Indoor allergens and asthma: report of the Third International Workshop.
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Air Pollution, Indoor↗
Biomedical subjects
M D Chapman
Publications and source records attributed to M D Chapman.
Immune memory in CD4+ CD45RA+ T cells.
This study addresses the question of whether human peripheral CD4+ CD45RA+ T cells possess antigen-specific immune memory. CD4+ CD45RA+ T cells were isolated by a combination of positive and negative selection. Putative CD4+ CD45RA+ cells expressed CD45RA (98.9%) and contained < 0.1% CD4+ CD45RO+ and < 0.5% CD4+ CD45RA+ CD45RO+ cells. Putative CD45RO+ cells expressed CD45RO (90%) and contained 9% CD45RA+ CD45RO+ and < 0.1% CD4+ CD45RA+ cells. The responder frequency of Dermatophagoides pteronyssinus-stimulated CD4+ CD45RA+ and CD4+ CD45RO+ T cells was determined in two atopic donors and found to be 1:11,314 and 1:8031 for CD4+ CD45RA+ and 1:1463 and 1:1408 for CD4+ CD45RO+ T cells. The responder frequencies of CD4+ CD45RA+ and CD4+ CD45RO+ T cells from two non-atopic, but exposed, donors were 1:78031 and 1:176,903 for CD4+ CD45RA+ and 1:9136 and 1:13,136 for CD4+ CD45RO+ T cells. T cells specific for D. pteronyssinus were cloned at limiting dilution following 10 days of bulk culture with D. pteronyssinus antigen. Sixty-eight clones were obtained from CD4+ CD45RO+ and 24 from CD4+ CD45RA+ T cells. All clones were CD3+ CD4+ CD45RO+ and proliferated in response to D. pteronyssinus antigens. Of 40 clones tested, none responded to Tubercule bacillus purified protein derivative (PPD). No difference was seen in the pattern of interleukin-4 (IL-4) or interferon-gamma (IFN-gamma) producing clones derived from CD4+ CD45RA+ and CD4+ CD45RO+ precursors, although freshly isolated and polyclonally activated CD4+ CD45RA+ T cells produced 20-30-fold lower levels of IL-4 and IFN-gamma than their CD4+ CD45RO+ counterparts. Sixty per cent of the clones used the same pool of V beta genes. These data support the hypothesis that immune memory resides in CD4+ CD45RA+ as well as CD4+ CD45RO+ T cells during the chronic immune response to inhaled antigen.
Allergens↗
Evaluation of German cockroach (Orthoptera:Blattellidae) allergen and seasonal variation in low-income housing.
Six apartments in a low-income housing project were evaluated for German cockroach. Blattella germanica (L.), infestation and concentration of an allergen derived from these cockroaches (Bla g II). Kitchen and living room samples were collected monthly for 1 yr. In addition, airborne sampling was carried out in 5 kitchens. The kitchen had the highest allergen concentration in 65% of visits and the highest number of cockroaches trapped in 69% of visits. In the kitchen, the highest cockroach levels were seen in June, whereas the values for Bla g II peaked in August. In keeping with this, the closest correlation was between Bla g II (microgram/g dust) and the number of cockroaches found 2 mo earlier. Airborne samples were assayed for 2 separate allergens. Bla g II and Bla g I. No allergen was detectable in the absence of disturbance. By contrast, during disturbance with a vacuum cleaner both Bla g II and Bla g I were detectable in the air of each apartment. Results suggest that immunochemical assay of a major allergen in dust samples from the kitchen floor may be used to monitor exposure to German cockroaches, also that cockroach levels may be used as an indicator or predictor of allergen in dust.
Allergens↗
Localization of antigenic sites on Der p 2 using oligonucleotide-directed mutagenesis targeted to predicted surface residues.
BACKGROUND: Understanding the molecular nature of allergen-antibody interactions is important to understanding the mechanism of conventional immunotherapy as well as to designing alternative immunotherapeutic strategies. Many important allergens have been cloned and expressed, making it possible to apply recombinant DNA techniques to dissect antigenic determinants. OBJECTIVE: The aim of this study was to use predictive algorithms and site-directed mutagenesis to investigate monoclonal antibody and IgE antibody epitopes of the major house dust mite allergen Der p 2. METHODS: Computer algorithms were used to assess the primary amino acid sequence of Der p 2 and to identify regions of hydrophilic and flexible sequence. Subsequently, site-directed mutagenesis was used to generate amino acid substitutions at hydrophilic residues at positions 44-46 and at position 100. The variants were tested in a competitive inhibition ELISA with four group 2-specific murine monoclonal antibodies and with human IgE antibody from mite allergic patients. RESULTS: Conservative amino acid substitutions at position 44-46 did not distinguish IgE antibody epitopes, but did suggest that these residues are involved in the epitope defined by one monoclonal antibody, 15E11. Non-conservative substitution of proline at this position reduced binding to all four monoclonal antibodies, as well as IgE antibody, by 50-80%. Point mutants at position 100 mapped the epitopes of two monoclonal antibodies, 7A1 and 13A4, previously shown to bind the same region of Der p 2. In addition, the two variants tested at this position showed distinct inhibition curves with these two monoclonal antibodies indicating differences in fine specificity. CONCLUSIONS: Using predictive algorithms, in the absence of tertiary structural information, we have been able to localize important B cell determinants on Der p 2. The results suggest that it is possible to modulate antibody recognition of allergens using site-directed mutagenesis and that this approach may provide a new strategy for allergen specific immunotherapy.
Antibodies, Monoclonal↗
Mouse/human chimeric IgG1 and IgG4 antibodies directed to the house dust mite allergen Der p 2: use in quantification of allergen specific IgG.
BACKGROUND AND OBJECTIVE: Chimeric mouse/human monoclonal IgG1 and IgG4 antibodies were developed against the house dust mite allergen Der p 2. These chimeric IgG antibodies, hIgG1-Dp2 A and hIgG4-Dp2 A, have the same binding characteristics as the previously reported chimeric hIgE-Dp2 A and are composed of the heavy chain variable domains and light chains of the original murine monoclonal antibody 2B12, whereas the heavy chain constant domains have been replaced by the human IgG1 or IgG4 heavy chain. The expression level of hIgG1-Dp2 A and hIgG4-Dp2 A was 1 and 3.5 microg/mL, respectively. METHODS AND RESULTS: Since all IgG in these culture supernatants is allergen-specific, they are useful reference reagents and enable the calculation of the amount of allergen specific IgG1 and IgG4 antibodies in absolute IgG amounts. The results obtained with two panels of sera from patients in immunotherapeutic treatment were evaluated and compared in Der p 2 IgE, IgG1 and IgG4 RAST and with reversed IgG4 RAST using labelled purified Der p 2. Close agreement between the results for the two IgG4 assays was found. CONCLUSION: With these chimeric reference reagents the quantities of isotype specific antiallergen antibodies can be calculated and compared.
Animals↗
Recombinant mite allergens. New technologies for the management of patients with asthma.
The introduction of molecular cloning techniques has led to advances in allergen identification and sequencing, production of recombinant allergens, identification of B-cell and T-cell epitopes, and tertiary structural analysis of allergen molecules. Over 10 groups of mite allergens have been cloned from Dermatophagoides spp., as well as several homologous allergens from Euroglyphus maynei, Lepidoglyphus destructor, and Blomia tropicalis. The availability of these allergens has made it feasible to consider their use for both diagnostic and therapeutic purposes. Several recombinant Dermatophagoides allergens show comparable reactivity on skin testing and in serologic assays to natural allergens, and cocktails of the recombinant proteins could be used as diagnostic reagents. New technologies have been developed for detection of allergen-specific IgE and for environmental allergen detection using rapid diagnostic tests. Novel approaches to immunotherapy are also being investigated, including T-cell-peptide based vaccines, allergen variants which lack IgE reactivity, and naked DNA vaccines. The application of allergen biotechnology should lead to improvements in the management of mite-allergic patients with asthma and represents a logical step toward reducing asthma mortality and morbidity.
Allergens↗
Aerodynamic properties of the major dog allergen Can f 1: distribution in homes, concentration, and particle size of allergen in the air.
Exposure and sensitization to dog allergen is a significant cause of asthma. In this study we investigated the distribution, aerodynamic characteristics, and particle-size distribution of the major dog allergen Can f 1. Dust samples were collected in 50 homes with a dog and 50 homes without dogs. Airborne Can f 1 concentration was measured in 28 homes with dogs and 36 homes without a dog. Particle-size distribution was determined by using 10 separate Andersen sampler measurements in a dog-handling facility, and in 10 homes with dogs, and by repeated measurements in a home with one dog. High levels of Can f 1 (> 10 microg/g) were found in dust in all but one home with a dog and in eight of 50 homes without dogs. Airborne Can f 1 levels varied greatly between the homes with dogs (range: 0.3 to 99 ng/m3). Low levels of airborne Can f 1 (range: 0.4 to 1.1 ng/m3) were detected in 11 of 36 homes without a dog. Can f 1 was predominantly associated with large particles collected on the first stage of the Andersen sampler (> 9 microm), which averaged 42 to 49% of the total allergen recovered in the dog-handling facility and in homes with dogs. Small particles (< 5 microm diameter) also carried Can f 1, and these particles comprised approximately 20% of the total airborne allergen load. There was an excellent concordance between the results obtained in different sampling areas, and between the total Can f 1 recovered on the Andersen sampler and on the parallel filter. In conclusion, airborne Can f 1 was detectable in undisturbed conditions in all homes with dogs and in almost one third of the homes without dogs. In houses with dogs, a significant proportion (approximately 20%) of airborne Can f 1 was associated with small particles (< 5 microm diameter). Owing to their aerodynamic characteristics, these particles would be expected to remain airborne for a long period and, when inhaled, could penetrate into the lower airways and initiate asthma attacks.
Air Pollution, Indoor↗
Sensitization to Blomia tropicalis in patients with asthma and identification of allergen Blo t 5.
In tropical and subtropical regions of the world, allergens produced by Blomia tropicalis are an important cause of IgE-mediated sensitization among patients with asthma. We compared the relative importance of sensitization to the two mite species among asthma patients from Florida, Puerto Rico, and Brazil (n = 83), who were concurrently exposed to B. tropicalis and D. pteronyssinus, with patients from the United States and from the United Kingdom (n = 56) exposed to D. pteronyssinus. In addition, molecular cloning techniques were used to clone and express a major B. tropicalis allergen. There were significant differences between IgE antibody responses to B. tropicalis and D. pteronyssinus that were related to exposure: only 22% of patients exposed to both species had a high ratio (> 10) of IgE D. pteronyssinus:B. tropicalis, whereas 68% of patients exposed only to D. pteronyssinus had a ratio of > 10 (p < 0.001). A major 14-kD allergen (Blo t 5), cloned from a B. tropicalis cDNA library, showed 43% sequence homology to D. pteronyssinus Der p 5. Recombinant Blo t 5 produced in E. coli reacted with 45 to 69% of sera from B. tropicalis-allergic asthmatics and induced positive immediate skin tests at 10(-3) to 1 microg/ml. In vivo and in vitro comparisons of IgE responses to B. tropicalis, D. pteronyssinus, rBlo t 5, and rDer p 5, showed that B. tropicalis has unique allergens that cause specific IgE responses. The results suggest that B. tropicalis is an independent cause of sensitization and that use of recombinant Blo t 5 should lead to a better understanding of the role of B. tropicalis in causing asthma in tropical environments.
Adolescent↗
Hypercoagulability and hypofibrinolysis in primary osteoarthritis.
Histologic evidence of venous thrombosis and lipid abnormalities have previously been reported in osteoarthritis. Hypofibrinolysis has been recorded in patients with ischemic necrosis of bone, and it has been proposed as a major cause of osteonecrosis. This study determines whether systemic evidence of coagulation and lipid abnormalities could be detected in osteoarthritis. Global and specific tests were used to assess coagulability and fibrinolysis in 44 patients with degenerative osteoarthritis of the hip and 52 matched control subjects. In patients with osteoarthritis, an increase in factor VIIlc, increased platelet sensitivity over a range of adenosine diphosphate concentrations (0.05 micromol/L-4 micromol/L) and elevated D dimer levels were found. Euglobulin clot lysis time was prolonged in this group and plasminogen activator inhibitor Type 1 activity was increased. Relative hyperlipidemia was observed in the osteoarthritis group, with increased cholesterol, low density lipoprotein cholesterol, and triglyceride levels. It is concluded that there is a hypercoagulable and prothrombotic condition in osteoarthritis, with hypofibrinolysis and indirect evidence of increased fibrin generation. The possible contribution of lipid abnormalities to hemostatic imbalance in osteoarthritis is discussed.
Aged↗
Structural and antigenic studies of cockroach allergens and their relevance to asthma.
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Allergens↗
Use of recombinant group 5 allergens to investigate IgE-mediated sensitization to Blomia tropicalis and Dermatophagoides pteronyssinus.
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Allergens↗
Reduction of IgE antibody binding to rDer p 2 variants generated by site-directed mutagenesis.
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Antigens↗
Exposure to house dust mite allergens and the clinical activity of asthma.
BACKGROUND: House dust mite allergens play an important role in inducing IgE-mediated sensitization and the development of bronchial hyperresponsiveness (BHR) and asthma. This study investigated the relationship between mite allergen exposure and the clinical activity and severity of asthma. METHODS: Nonsmoking adult patients with asthma (n = 53) were randomly recruited from the asthma registry of two large family practitioner surgeries. Each participant underwent skin testing with common inhalant allergens, a methacholine bronchoprovocation test, and pulmonary function testing on up to 3 separate occasions over a 4-week period. BHR was expressed both as PD20 and dose-response ratio (DRR), and the patients with patients with PD20 of less than 12.25 mumol methacholine were classified as methacholine reactors. Patients were also asked to record peak expiratory flow rate (PEFR) values at 2-hour intervals during waking hours for 1 month. Daily PEFR variability was calculated as amplitude percent mean. Dust samples were collected by vacuuming bedding, bedroom carpets and mattresses. In addition, in the homes of 32 subjects with positive skin test responses to mites, airborne samples were taken overnight for 8 hours with a personal sampler attached to each subject's pillow. Der p 1 and Der p 2 levels were determined by a two-site monoclonal antibody-based ELISA. RESULTS: No difference in mite exposure was found between subjects who were sensitive to mites and those who were not. However, mite-sensitive methacholine reactors were exposed to significantly higher concentrations of Der p 1 in beds than mite-sensitive methacholine nonreactors (13.2 micrograms/gm and 1.45 micrograms/gm, respectively; p < 0.02). Der p 1 and Der p 2 were undetectable in 30 of 32 airborne samples. In mite-sensitive patients both Der p 1 and Der p 2 in beds significantly correlated with BHR (PD20: r = -0.49, DRR, r = 0.49; PD20: r = -0.46, DRR: r = 0.43) and amplitude percent mean PEFR (r = 0.38, r = 0.41) for Der p 1 and Der p 2, respectively. There was a significant negative correlation between exposure to Der p 1 and percent predicted FEV1 (r = -0.43). The correlation between Der p 2 and percent predicted FEV1 just failed to reach a significant level but showed a clear trend ( r = -0.35, p = 0.068). CONCLUSIONS: Clinical activity and severity of asthma (measured by the level of BHR, PEFR variability, and percent predicted FEV1) in mite-sensitive patients is related to exposure to mite allergens in the dust reservoir, with levels in bed being an important indicator that correlated with disease activity.
Adult↗
New mattresses: how fast do they become a significant source of exposure to house dust mite allergens?
BACKGROUND: Sensitization and exposure to mite allergens is a major risk factor for asthma. Little is known about the rate of build-up of allergens in the mite micro-habitats. OBJECTIVES: To investigate the rate of increase in mite allergen levels in new mattresses. METHODS: Der p 1 was measured in the dust samples collected from six identical new single mattresses over a period of 2 years. RESULTS: Der p 1 increased significantly at 4 months as compared with baseline level (P < 0.01), but no difference was found between the concentrations at 4, 8, 12 and 24 months. There was a significant correlation between Der p 1 concentration in mattresses at 4, 8, 12 and 24 months and Der p 1 levels in the bedroom carpet at the beginning of the study. CONCLUSIONS: New mattresses can become a significant source of exposure to mite allergens after a short period of time (< 4 months). There is little justification for advising mite sensitive patients to replace their mattresses as a part of avoidance regime.
Allergens↗
Domestic allergens in public places. II: Dog (Can f1) and cockroach (Bla g 2) allergens in dust and mite, cat, dog and cockroach allergens in the air in public buildings.
BACKGROUND: Sensitization and exposure to indoor allergens are the major risk factors for asthma. It is possible that significant exposure to domestic allergens occurs outside the home. OBJECTIVES: To investigate the levels of Can f 1 and Bla g 2 in the dust from carpeted floors and upholstered seats in public buildings and public transport and the airborne concentrations of Der p 1, Fel d 1, Can f 1 and Bla g 2 in schools and offices. METHODS: Can f 1 and Bla g 2 were measured in the dust collected by vacuuming a 1 m2 area of carpet, as well as upholstered seats in five schools, six hotels, four cinemas, six pubs, three buses and two trains. Dust was also collected from the bedroom carpet, living room carpet, mattress and sofa in 20 homes with and 20 homes without a dog in the same area. Personal airborne sampling (2 L/min) was conducted for 8 h in offices (n = 16) and classrooms (n = 9). In addition, airborne samples in schools were collected using a high volume pump (60 L/min) for 1 h in three classrooms immediately after the children vacated the school. Can f 1, Bla g 2, Der p 1 and Fel d 1 were assayed using a two-site monoclonal antibody-based ELISA. RESULTS: Can f 1 was detected in all dust samples from public places, ranging from 0.2 to 52.5 micrograms/g. Significantly higher levels were found in upholstered seats (geometric mean--GM 9.4 micrograms/g) than in carpets (GM 1.5 micrograms/g; P < 0.001), and levels of Can f 1 > 10 micrograms/g were found in 40% of upholstered seats in public places. Can f 1 was significantly higher in upholstered seats in public places than in sofas in homes without a dog (GM 1.8 micrograms/g; P < 0.001). Detectable levels of Bla g 2 were found in all of the schools (GM 2.4 U/g, range 0.8-4.4 U/g). Bla g 2 concentration greater than 2U/g (provisional threshold level representing risk of sensitization) was measured in 65% of the classrooms sampled. Der p 1 and Bla g 2 were below the detection limit in all airborne samples. However, airborne Fel d 1 and Can f 1 were detected in schools and offices, albeit in low concentrations. CONCLUSIONS: Upholstered seats from public places constitute a reservoir for the accumulation of dog allergen, and a source of exposure to Can f 1 inside public buildings or on public transport. Exposure to cockroach allergens in schools may be important for cockroach sensitized asthmatic children.
Air Pollution, Indoor↗
Changing concepts of allergic disease: the attempt to keep up with real changes in lifestyles.
In the past 100 years, changes have occurred in the outdoor environment and in houses that have contributed to the increased prevalence of hay fever and asthma. Much evidence indicates that exposure to indoor allergens is an important cause of asthma. Changes in housing that have contributed to the increased prevalence and severity of asthma include increased temperature, decreased ventilation, and permanent carpeting. In addition to these changes, geographic differences in allergens, deficiencies in cleanliness, poor health care, passive smoke, and lack of exercise also contribute to the increase in severity of asthma that has occurred. The management of asthma includes controlling exposure to indoor allergens and seeking additional treatable causes of asthma (e.g., fungal allergens). Changes will continue to occur, and physicians who treat allergic diseases should become involved in the design of houses to limit exposure. Many questions regarding allergen measurement and control remain.
Allergy and Immunology↗
Cloning of cockroach allergen, Bla g 4, identifies ligand binding proteins (or calycins) as a cause of IgE antibody responses.
An allergen cloned from a Blattella germanica (German cockroach) cDNA library, encoded a 182-amino acid protein of 20,904 Da. This protein, designated B. germanica allergen 4 (Bla g 4), was expressed as a glutathione S-transferase fusion protein in Escherichia coli and purified by affinity chromatography and high-performance liquid chromatography. The prevalence of serum IgE antibody to recombinant Bla g 4 in 73 cockroach allergic patients with asthma ranged from 40% (antigen binding radioimmunoassay) to 60% (plaque immunoassay). Cockroach allergic patients gave positive intradermal skin tests to recombinant Bla g 4 at concentrations of 10(-3)-10(-5) micrograms/ml, whereas non-allergic controls, or cockroach allergic patients with no detectable serum IgE antibody to Bla g 4, gave negative skin tests to 1 microgram/ml. Polymerase chain reaction and Southern analysis identified a 523-base pair DNA encoding Bla g 4 in both B. germanica and Periplaneta americana (American cockroach). However, Northern analysis showed that mRNA encoding Bla g 4 was transcribed in B. germanica but not in P. americana, suggesting that allergen expression was species specific. Sequence similarity searches showed that Bla g 4 was a ligand binding protein or calycin and unexpectedly revealed that this family contained several important allergens: beta-lactoglobulin, from cow milk, and rat and mouse urinary proteins. Although the overall sequence homology between these proteins was low (approximately 20%), macromolecular modeling techniques were used to generate two models of the tertiary structure of Bla g 4, based on comparisons with the x-ray crystal coordinates of bilin binding protein and rodent urinary proteins. The results show that members of the calycin protein family can cause IgE antibody responses by inhalation or ingestion and are associated with asthma and food hypersensitivity.
Allergens↗
Molecular cloning of a major cockroach (Blattella germanica) allergen, Bla g 2. Sequence homology to the aspartic proteases.
Inhalation of allergens produced by the German cockroach (Blattella germanica) elicits IgE antibody formation and the development of asthma in genetically predisposed individuals. We compared the allergenic importance of two cockroach (CR) allergens, Bla g 1 and Bla g 2, and determined the complete amino acid sequence of the major 36-kDa allergen, Bla g2. A survey of 106 sera from CR allergic patients showed the prevalence of IgE antibodies to Bla g 1 and Bla g 2 to be 30.2% and 57.6%, respectively. Immediate skin tests on 7 selected patients gave positive reactions using 10(-3) micrograms/ml either allergen, whereas controls showed no response to 10 micrograms/ml. Natural Bla g 2 was purified and the sequence of the NH2 terminus and tryptic peptides, comprising 36% of the molecule, was determined. The cDNA for Bla g 2 was cloned from a B. germanica expression library and encoded a 24-amino acid signal peptide and a 328-amino acid mature protein, which showed the highest degree of identity to mosquito (Aedes aegypti) lysosomal aspartic protease (30.8%), with similar identity to pepsin, cathepsins D and E, renin, and chymosin. Bla g 2 mRNA and protein were detected in B. germanica, but not in Periplaneta americana, the other principal domiciliary CR species in the U.S. High concentrations of Bla g 2 were found in CR digestive organs (esophagus, gut, and proventriculus). The results show that Bla g 2 is a major species-specific allergen of B. germanica and suggest that the allergen functions as a digestive enzyme in the cockroach.
Allergens↗