John W. Vaughan

Sensitisation, asthma, and a modified Th2 response in children exposed to cat allergen: a population-based cross-sectional study

BACKGROUND Although asthma is strongly associated with immediate hypersensitivity to indoor allergens, several studies have suggested that a cat in the house can decrease the risk of asthma. We investigated the immune response to cat and mite allergens, and asthma among children with a wide range of allergen exposure. METHODS We did a population-based cross-sectional study of children (aged 12-14 years), some of whom had symptoms of asthma and bronchial hyper-reactivity. Antibodies to mite (Der f 1) and cat (Fel d 1) allergens measured by isotype (IgG and IgG4) specific radioimmunoprecipitation assays were compared with sensitisation and allergen concentrations in house dust. FINDINGS 226 children were recruited, 47 of whom had symptoms of asthma and bronchial hyper-reactivity. Increasing exposure to mite was associated with increased prevalence of sensitisation and IgG antibody to Der f 1. By contrast, the highest exposure to cat was associated with decreased sensitisation, but a higher prevalence of IgG antibody to Fel d 1. Thus, among children with high exposure, the odds of sensitisation to mite rather than cat was 4.0 (99% CI 1.49-10.00). Furthermore, 31 of 76 children with 23 microg Fel d 1 at home, who were not sensitised to cat allergen had >125 units of IgG antibody to Fel d 1. Antibodies to Fel d 1 of the IgG4 isotype were strongly correlated with IgG antibody in both allergic and non-allergic children (r=0.84 and r=0.66, respectively). Sensitisation to mite or cat allergens was the strongest independent risk factor for asthma (p<0.001). INTERPRETATION Exposure to cat allergen can produce an IgG and IgG4 antibody response without sensitisation or risk of asthma. This modified T-helper-2 cell response should be regarded as a form of tolerance and may be the correct objective of immunotherapy. The results may also explain the observation that animals in the house can decrease the risk of asthma.

Exhaled breath condensate pH is a robust and reproducible assay of airway acidity

Exhaled breath condensate (EBC) pH is low in several lung diseases and it normalises with therapy. The current study examined factors relevant to EBC pH monitoring. Intraday and intraweek variability were studied in 76 subjects. The pH of EBC collected orally and from isolated lower airways was compared in an additional 32 subjects. Effects of ventilatory pattern (hyperventilation/hypoventilation), airway obstruction after methacholine, temperature (−44 to +13°C) and duration of collection (2–7 min), and duration of sample storage (up to 2 yrs) were examined. All samples were collected with a disposable condensing device, and de-aerated with argon until pH measurement stabilised. Mean EBC pH (n=76 subjects, total samples=741) was 7.7±0.49 (mean±sd). Mean intraweek and intraday coefficients of variation were 4.5% and 3.5%. Control of EBC pH appears to be at the level of the lower airway. Temperature of collection, duration of collection and storage, acute airway obstruction, subject age, saliva pH, and profound hyperventilation and hypoventilation had no effect on EBC pH. The current authors conclude that in health, exhaled breath condensate pH is slightly alkaline, held in a narrow range, and is controlled by lower airway source fluid. Measurement of exhaled breath condensate pH is a simple, robust, reproducible and relevant marker of disease.

Quantitative measurement of airborne allergens from dust mites, dogs, and cats using an ion-charging device.

Background Increasing evidence suggests that children raised with an animal(s) in the house have a decreased risk of becoming sensitized. However, it is not clear whether this phenomenon is related to airborne exposure.

Exhaled breath condensate pH assays are not influenced by oral ammonia.

Background: Measurement of pH in exhaled breath condensate (EBC) is robust and simple. Acidic source fluid (airway lining fluid) traps bases while volatilising acids, leading to EBC acidification in many lung diseases. Lower airway ammonia is one determinant of airway lining fluid pH, raising the concern that addition of the base ammonia by contamination from the mouth might confound EBC pH assays. Methods: Three discrete methods were used to limit oral ammonia contamination of EBC collections: endotracheal intubation, oral rinsing, and −40°C condenser temperatures. Separately, ammonia was removed from collected EBC samples by lyophilisation and resuspension. Intraweek and intraday variability of ammonia concentration was determined in 76 subjects, and ammonia and pH from a further 235 samples were graphically compared. Ammonia was assayed spectrophotometrically and pH was assessed after deaeration. Results: Data from 1091 samples are presented. Ammonia was reduced in EBC by all methods. Endotracheal intubation decreased EBC ammonia from a mean (SD) of 619 (124) μM to 80 (24) μM (p<0.001, n = 32). Oral rinsing before collection also led to a decline in EBC ammonia from 573 (307) μM to 224 (80) μM (p = 0.016, n = 7). The colder the condensation temperature used, the less ammonia was trapped in the EBC. Lyophilisation removed 99.4 (1.9)% of ammonia. Most importantly, the pH of EBC never decreased after removal of ammonia by any of these methods. Intraweek and intraday coefficients of variation for ammonia were 64 (27)% and 60 (32)%, which is substantially more variable than EBC pH assays. Conclusions: Although ammonia and pH appear to correlate in EBC, the oral ammonia concentration is not an important determinant of EBC pH. No precautions need to be taken to exclude oral ammonia when EBC pH is of interest. The low pH and low ammonia found in EBC from patients with lung diseases appear to be independent effects of volatile compounds arising from the airway.

Serum IgG and IgG4 Antibodies to Fel d 1 among Children Exposed to 20 μg Fel d 1 at Home: Relevance of a Nonallergic Modified Th2 Response

Exposure to foreign antigens is an essential element of all immune responses, including allergic sensitization. For some allergens (e.g. mite and cockroach), the prevalence of sensitization is directly correlated with exposure. However, for allergens derived from domestic animals, several studies have suggested that children with a cat in the home have a decreased risk of sensitization and asthma. We have now shown that many children exposed to greater than 20 µg of Fel d 1/g of dust at home made an IgG and IgG4 antibody response to Fel d 1 without IgE antibody. This modified Th2 response is not associated with symptoms and should be regarded as a form of immunological tolerance. The fact that the dose-response relationship between cat exposure and sensitization is bell shaped, while that for mite exposure and sensitization is linear, is highly relevant to understanding the role of allergens in the increase in allergic disease.

Evaluation of materials used for bedding encasement : Effect of pore size in blocking cat and dust mite allergen

BACKGROUND Mattress and pillow encasings are recommended for patients allergic to dust mites. Many encasements block allergen and are vapor permeable but do not allow free passage of air through the material. Recently, breathable fabrics made from tightly woven synthetic fibers or nonwoven synthetics have been recommend as encasements. OBJECTIVE The purpose of this study was to develop a method for testing encasement materials made of breathable fabrics. METHODS Dust samples containing a known quantity of allergen (Der f 1, Der p 1, and Fel d 1) were pulled across a variety of fabrics using a modified dust trap. Airflow through the dust trap was controlled with a vacuum pump. Five minutes after dust was introduced, the pump was shut off. A filter located downstream of the fabric collected allergen passing through the fabric during the test and was assayed with ELISA for the relevant allergen. Fabrics to be tested were obtained from manufacturers and specialty catalogs. RESULTS As the average pore size decreases, the airflow through a fabric becomes restricted, and the pressure differential created by the vacuum pump increases. Dust mite allergens (Der f 1 and Der p 1) were blocked below detectable limits by fabrics of less than 10 microm in pore size. Fabrics with an average pore size of 6 microm or less blocked cat allergen (Fel d 1). CONCLUSION The method we developed provided a rigorous and reliable test for leakage of common indoor allergens through breathable barrier fabrics. Our results show that tightly woven fabrics and nonwoven synthetic fabrics can block common indoor allergens but still allow airflow.

Assessment of vacuum cleaners and vacuum cleaner bags recommended for allergic subjects.

BACKGROUND High-quality vacuum cleaners and vacuum cleaner bags are often recommended to allergic patients as a means of reducing indoor allergen exposure. A number of vacuum cleaners on the market today claim to capture 99.9% of particles 0.3 microm or larger entering the vacuum cleaner, and many vacuum cleaner bags are now being sold as microfiltration bags. OBJECTIVE The purpose of this study was to compare the allergen-trapping abilities of vacuum cleaners and to use a new technique for testing vacuum cleaner bags that are recommended for allergic patients. METHODS Vacuum cleaners were tested in an 18-m(3) laboratory room permeated with dust containing high levels of cat allergen by using techniques previously described. Air was sampled with parallel filters in conjunction with a particle counter. The filters were assayed by ELISA for cat allergen (Fel d 1). Vacuum cleaner bags were tested by using a modified dust trap to pull sieved house dust containing a known amount of Fel d 1 across the material used for the bag. Allergen passing through the bag was trapped on a filter covering the exit of the trap and analyzed for Fel d 1. RESULTS In general, vacuum cleaners designed for allergic patients leaked lower amounts of allergen (<0.5-4.04 ng/m(3)) than that found in our previous studies (<0.5-100 ng/m(3)). Single-layer vacuum cleaner bags performed poorly (1250-2640 ng recovered) compared with most of the 2- and 3-layer microfiltration bags (0.53-2450 ng recovered). The range of allergen recovered from the 2-layer bags (0.93-2450 ng recovered) highlighted the variability found between manufacturers. CONCLUSION The results suggest that although allergen leakage has been reduced, there is still room for improvement. A method of testing allergen leakage by using Fel d 1 should be applied to vacuum cleaners and bags recommended for allergic patients.

Decreased prevalence of asthma among children with high exposure to cat allergen: relevance of the modified Th2 response.

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New approaches to environmental control

ConclusionAllergic disease is characterized by inflammation of the respiratory tract (nose or lung) or the skin. Exposure to allergens such as dust mite is the best-established cause of this inflammation (1,42). Thus, controlling exposure to indoor allergens is the logical first line of treatment for these diseases. However, the methods that should be recommended for avoidance are dependent on understanding how allergens accumulate in the home as well as identifying the specific sensitivity of the patient. In addition, the new materials and new methods proposed need careful evaluation. There are three important developments: new breathable materials for covering beds and pillows, vacuum cleaners designed for allergic patients, and techniques for reducing exposure to animal dander with the animal remaining in the house.

Consumption of S-nitrosoglutathione by polymorphonuclear leukocyte lysates

John F Hunt, Elizabeth Erwin, John W Vaughan, Benjamin M Gaston University of Virginia, Charlottesville, VA S-nitrosoglutathione (GSNO) is an endogenous bronchodilator depleted in the severe asthmatic airway. Eosinophils and neutrophils are found in elevated levels in bronchoalveolar lavage and biopsies of asthmatic lung, and exhaled nitric oxide (NO) levels are high. We hypothesized that polymorphonuclear leukocytes possess a catabolic enzyme for GSNO that releases NO. In this study, we isolated human peripheral blood eosinophils and neutrophils by density gradient and subsequent magnetic separation and subjected the cells to detergent lysis and homogenization in protease inhibitors in cold conditions. Homogenized cell lysates were suspended in 0.5% fetal bovine serum with 2 mM EDTA. Fifty micromolar GSNO was added to the cell homogenates which then were assayed over time for residual GSNO by gas-phase chemiluminescent detection after reduction to NO in Cu +1 -cysteine. Both eosinophil and neutrophil homogenates rapidly consumed GSNO compared to control (0.5% FBS + EDTA), with 33-50% comparative reduction in GSNO by 6 minutes, but limited difference from control in rate of consumption thereafter. In contrast, there were no differences from control in GSNO consumption by cell lysates after allowing them to degrade in a 37 degree water bath for 2 hours prior to addition of GSNO. These initial results suggest there is a GSNO consumptive pathway present in peripheral blood polymorphonuclear cell lysates. This pathway can be degraded with time, likely through proteolysis (if enzymatic) or elimination of NO-reactive species such as superoxide. If enzymatic, then consumption of a necessary cofactor could explain the cessation of effect after 6 minutes. This work supported by AAAAI Education and Research Trust Faculty Development Award, GlaxoSmithKline, Virginia Thoracic Society, NIH. JH is a Parker B. Francis Fellow in Pulmonary Medicine. 991 sisRelati°nshiPof Neuroblastoma°f TRK-A Statement to Clinical Stage and Progno-