Martin D. Chapman

Indoor allergens and asthma:Report of the Third International Workshop

In parallel with changes in lifestyle over the last 50 years (sedentary living in warm houses with extensive furnishing and low ventilation rates), there has been a progressive increase in the prevalence and morbidity of asthma in many parts of the world. The increase has been in perennial rather than seasonal asthma, and a large proportion of the patients are sensitized to one or more of the allergens found predominantly inside houses, that is, indoor allergens. The Third International Workshop on Indoor Allergens and Asthma was designed to discuss recent progress in basic and clinical research in this area, to formulate recommendations for allergen-specific management of asthma, and to consider future research directions. As with the two previous workshops, discussion topics included biology; allergen immunochemistry; molecular biology and immune response; epidemiology of asthma; and the role of allergen avoidance, a, 2 Because of dramatic progress in recent years, the Third International Workshop was expanded to cover not only house dust mite allergens but also allergens from cat, dog, and cockroach, for which immunochemical and epidemiologic data are available. Over the past 5 years there have been significant advances in several areas of research on indoor allergens, including: (1) cloning and expression of recombinant allergens, 3-7 (2) analysis of T-cell responses to indoor allergens, derivation of T-cell clones, and analysis of T-cell epitope specificity and cytokine profiles, s, 9 (3) investigation of the dose-response relationship between exposure to mite, cat, and cockroach allergens and sensitization, 1°-13 and (4) epidemiologic studies on indoor allergens as risk factors for the symptoms of asthma and bronchial hyperreactivity (BHR)? 4-17 Better definition of the allergens has made it possible to analyze their structure and biologic function and to define epitopes recognized by antibodies or T cells. Information obtained from those studies has provided exciting possibilities for developing new vaccines for safe and effective immunotherapy. 9, 18. 19 Studies of T-cell responses to dust mites have confirmed the dominance of T-helper cell (Tin) responses in allergic individuals.

A two-site monoclonal antibody ELISA for the quantification of the major Dermatophagoides spp. allergens, Der p I and Der f I

A two-site monoclonal antibody (Mab) ELISA was developed to measure the Group I allergens from Dermatophagoides spp., Der p I from D. pteronyssinus and Der f I from D. farinae. Species-specific Mabs were used to coat microtiter plates which were then incubated with allergen or house dust extracts. Bound allergen was detected using a biotinylated Mab which recognized a common epitope on both Der p I and Der f I, followed by the addition of streptavidin-peroxidase and ABTS/H2O2 substrate. The assay had low non-specific binding (approximately 0.08 absorbance units) and had a sensitivity of 5 ng/nl for aqueous allergen extracts (equivalent to 0.1 microgram allergen/g dust). 53 dust samples were assayed using the Mab ELISA and an RIA previously described using 125I-labelled Mab. The results showed a very good quantitative correlation between the assays (r = 0.96, p less than 0.001 for Der p I; r = 0.92, P less than 0.001 for Der f I). A further 132 dust samples from a different geographical areas were also assayed by both methods and gave correlation coefficients of 0.90 (P less than 0.001) and 0.86 (P less than 0.001) for Der p I and Der f I, respectively. The Mab ELISA will be useful in epidemiological studies of allergic asthma, both in the assessment of levels of dust mite allergen present in houses and the efficacy of allergen avoidance regimes.

Microarrayed allergen molecules: diagnostic gatekeepers for allergy treatment

Type I allergy is an immunoglobulin E (IgE)‐mediated hypersensitivity disease affecting more than 25% of the population. Currently, diagnosis of allergy is performed by provocation testing and IgE serology using allergen extracts. This process defines allergen‐containing sources but cannot identify the disease‐eliciting allergenic molecules. We have applied microarray technology to develop a miniaturized allergy test containing 94 purified allergen molecules that represent the most common allergen sources. The allergen microarray allows the determination and monitoring of allergic patients’ IgE reactivity profiles to large numbers of disease‐causing allergens by using single measurements and minute amounts of serum. This method may change established practice in allergy diagnosis, prevention, and therapy. In addition, microarrayed antigens may be applied to the diagnosis of autoimmune and infectious diseases.

Risk factors for asthma in inner city children

Inner city children have the highest prevalence and the highest mortality rates for asthma in the United States. The purpose of this study was to evaluate sensitization and exposure to common indoor allergens among children aged 3 years to 15 years seen for treatment of asthma at Grady Memorial Hospital, Atlanta, Ga. Eighty children in this study were enrolled in the emergency department and 64 in hospital clinics. Dust from 57 homes, assayed for three indoor allergens (dust mite, cat, and cockroach), revealed similar exposure for asthma and control groups. Sixty-nine percent of the children with asthma had IgE antibodies to dust mite, cockroach, or cat; only 27% of the control subjects were similarly sensitized (p < 0.001). Of 35 children with asthma 21 had both sensitization and significant exposure to the relevant allergen; this was true for only 3 of 22 control subjects (odds ratio, 9.5; p < 0.001). Neither sensitization nor exposure to cat allergen was common in this population. The results show that black children in inner city Atlanta are exposed to high levels of mite and cockroach allergens and that a high proportion of the children with asthma are sensitized to these allergens; the combination of sensitization and exposure is a major risk factor for asthma in this population.

Epidemiology of acute asthma: IgE antibodies to common inhalant allergens as a risk factor for emergency room visits

In recent years the morbidity and mortality of asthma has increased, although the etiology is still poorly understood. Most patients with asthma suffer acute attacks that are commonly treated in hospital emergency rooms (ER). In the present study, asthma in adults was studied with acute attacks as a marker for the disease; 102 patients first observed at a university hospital ER with acute airway obstruction were compared to 118 patients observed at the same ER with any diagnosis other than shortness of breath to evaluate allergy as a risk factor for asthma in adults. Sera were assayed for IgE antibody (Ab) to dust mites, cockroach, cat dander, and grass and ragweed pollen. The results demonstrate that in adults younger than 50 years of age, the prevalence of IgE Abs was fourfold greater among subjects with asthma than among control subjects (46/67 versus 12/81; odds ratio, 10.1; 95% confidence interval, 4.9 to 20.7). The population attributable risk for the presence of IgE Ab to one of the five allergens was greater than 50%. Among individuals older than 50 years of age, the prevalence of serum IgE Abs was not significantly increased among patients with acute airway obstruction. In the whole group, the prevalence of IgE Abs to different allergens demonstrated significant seasonal and socioeconomic differences, suggesting that the associated risk is related to exposure to those allergens. The results establish that, with acute attacks of asthma as a marker for adult asthma, the presence of serum IgE Abs to common inhalant allergens is a major risk factor.(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitative assessment of exposure to dog (Can f 1) and cat (Fel d 1) allergens: Relation to sensitization and asthma among children living in Los Alamos, New Mexico

BACKGROUND Our objective was to identify the allergens associated with asthma among schoolchildren in an area of the United States where dust mite growth is expected to be poor. Los Alamos, N.M., was chosen because it has low rainfall and is at high altitude (7200 feet) making it very dry. One hundred eleven children (12 to 14 years old) from the middle school who had been previously classified according to bronchial hyperreactivity to histamine (BHR) were studied. METHODS Sera were assayed for IgE antibodies to mite, cat, dog, cockroach, Russian thistle, and grass pollen, with both CAP system fluoroimmunoassay (Kabi Pharmacia, Uppsala, Sweden) and conventional RAST. Allergens were measured in dust samples from 108 homes with two-site assays for mite (Der p 1 and Der f 1), cat (Fel d 1), dog (Can f 1), and cockroach (Bla g 2). RESULTS Concentrations of dog and cat allergens were elevated in almost all houses with pets but were also high in a significant proportion of the houses without pets. Levels of mite allergen were less than 2 micrograms/gm in 95% of the houses, and cockroach was undetectable in all but two of the houses. Among the 21 with BHR who had symptoms, 67% had IgE antibody to dog and 62% had IgE antibody to cat. For these allergens IgE antibody was strongly associated with asthma (p < 0.001). By contrast, the presence of IgE antibody to mite, cockroach, or grass pollen was not significantly associated with asthma. CONCLUSION The high prevalence of IgE antibody to cat and dog allergens among these children is in keeping with the presence of cat and/or dog allergen in most of the houses. Furthermore, sensitization (as judged by IgE antibodies) to cat and dog allergens was strongly associated with asthma. On the other hand, no clear relationship was found between sensitization or symptoms and the current level of allergen in individual houses. The results show that in this mite-and cockroach-free environment sensitization to domestic animals was the most significant association with asthma.

Antigenic and structural analysis of group II allergens (Der f II and Der p II) from house dust mites (Dermatophagoides spp)

Monoclonal antibody affinity chromatography was used to purify two homologous mite allergens, Der f II from Dermatophagoides farinae and Der p II from D. pteronyssinus. They have the same molecular weight (MW) (15 kd) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, they have similar amino acid compositions, and their N-terminal amino acid sequences differ in only four of the first 35 residues. An excellent correlation was observed between IgE antibody to Der f II and Der p II measured in sera from 65 mite-allergic patients (r = 0.94; p less than 0.001) and between quantitative intradermal skin tests to both allergens. A third allergen (Der f III, MW 29 kd) was purified from D. farinae by repeated gel filtration. In sera from 51 mite-allergic patients, IgE antibody to Der f II, Der f III, and previously purified Der f I (MW 24 kd) was detected in 92%, 16%, and 78% of the sera by radioimmunoassay, respectively. Most patients, 41/51 (80%), demonstrated IgE antibody to more than one allergen. With monoclonal antibodies fully cross-reactive with Der f II and Der p II, a two-site immunoassay was developed for measuring absolute quantities (nanograms or micrograms) of these allergens. In extracts rich in mite-fecal material (n = 5), Der f I and Der p I (group I allergens) and Der f II and Der p II (group II allergens) were measured in ratios of 11:1 to 35:1. Lower ratios (1.1:1 to 7:1) were observed in mite body extracts (n = 6). These experiments clearly define a second group of major dust mite allergens that demonstrate extensive structural and antigenic homology.

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 subjects 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.

The effect of cat removal on allergen content in household-dust samples

To evaluate the effect of cat removal on cat-allergen content in the home, serial house dust samples were collected from 15 homes during a 9- to 43-week period after cat removal. Samples were obtained with a hand-held vacuum cleaner, and allergen content was quantitated by a radioimmunoassay specific for the major cat allergen, Fe1 d I. Baseline Fe1 d I content ranged from 7.8 Food and Drug Administration units per gram of dust to 436.7 U/gm (median 61.2 U/gm), consistent with levels found in homes with a pet cat. Fe1 d I levels declined gradually in most homes, and by 20 to 24 weeks after cat removal, eight of 15 reached levels consistent with levels found in control homes without cats. In two of those homes, allergen levels fell much more rapidly after aggressive environmental control measures were undertaken. In the other seven homes, however, the decline occurred at a much slower rate, with three homes demonstrating persistent elevations in Fe1 d I content for 20 or more weeks. These data demonstrate that the task of allergen elimination from an indoor environment is extremely difficult, even when the source of a specific allergen can be identified and removed.

Environmental exposure to cockroach allergens: Analysis with monoclonal antibody-based enzyme immunoassays

Quantitative two-site monoclonal antibody (MAb)-based enzyme-linked immunoassays for two cockroach (CR) allergens, Bla g I and Bla g II, have been developed and used to measure allergen levels in house-dust samples. Dust collected from the CR-infested homes of two patients with asthma from Charlottesville, Va., demonstrated wide variation in the levels of Bla g I, depending on the location of dust collection. Dust from kitchen floors and cabinets contained 50-fold more allergen (mean, 10,755 U/gm of dust) than dust from bedrooms and upholstered furniture (mean, 204 U/gm). One hundred forty-five dust samples were collected from the bedrooms and living rooms of 22 children with asthma and 16 control subjects without asthma living in Atlanta, Ga. Twenty-seven of the 38 homes (17/22 children with asthma; 10/16 control subjects) had detectable Bla g I (4 to 1340 U/gm of dust). Bla g II levels were assayed in 40 kitchen, bedroom, and living room samples from homes in Wilmington, Del. Highest levels of Bla g II were detected in kitchen-floor dust (300 U/gm of dust). Additionally, approximately 20% of homes with no visual evidence of CR infestation had significant levels of Bla g II in at least one dust sample (greater than 4 U/gm of dust). Our results demonstrate that CR may be an occult allergen in homes. The kitchen appears to be the primary site of CR-allergen accumulation, but significant CR-allergen levels can also be found at other sites in the home. The MAb-based assays can be used for quantitation of environmental exposure to CR allergens.(ABSTRACT TRUNCATED AT 250 WORDS)