model.

Background: Allergy and helminth infections share key immunological features in terms of Th2 responses. Although in industrialized countries clustering of allergic disorders within families has been frequently reported, such information is lacking from areas where helminth infections are endemic. Methods: A total of 466 subjects from 29 families and 112 households participated in this study. Filarial infection, skin test reactivity and IgE to mite as well as total IgE were measured in all samples. Clustering of the allergy-related outcomes due to genetic and household factors was tested. Results: Genetic factors contributed significantly to the clustering of total IgE and allergen-specific IgE, whereas only household factors contributed to the clustering of SPT positivity. Conclusion: Similar to several studies conducted in western populations, total IgE and allergen-specific IgE are influenced by genetic factors in a population resident in a helminth endemic area. However, clustering of SPT positivity due to genetic factors was not significant in the current study raising the question of whether the presence of helminth infections may override genes that are associated with the expression of tissue reactivity to allergens in the west.

Background: It remains to be established which factors contribute to the occurrence of asthma in allergic individuals. We hypothesized that differences in the late allergic inflammatory reaction to allergen between asthmatic and non-asthmatic house dust mite-allergic individuals might contribute to the difference in the clinical presentation of allergy. Aim: To compare allergen-induced changes in parameters for cellular inflammation during the phase of the late allergic reaction in the skin and nose, in house dust mite-allergic individuals with or without asthma. Material and Methods: Nasal and dermal allergen challenges with house dust mite (Dermatophagoides pteronyssinus) extract were performed in 52 house dust mite-allergic individuals, of whom 26 had mild to moderate persistent asthma and 26 had perennial rhinitis without current or past asthmatic symptoms. Serial nasal lavage samples were analyzed for the presence of inflammatory cells (eosinophils and neutrophils) and soluble markers associated with cellular inflammation [interleukin-5 (IL-5), interleukin-8 (IL-8), eosinophil cationic protein (ECP) and myeloperoxidase (MPO)]. Macroscopic late phase skin reactions were studied after intracutaneous skin tests with house dust mite extract. Results: Fixed dose nasal allergen provocation elicited a similar degree of immediate allergic reaction as judged by plasma protein exudation and histamine concentrations in asthma and non-asthmatic rhinitis. Subsequently, no differences between groups were found during the phase of the late allergic reaction (4–24 h) in inflammatory cell influx, plasma protein leakage, ECP or MPO. Likewise, there were no differences in levels of chemotactic cytokines IL-5 and IL-8. In agreement with the results of nasal challenge, the late skin reaction after dermal challenge with a fixed allergen dose and after an allergen dose 10,000 times above the skin threshold for an early skin reaction did not differ between the groups. Conclusion: House dust mite-allergic patients with or without asthma have very similar late allergic inflammatory reactions in the skin and in the nose after allergen challenge. Hence, it is unlikely that the occurrence of pulmonary symptoms in asthma is explained by a general tendency of asthmatics to have an enhanced late allergic cellular inflammatory response. Nasal and dermal allergen provocations are adequate models to study allergen-induced inflammation but probably lack the pivotal link which is essential for the development of asthma.

Urokinase-type plasminogen activator (uPA) and its receptor uPAR are compo- nents of the fibrinolytic system and are important for an adequate immune re- sponse to respiratory tract infection, in part through their role in the migration of inflammatory cells. PA inhibitor-1 (PAI-1) is the predominant inhibitor of soluble and receptor-bound uPA. To determine the role of PAI-1 in host defense against pneumococcal pneumonia, the following studies were performed: (1) Patients with unilateral community-acquired pneumo- nia demonstrated elevated PAI-1 concen- trations together with decreased PAactiv- ity in bronchoalveolar lavage fluid (BALF) obtained from the infected, but not from the contralateral, site. (2) Mice with Strep- tococcus pneumoniae pneumonia dis- played elevated PAI-1 protein and mRNA levels in their lungs. (3) PAI-1 gene- deficient mice, however, had an unaltered immune response to pneumococcal pneu- monia, as measured by cell recruitment into lungs, bacterial outgrowth, and sur- vival. Furthermore, plasminogen-gene- deficient mice also had an unremarkable defense against pneumococcal pneumo- nia. These data indicate that pneumonia is associated with inhibition of thefibrino- lytic system at the site of the infection secondary to increased production of PAI-1; an intact fibrinolytic response is not required for an adequate host re- sponse to respiratory tract infection, how- ever, suggesting that the previously de- scribed role of uPA and uPAR are restricted to their function in cell migra- tion. (Blood. 2003;102:934-939)

Background: Tuberculosis (TB) is an important cause of morbidity and mortality worldwide. Toll-like-receptors (TLRs) are important for the recognition of the causative agent Mycobacterium tuberculosis. Negative regulation of TLRs is necessary to control deleterious inflammatory damage, but could provide a means of immune evasion by M. tuberculosis as well. Methods: To obtain insight in the extent of expression of inhibitory regulators of immunity in patients with active TB, peripheral-blood-mononuclear-cells (PBMCs) and plasma were obtained from 54 TB patients and 29 healthy blood donors from Chittagong, Bangladesh. Bilateral alveolar macrophages were obtained from an infected versus a contralateral normal lung segment of 9 patients. Statistical analyses were performed using Mann-Whitney U and Wilcoxon matched pairs testing. Correlations were calculated using the Spearman rho test. Results: PBMCs harvested from TB patients demonstrated increased mRNA expression of IL-1-receptor-associated-kinase-M, suppressor-of-cytokine-signalling-3 and Toll-interacting-protein. Flow cytometry revealed enhanced expression of IL-1-receptor-like-1 (ST2) on lymphocytes. Plasma soluble ST2 was elevated in patients with TB and correlated with established TB biomarkers, most strongly with soluble interleukin-2 receptor subunit a and interleukin-8. Alveolar macrophage mRNA expression of negative TLR regulators did not differ between the infected and contralateral lung side. Conclusion: These results show enhanced expression of distinct negative regulators of innate immunity in PBMCs of patients with TB and identify plasma soluble ST2 as a potential novel biomarker for TB disease activity

It has been reported that serum immunoglobulin E (IgE) from certain atopic patients can sensitize basophils to release histamine in response to IgE-dependent histamine-releasing factors (HRFs). It has also been shown that patients suffering from severe forms of atopy may contain IgE autoantibodies. It was investigated whether HRF-responsive sera contained IgE autoantibodies and if there was an association between IgE autoreactivity and IgE-dependent responsiveness to HRF. The presence of HRF-responsive IgE (IgE+) in serum of patients with respiratory atopy was determined by stimulating stripped human basophils sensitized by serum with peripheral blood mononuclear cell (PBMC)-derived HRF, and measuring the release of histamine. In parallel, these sera were screened for the presence of IgE autoantibodies to nitrocellulose-blotted human cellular extracts. The capacity of IgE autoantigen-containing preparations to induce histamine release was tested in the stripped basophil assay. Eleven out of 52 sera contained IgE autoantibodies to blotted cellular extracts of human PBMCs or of the human epithelial cell line A431. No significant association was found between IgE autoreactivity and IgE-dependent responsiveness to HRF: 7/26 IgE+ sera contained IgE to human cellular extracts, and 4/26 of the sera without IgE+ did also. IgE autoantigen-containing extracts did not induce histamine release of appropriately sensitized basophils. By size-exclusion chromatography it was shown that a 32 000 MW autoantigen eluted in the >55 000 MW fraction, which indicates that this protein forms polymers or complexes with other macromolecules. This might explain the discrepancy between binding and histamine-releasing activity. A 20 000 MW IgE-defined autoantigen cross-reacted with a shrimp allergen. Our results indicate that IgE-reactivity to immunoblotted human protein and IgE-dependent HRF activity are distinct entities that may co-occur in atopic patients.

Summary Objectives Human tuberculosis (TB) remains an important cause of death globally. Bangladesh is one of the most affected countries. We aimed to investigate the impact of pulmonary TB on pro- and anticoagulant mechanisms. Methods This prospective study was conducted in Chittagong, Bangladesh. We performed an in-depth analysis of coagulation activation and inhibition in plasma obtained from 64 patients with primary lung TB and 11 patients with recurrent lung TB and compared these with 37 healthy controls. Additionally, in nine patients coagulation activation was studied in bronchoalveolar lavage fluid (BALF) harvested from the site of infection and compared with BALF from a contralateral unaffected lung subsegment. Results Relative to uninfected controls, primary and recurrent TB were associated with a systemic net procoagulant state, as indicated by enhanced activation of coagulation (elevated plasma levels of thrombin-antithrombin complexes, D-dimer and fibrinogen) together with impaired anticoagulant mechanisms (reduced plasma levels of antithrombin, protein C activity, free protein S, and protein C inhibitor). Activation of coagulation did not correlate with plasma concentrations of established TB biomarkers. Coagulation activation could not be detected at the primary site of infection in a subset of TB patients. Conclusions Pulmonary TB is associated with a systemic hypercoagulable state.

Background: IL-3, IL-5, and GM-CSF are not able to induce histamine release in purified basophils of nonallergic donors. However, we have recently found that preincubation with 2 μmol/L thapsigargin, which induces a rise in intracellular free calcium ions, renders human basophils extremely sensitive for IL-3, IL-5, or GM-CSF, leading to enhanced histamine release. Histamine release was also induced in the reverse order (first cytokine and then thapsigargin). Objective: Because these cytokines are supposed to be increased in allergic inflammation, we examined whether basophils of patients with allergic asthma showed an enhanced response to thapsigargin. Methods: We measured the histamine release induced by thapsigargin in a group of allergic asthmatic subjects (n = 24) and compared this response with those of 3 control groups. The control groups consisted of healthy control subjects (group 1, n=21); patients with a nonallergic, nonasthmatic lung disease (group 2, n=22); and patients with nonallergic asthma (group 3, n=9). Results: There was no difference in spontaneous histamine release. Also, no significant difference in histamine release was found when anti-IgE or formyl-methionyl-leucyl-phenylalanine was used as a stimulus. Histamine release induced by IL-3 alone or a combination of IL-3 and thapsigargin also did not differ. In contrast, basophils from the group with allergic asthma showed a significantly higher percentage of histamine release induced by thapsigargin (38.2% ± 13.2%) than did basophils from the 3 control groups (healthy control subjects, 22.5% ± 6.9%; subjects with lung disease, 24.9% ± 8.9%; subjects with nonallergic asthma 15.0% ± 3.0%; all mean ± SD). Conclusion: These data indicate that basophils in peripheral blood of subjects with allergic asthma have a primed phenotype and that thapsigargin-induced histamine release is a practical tool to study this phenomenon. (J Allergy Clin Immunol 1999;104:1000-7.)

Background: The use of allergen extracts will hamper studies into quantitative aspects of allergic responses because the precise amount of relevant allergen for each patient is unknown. Objective : We applied isolated IgE-binding components (major allergens) in the technique of bronchial allergen challenge to determine the role of patient characteristics in the early asthmatic response (EAR). Methods: In 30 patients with mild-to-moderate asthma, the EAR was investigated after inhalation of an isolated major allergen of Dermatophagoides pteronyssinus (i.e., Der p 1 [ n = 16] or Der p 2 [ n = 14]). The degree of early-phase bronchial responsiveness to allergen (the cumulated dose of allergen causing a 20% fall in FEV 1 [PD 20 allergen]) was related to the degree of nonspecific bronchial responsiveness (the concentration of histamine causing a 20% fall in FEV 1 [PC 20 histamine]) and the level of specific IgE or allergen thresholds as found in skin tests and basophil histamine release assays. Results: Twenty-seven patients with an immediate response during allergen and histamine challenges (ΔFEV 1 , ≥20%) were analyzed. In these patients, a strong correlation was found between PD 20 allergen and PC 20 histamine ( r = 0.81, p 20 allergen and the level of specific IgE ( r = –0.36, p = 0.07) or allergen thresholds as found in skin tests (skin prick test, r = 0.36 and p = 0.07; intracutaneous test, r = 0.49 and p = 0.01) or basophil histamine release assays ( r = 0.37, p = 0.08). Moreover, no significant contribution of these indices of IgE-mediated hypersensitivity to the prediction of PD 20 allergen by multilinear regression models with PC 20 histamine was found. Conclusion: In asthmatic patients allergic to house dust mites the degree of nonspecific bronchial hyperresponsiveness is the main determinant of early-phase bronchial responsiveness to allergen. In these patients the degree of allergic sensitivity does not contribute to the prediction of the EAR after allergen inhalation. (J Allergy Clin Immunol 1998;102:24-31.)

Background: In our outpatient population, approximately one third of patients sensitized to grass pollen were found to have significant serum levels of anti-peanut IgE in the RAST, without positive peanut skin prick test (SPT) response and without peanut-related allergic symptoms. It was suggested earlier that poor biologic activity of IgE antibodies directed to cross-reactive carbohydrate determinants (CCD) of glycoproteins might explain these discrepancies. Objective: In this study we investigated the biologic activity of IgE directed to CCD. Methods: Sera of 32 patients allergic to grass pollen with significant levels of anti-peanut IgE, a negative response on peanut SPT, and no symptoms of peanut allergy were tested for the presence of anti-CCD IgE. Eleven of these patients with greater than 3.0 IU/ml anti-peanut IgE (patients 1 to 11) were selected together with four control patients allergic to peanut, on the basis of a positive response on peanut SPT and a history of peanut allergy (patients 12 to 15). Inhibition of the peanut RAST was performed by using proteinase K–treated grass pollen extract as a CCD source. Basophil histamine release assays (BHRAs) were performed with peanut extract and the isolated peanut major allergens Ara h 1 and Ara h 2. In addition, intracutaneous tests with peanut extract were performed. Results: In 29 (91%) of 32 patients with discrepant peanut RAST and SPT responses, anti-CCD IgE (≥0.1 IU/ml) was detected. In patients 1 to 11 almost complete inhibition of the peanut RAST with CCD was found (94.3% ± 5.5%; mean ± SD). In contrast, in the patients allergic to peanut only partial inhibition (59%) was found in one subject ( p = 0.002, Mann-Whitney test). In the BHRAs and the intracutaneous tests of patients with discrepant peanut RAST and SPT results, reactivity was found only at high concentrations of peanut allergens. When related to specific IgE levels, reactivity to peanut allergens in the BHRAs of these patients was found to be at least a factor of 1000 less when compared with reactivity to control inhalant allergens. Conclusion: We conclude that cross-reactive IgE directed to carbohydrate determinants of glycoproteins, as found in grass pollen–sensitized patients, has poor biologic activity. It can therefore cause positive RAST results without apparent clinical significance. (J Allergy Clin Immunol 1997;100:327-34.)

BACKGROUND: Mast cells are implicated in allergic and innate immune responses in asthma, although their role in models using an allergen relevant for human disease is incompletely understood. House dust mite (HDM) allergy is common in asthma patients. Our aim was to investigate the role of mast cells in HDM-induced allergic lung inflammation. METHODS: Wild-type (Wt) and mast cell-deficient Kit(w-sh) mice on a C57BL/6 background were repetitively exposed to HDM via the airways. RESULTS: HDM challenge resulted in a rise in tryptase activity in bronchoalveolar lavage fluid (BALF) of Wt mice, indicative of mast cell activation. Kit(w-sh) mice showed a strongly attenuated HDM- induced recruitment of eosinophils in BALF and lung tissue, accompanied by reduced pulmonary levels of the eosinophil chemoattractant eotaxin. Remarkably, Kit(w-sh) mice demonstrated an unaltered capacity to develop lung pathology and increased mucus production in response to HDM. The increased plasma IgE in response to HDM in Wt mice was absent in Kit(w-sh) mice. CONCLUSION: These data contrast with previous reports on the role of mast cells in models using ovalbumin as allergen in that C57BL/6 Kit(w-sh) mice display a selective impairment of eosinophil recruitment without differences in other features of allergic inflammation.

Background In patients with prednisone-dependent asthma the dose of oral corticosteroids should be adjusted to the lowest possible level to reduce long-term adverse effects. However, the optimal strategy for tapering oral corticosteroids is unknown. Objective To investigate whether an internet-based management tool including home monitoring of symptoms, lung function and fraction of exhaled nitric oxide (FENO) facilitates tapering of oral corticosteroids and leads to reduction of corticosteroid consumption without worseningasthmacontrolorasthma-relatedqualityoflife. Methods In a 6-month pragmatic randomised prospective multicentre study, 95 adults with prednisone-dependent asthma from six pulmonary outpatient clinics were allocated to two tapering strategies: according to conventional treatment (n¼43) or guided by a novel internet-based monitoring system (internet strategy) (n¼52). Primary outcomes were cumulative sparing of prednisone, asthma control and asthma-related quality of life. Secondary outcomes were forced expiratory volume in 1 s (FEV1), exacerbations, hospitalisations and patient’s satisfaction with the tapering strategy. Results Median cumulative sparing of prednisone was 205 (25e75th percentile � 221 to 777) mg in the internet strategy group compared with 0 (� 497 to 282) mg in the conventional treatment group (p¼0.02). Changes in prednisone dose (mixed effect regression model) from baseline were � 4.79 mg/day and +1.59 mg/day, respectively (p

Monocyte chemoattractant protein 1 (MCP-1) plays an important role in leukocyte recruitment to sites of infection and inflammation. In addition, MCP-1 may attenuate inflammation by virtue of its capacity to inhibit the production of proinflammatory cytokines. We here investigated the role of MCP-1 in lung inflammation induced by lipopolysaccharide (LPS) or lipoteichoic acid (LTA), constituents of the gram-negative and gram-positive bacterial cell wall, respectively. Healthy humans demonstrated elevated MCP-1 concentrations in their bronchoalveolar lavage fluid (BALF) 6 h after inhalation of LPS. Similarly, intranasal administration of LPS or LTA to mice resulted in a rise in BALF MCP-1 levels. Murine alveolar macrophage-like cells released significant amounts of MCP-1 upon stimulation with LPS or LTA in vitro. Compared to Wt mice, MCP-1(-/-) mice demonstrated lower TNF-alpha levels and a diminished neutrophil influx into their bronchoalveolar space after either LPS or LTA instillation. After intrapulmonary delivery of LPS MCP-1(-/-) mice had decreased interleukin-6 and KC concentrations and less severe lung inflammation upon histopathological examination. Remarkably, MCP-1 deficiency was associated with an early enhancement of interleukin-10 release in BALF after both LPS and LTA instillation. These data suggest that MCP-1 is a proinflammatory mediator during pulmonary inflammation induced by either LPS or LTA. (C) 2011 Elsevier Ltd. All rights reserved

RATIONALE: Recognition of pathogen-associated molecular patterns by Toll-like receptors (TLRs) is considered to be important for an appropriate immune response against pathogens that enter the lower airways. OBJECTIVES: We studied the effects of two different TLR agonists relevant for respiratory infections in the human lung: lipoteichoic acid (LTA; TLR2 agonist, component of gram-positive bacteria) and lipopolysaccharide (LPS; TLR4-agonist, component of gram-negative bacteria). METHODS: Fifteen healthy subjects were given LPS or LTA: by bronchoscope, sterile saline was instilled into a lung segment followed by instillation of LTA or LPS into the contralateral lung. After 6 hours, a bronchoalveolar lavage was performed and inflammatory parameters were determined. Isolated RNA from purified alveolar macrophages was analyzed by multiplex ligation-dependent probe amplification. In addition, spontaneous cytokine release by alveolar macrophages was measured. MEASUREMENTS AND MAIN RESULTS: Marked differences were detected between LTA- and LPS-induced lung inflammation. Whereas both elicited neutrophil recruitment, only LPS instillation was associated with activation of neutrophils (CD11b surface expression, degranulation product levels) and consistent rises of chemo-/cytokine levels. Moreover, LPS but not LTA activated alveolar macrophages, as reflected by enhanced expression of 10 different mRNAs encoding proinflammatory mediators and increased spontaneous cytokine release upon incubation ex vivo. Remarkably, only LTA induced C5a release. CONCLUSIONS: This is the first study to report the in vivo effects of LTA in men and to compare inflammation induced by LTA and LPS in the human lung. Our data suggest that stimulation of TLR2 or TLR4 results in differential pulmonary inflammation, which may be of relevance for understanding pathogenic mechanisms at play during gram-positive and gram-negative respiratory tract infection

OBJECTIVE: Various lung diseases are associated with local activation of coagulation and concurrent inhibition of fibrinolysis. Although salmeterol, a beta2-adrenoceptor agonist with profound bronchodilatory properties, has been studied extensively, the effects of this compound on the pulmonary hemostatic balance are not elucidated. DESIGN: A single-blinded, placebo-controlled study. SETTING: University hospital and laboratory. SUBJECTS: A total of 32 human volunteers. INTERVENTIONS: Subjects inhaled 100 microg of salmeterol or placebo (t = -30 mins) followed by 100 microg of lipopolysaccharide (LPS) or normal saline (t = 0 mins; n = 8 per group). MEASUREMENTS AND MAIN RESULTS: Measurements were performed in bronchoalveolar lavage fluid obtained 6 hrs postchallenge. Inhalation of LPS enhanced pulmonary coagulation as determined by an increase in the concentrations of thrombin-antithrombin complexes, factor VIIa, and soluble tissue factor in bronchoalveolar lavage fluid (all p < .05 vs. saline). LPS concurrently inhibited pulmonary fibrinolysis, as reflected by a decrease in bronchoalveolar lavage fluid plasminogen activator activity together with an increase in plasminogen activator inhibitor type 1 (both p < .05 vs. saline). Moreover, LPS inhalation was associated with a suppression of the anticoagulant protein C pathway, as indicated by an increase in soluble thrombomodulin and decreases in protein C and activated protein C levels in bronchoalveolar lavage fluid (all p < .05 vs. saline). Salmeterol, either with or without LPS inhalation, enhanced fibrinolysis (plasminogen activator activity and tissue-type and urokinase-type plasminogen activator levels) but did not influence LPS-induced changes in coagulation or the protein C pathway. CONCLUSIONS: Salmeterol has profibrinolytic properties in the normal lung and when applied in a model of sterile pulmonary inflammation

SummaryPneumonia is frequently associated with changes in coagulation and fibrinolysis in the bronchoalveolar space. To determine the effect of lipopolysaccharide (LPS) on the hemostatic balance in the human lung, six healthy subjects inhaled nebulized LPS or saline in a randomized cross-over study and bronchoalveolar lavage fluid was obtained six hours thereafter. LPS induced soluble tissue factor and thrombin-antithrombin complexes and inhibited plasminogen activator activity in BALF. Additionally plasminogen activator inhibitor type 1 production was upregulated after LPS inhalation. LPS also elicited local activation of neutrophils (release of elastase, myeloperoxidase and bactericidal/permeability increasing protein) and secretion of interleukin (IL)-6 and IL-8. Inhalation of LPS by healthy humans reproduces major features of the procoagulant response to inflammatory and infectious lung diseases and may be used as a novel model to evaluate pathogenetic mechanisms and new interventions.

Background Current guidelines generally recommend a combination of inhaled corticosteroids and a Beta(2)-agonist for persistent asthma. The adjustment of anti-inflammatory therapy in persistent asthma is advised to be guided mainly by the presence of symptoms. Objective To investigate whether clinically masked increases in bronchial inflammation occur in guideline-treated, persistent asthma following allergen exposure. Methods After a 4-week steroid-run-in period (fluticasone 250 μg twice daily) 48 allergic patients with persistent asthma underwent a bronchial challenge with a single dose of allergen, after inhalation of salbutamol (400 μg, nebulized dose). FEV 1 and sputum markers of bronchial inflammation were measured before and after allergen challenge. Furthermore, additional rescue-salbutamol usage was recorded following allergen challenge. Results After allergen challenge there was a significant increase in sputum eosinophil numbers (geometric mean number ×10 4 /g [95% CI]: 0.5 [0.3; 1.0] before, and 2.4 [1.3; 4.2] after challenge, p = 0.01 ). The mean change in FEV 1 between 4 and 8 h after challenge relative to baseline was −0.04% [95% CI−2.3; 2.2], p > 0.9 . None of the patients took additional rescue salbutamol over 8 h after allergen challenge. Conclusions Clinically masked increases in bronchial inflammation occur in guideline-treated, persistent asthma following allergen exposure. This finding underscores the need for additional guides for the adjustment of anti-inflammatory therapy in persistent asthma.

BACKGROUND: Adding a long-acting beta(2)-agonist to inhaled corticosteroids results in better symptomatic asthma control than increasing the dose of inhaled corticosteroids. OBJECTIVE: Investigating whether adding the long-acting beta(2)-agonist salmeterol to the inhaled corticosteroid fluticasone propionate has an effect on allergen-induced allergic inflammation in asthma. METHODS: Bronchial allergen challenges were performed in 26 patients with allergic asthma, pretreating them with a single dose of either fluticasone/salmeterol (100/50 microg) or fluticasone alone (100 microg), in a double-blind, randomized, cross-over design. Sputum and serum markers of bronchial inflammation were measured after allergen challenge, as well as lung function parameters. Primary outcomes were sputum eosinophil numbers and eosinophil cationic protein. RESULTS: Asthmatic responses after allergen challenge were significantly reduced after pretreatment with fluticasone/salmeterol relative to fluticasone alone. Sputum inflammatory markers after allergen challenge were not significantly affected by fluticasone/salmeterol pretreatment. By contrast, serum IL-5 was significantly reduced (geometric mean serum IL-5 [SEM]: 0.5 [0.3] vs 1.1 [0.3] pg/mL 1 hour and 0.6 [0.3] vs 1.1 [0.3] pg/mL 6 hours after challenge with fluticasone/salmeterol vs fluticasone alone pretreatment, respectively; P values < .05). Also, peripheral blood eosinophils were significantly reduced (geometric mean number x 10(6)/L [SEM]: 172 [0.1] vs 237 [0.1] at 6 hours and 271 [0.1] vs 351 [0.1] at 24 hours with fluticasone/salmeterol vs fluticasone alone pretreatment, respectively; P < .05). CONCLUSION: Adding salmeterol to fluticasone reduces allergen-induced serum IL-5 and peripheral blood eosinophils. This phenomenon may contribute to the improved clinical outcomes that result from adding a long-acting beta(2)-agonist to inhaled corticosteroids

For the diagnosis of asthma in young children, GPs have to rely on history taking and physical examination, as spirometry is not possible. The additional diagnostic value of specific immunoglobulin E (IgE) to inhalent allergens remains unclear.To assess the predictive accuracy of specific IgE to cat, dog, and/or house dust mites in young children for the subsequent development of asthma at the age of 6 years.Prospective follow-up study.Seventy-two general practices.A total of 654 children, aged 1-4 years, visiting their GPs for persistent coughing (/= 5 days), were tested for IgE antibodies by radio allergosorbent testing (RAST). Parents completed a questionnaire on potential risk indicators. Those children who showed an IgE-positive status (12.7%) and a random sample of those with an IgE-negative status (0.5 U/ml) were followed up to the age of 6 years when the asthma status was established. The main outcome measure was asthma at the age of 6 years (combination of both symptoms and/or use of asthma medication, and impaired lung function).Addition of RAST results to a prediction model based on age, wheeze, and family history of pollen allergy increased the area under the receiver operating characteristic (ROC) curve from 0.76 to 0.87. Furthermore, RAST improved patient differentiation as indicated by a change in the range of asthma probabilities from 6-75% before the IgE test, to 1-95% after the IgE-test.Sensitisation to inhalant allergens in 1-4-year-olds, as shown by RAST, is a useful diagnostic indicator for the presence of asthma at the age of 6 years, even after a clinical history has been obtained. This model should preferably be validated in a new population before it can be applied in practice.

Rationale: Salmeterol is a beta(2)-adrenoreceptor agonist used in the treatment of obstructive pulmonary disease. Salmeterol inhibits inflammatory responses by neutrophils and mononuclear cells in vitro and in mouse models of lung inflammation in vivo. Objective: To determine the effect of salmeterol on LPS-induced lung inflammation in humans. Methods: Thirty-two healthy subjects were enrolled in a single-blinded, placebo-controlled study. Subjects inhaled 100 mu g salmeterol or placebo (t = -0.5 h) followed by 100 mu g LPS or normal saline (t = 0 h; n = 8/group). Measurements were performed in bronchoalveolar lavage fluid and purified alveolar macrophages obtained 6 h post-challenge. Measurements and Main Results: Inhalation of LIPS was associated with neutrophil influx, neutrophil degranulation (myeloperoxidase, bactericidal/permeability-increasing protein and elastase), release of cytokines (tumor necrosis factor a and interleukin 6) and chemokines (interleukin 8, epithelial cell-derived neutrophil attractant 78, macrophage inflammatory proteins 1 alpha and 1 P), activation of alveolar macrophages (upregulation of HLA-DR and CD71; enhanced expression of mRNAs for 13 different mediators of inflammation), and protein leakage (all p

Background. Several studies have shown an inverse association between helminth infections and atopy, but none have clearly established that the pathogens themselves, rather than other associated factors, cause the suppression of atopy. To show a direct link, prospective intervention studies are required. Methods. A randomized, controlled trial was performed to study whether repeated anthelminthic treatment results in increased allergic sensitivity to house dust mites (HDMs) in chronically infected children. The trial population consisted of 317 Gabonese schoolchildren with a high prevalence of intestinal helminths. Intervention consisted of treatment every 3 months with praziquantel and mebendazole and with placebo in the control group. Follow-up lasted 30 months: at 6-month intervals, skin-test sensitivity to mites, helminth infection status, and levels of total IgE were determined. Results. Treatment resulted in a significant increase in the rate of developing skin sensitivity to HDMs ( hazard ratio, 2.51; 95% confidence interval, 1.85-3.41), which was mediated, in part, by reductions in Ascaris and/or Trichuris infections. Levels of total IgE were reduced, but this did not mediate the effect of treatment on skin-test reactivity. Conclusions. Anthelminthic treatment of chronically infected children results in increased atopic reactivity, which indicates that helminths directly suppress allergic reactions

Background and Objective: The goal of this study was to determine the influence of chronic obstructive pulmonary disease (COPD) on health-related quality of life (HRQL) independent of comorbidity. Methods: Patients with COPD in general practice, greater than or equal to 40 years, were selected. To recruit controls, a random sample of persons without COPD and greater than or equal to 40 years, was taken. HRQL was assessed with the SF-36 and comorbidity was determined by questionnaire. Results: The influence of COPD on HRQL independent of comorbidity (represented by adjusted regression coefficients) was significant for physical functioning (-27.6), role functioning due to physical problems (-21.6), vitality (-14.4), and general health (-25.7), and was minor and not significant for social functioning (-5.6), mental health (-1.3), role functioning due to emotional problems (-2.7), and bodily pain (-2.5). Comorbidity contributed significantly to the HRQL of all domains (-7.6 to -27.1). Conclusion: COPD patients can be impaired in all domains of HRQL. However, impairments in physical functioning, vitality, and general health are related to COPD and to some extent to comorbidity, while impairments in social and emotional functioning do not seem to be related to COPD, but only to comorbidity. (C) 2003 Elsevier Inc. All rights reserved

It has been reported that serum immunoglobulin E (IgE) from certain atopic patients can sensitize basophils to release histamine in response to IgE-dependent histamine-releasing factors (HRFs). It has also been shown that patients suffering from severe forms of atopy may contain IgE autoantibodies. It was investigated whether HRF-responsive sera contained IgE autoantibodies and if there was an association between IgE autoreactivity and IgE-dependent responsiveness to HRF. The presence of HRF-responsive IgE (IgE+) in serum of patients with respiratory atopy was determined by stimulating stripped human basophils sensitized by serum with peripheral blood mononuclear cell (PBMC)-derived HRF, and measuring the release of histamine. In parallel, these sera were screened for the presence of IgE autoantibodies to nitrocellulose-blotted human cellular extracts. The capacity of IgE autoantigen-containing preparations to induce histamine release was tested in the stripped basophil assay. Eleven out of 52 sera contained IgE autoantibodies to blotted cellular extracts of human PBMCs or of the human epithelial cell line A431. No significant association was found between IgE autoreactivity and IgE-dependent responsiveness to HRF: 7/26 IgE+ sera contained IgE to human cellular extracts, and 4/26 of the sera without IgE+ did also. IgE autoantigen-containing extracts did not induce histamine release of appropriately sensitized basophils. By size-exclusion chromatography it was shown that a 32 000 MW autoantigen eluted in the >55 000 MW fraction, which indicates that this protein forms polymers or complexes with other macromolecules. This might explain the discrepancy between binding and histamine-releasing activity. A 20 000 MW IgE-defined autoantigen cross-reacted with a shrimp allergen. Our results indicate that IgE-reactivity to immunoblotted human protein and IgE-dependent HRF activity are distinct entities that may co-occur in atopic patients.

Background: Activity of immunoglobulin (Ig)E-dependent histamine-releasing factor (HRF) is dependent on the IgE molecules bound to the surface of basophils. Sera capable of passively sensitizing basophils to release histamine to HRF were designated IgE(+) sera. IgE(+) and HRF have been suggested to play a role in late allergic reaction (LAR). Objective: The working hypothesis was tested that IgE(+) induces a LAR. Further, activity of HRF produced by mononuclear cells (HRFmn) was compared with that of recombinant HRF p23. Methods: Atopic patients (n = 82) were bronchially provoked with Dermatophagoides pteronyssinus extract and the change in forced expiratory volume in I second was monitored. A LAR was defined as forced expiratory volume in 1 second as percentage of baseline

The 5q31-33 region contains a number of genes, including IL-13, that could be involved in the aetiology of allergic asthma. Other genes of interest in this region are the Th2 cytokines IL-4 and IL-5; the p40 chain of the Th1 cytokine inducer IL-12; IL-3; IL-9; CD14; the s2-adrenergic receptor; the corticosteroid receptor and the transcription factors interferon regulatory factor 1 (IRF1) and transcription factor 7 (TCF7). Other diseases such as schistosomiasis may also be (at least partly) controlled by this region. In humans, the severity of infection caused bySchistosomamansoniis linked to a marker on chromosome 5g31 [1]. In mice, resistance to this disease is regulated by Th2 cytokines; in particular, neutralisation of IL-13 leads to reduced pulmonary granuloma formation and total serum IgE levels, while Th2 cytokine production (IL-4, IL-5, and IL-13) remains intact [2]. The important role of Th2 cytokines in human asthma is underscored by the fact that T cells, acquired from bronchial biopsies, display an increased capacity to produce IL-4, IL-5, and IL-13 [3-5]. Interestingly, pulmonary expression of IL-13 is observed in both allergic and non-allergic asthma [6], while IL-4 expression may be more restricted to allergic asthma [7]. Because of their biological effects, IL-4 and IL-13, located at close proximity on 5q31, are very likely candidates to be involved in the inheritance of asthma. Firstly, for an antibody isotype switch to IgE, B cells require stimulation by either IL-4 or IL-13 [8, 9], and secondly these cytokines induce VCAM-1 expression on endothelial cells and pulmonary fibroblasts [10, 11], which may cause the accumulation of eosinophils at the site of the allergic reaction [12]. Recently, other effector functions of IL-4 and IL-13 have been described, independent of IgE and eosinophils.

In the liver, paracrine interaction between Kupffer cells and hepatocytes influences glucose metabolism. In vitro in rats, nitric oxide (NO), a paracrine mediator, inhibits several pathways of hepatic glucose production. The role of NO on glucose production has not been studied in vivo in humans. Glucose production was measured during N-G-monomethyl-L-arginine, monoacetate salt (L-NMMA) infusion, an inhibitor of NO synthesis in vivo, in 6 healthy men fasting 23 hours in a saline-controlled crossover study. During L-NMMA infusion, NO output decreased 40% to 50%, peripheral vascular resistance increased approximately 22%, and cardiac output (CO) decreased approximately 14%. The decrease in glucose production was not different between L-NMMA and saline. Glucose concentration, substrate supply, and glucoregulatory hormone concentrations were not different; epinephrine was lower with L-NMMA. A 40% to 50% inhibition of NO synthesis in vivo in humans does not affect glucose production during short-term fasting. The hypothesis that NO is an important modulator of basal glucose production in healthy humans in vivo should therefore be rejected. Copyright (C) 2002 by W.B. Saunders Company

We have investigated whether increased plasma protein leakage is present early after segmental allergen challenge in allergic asthma. Seven asthmatic subjects with mild allergy (AA group) and 5 non-asthmatic subjects with allergy (ANA group) were challenged with allergen doses based on similar early skin reactions; 5 healthy control subjects without allergy (C group) were challenged with the highest dose applied in the subjects with allergy. Bronchoalveolar lavage (BAL) fluid was obtained before, at 5 minutes after, and at 4 hours after challenge from different segments. Levels of albumin (Alb) and alpha2-macroglobulin (A2M) were measured in BAL fluid and serum. In addition, we calculated the relative coefficient of excretion as follows: RCE = ((A2M in BAL fluid)/(A2M in serum))/((Alb in BAL fluid)/(Alb in serum)). Also, levels of tryptase as a marker of mast cell activation and tumor necrosis factor-alpha (TNF-alpha), a possible inducer of plasma protein leakage, were determined. At 5 minutes after challenge, in none of the groups was a significant change found in the parameters for protein leakage. Levels of tryptase were increased in the subjects with allergy at 5 minutes after challenge only (P = .004). At 4 hours after challenge, levels of Alb (P = .03) and A2M (P = .04) and the RCE (P = .04) were increased in the AA group only. At 4 hours, levels of TNF-alpha were increased, with no significant differences among the three groups. In the asthmatic subjects with allergy, levels of TNF-alpha correlated with levels of Alb (r = 0.85, P = .02). In conclusion, at 4 hours after segmental allergen challenge, plasma protein leakage was increased in the asthmatic subjects only. The increase in levels of TNF-alpha in all groups indicates that the presence of TNF-alpha alone was not sufficient to cause plasma protein leakage within 4 hours after allergen challenge. Our results confirm the concept that plasma exudation after allergen exposure is a pathophysiologic event associated with asthma.