Pteronyssinus Allergen Research Articles

Effect of mite allergen immunotherapy on the altered phenotype of dendritic cells in allergic asthmatic children

Allergic asthma is a T(H)2 inflammatory disease. Dendritic cells (DCs) play key roles in the T(H)1/T(H)2 balance. Allergen specific immunotherapy (SIT) has the potential to modify the course of allergy because the ratio of T(H)1 to T(H)2 cytokines produced is increased after SIT. To determine how SIT affects DCs in children and to define novel parameters of this treatment. We investigated the changes of phenotypic and functional variations of monocyte-derived DCs fromallergic asthmatic children undergoing complete mite SIT. Peripheral blood monocytes from SIT allergic asthmatic children, allergic asthmatic controls, and healthy controls were cultured with granulocyte-macrophage colony-stimulating factor and interleukin 4 and then stimulated with Dermatophagoides pteronyssinus (Der p) allergen or lipopolysaccharide (LPS). The expressions of surface molecules on monocyte-derived DCs were assessed by flow cytometry. Cytokine production by cultured monocyte-derived DCs was determined by enzyme-linked immunosorbent assay. After LPS stimulation, monocyte-derived DCs of the allergic asthmatic group had a higher CD86 and lower HLA-DR expression than the healthy controls. In SIT patients, the expression was similar to that of the healthy controls. After Der p stimulation monocyte-derived DCs of the allergic asthmatic patients displayed lower Toll-like receptor 4 (TLR4), whereas again in SIT patients the expression was similar to that of healthy controls. These findings indicate that SIT normalizes the expression of CD86, HLA-DR, and TLR4 on DCs. Moreover, CD86, HLA-DR, and TLR4 may be useful parameters for monitoring SIT. Decreased TLR4 expression in allergic asthmatic patients might be compensated by TLR4 agonists, with the potential of amplifying the effects of SIT.

Varying Allergen Composition and Content Affects the in vivo Allergenic Activity of Commercial Dermatophagoides pteronyssinus Extracts

Background: Diagnosis and immunotherapy of house-dust mite (HDM) allergy is still based on natural allergen extracts. The aim of this study was to analyze commercially available Dermatophagoides pteronyssinus extracts from different manufacturers regarding allergen composition and content and whether variations may affect their allergenic activity. Methods: Antibodies specific for several D. pteronyssinus allergens (Der p 1, 2, 5, 7, 10 and 21) were used to analyze extracts from 10 different manufacturers by immunoblotting. Sandwich ELISAs were used to quantify Der p 1 and Der p 2 in the extracts. Mite-allergic patients (n = 45) were skin-tested with the extracts and tested for immunoglobulin E (IgE) reactivity to a panel of 10 mite allergens (Der p 1, 2, 4, 5, 7, 8, 10, 14, 20 and 21) by dot blot. Results: Only Der p 1 and Der p 2 were detected in all extracts but their concentrations and ratios showed high variability (Der p 1: 6.0–40.8 µg ml^–1; Der p 2: 1.7–45.0 µg ml^–1). At least 1 out of 4 allergens (i.e. Der p 5, 7, 10 and 21) was not detected in 8 of the studied extracts. Mite-allergic subjects showed different IgE reactivity profiles to the individual mite allergens, the extracts showed different allergenic activity in skin-prick tests and false-negative results. Conclusions: Commercially available D. pteronyssinus extracts lack important allergens, show great variability regarding allergen composition and content and some gave false-negative diagnostic test results in certain patients.

Carrier‐bound nonallergenic Der p 2 peptides induce IgG antibodies blocking allergen‐induced basophil activation in allergic patients

More than 90% of house dust mite-allergic patients are sensitized to the major Dermatophagoides pteronyssinus allergen, Der p 2. The aim of this study was to develop and characterize an allergy vaccine based on carrier-bound Der p 2 peptides, which should allow reducing IgE- and T-cell-mediated side-effects during specific immunotherapy (SIT). Five Der p 2 peptides (P1-P5) were synthesized and analyzed regarding IgE reactivity and allergenic activity. Lymphoproliferative and cytokine responses induced with Der p 2 and Der p 2 peptides were determined in peripheral blood mononuclear cells from mite-allergic patients. Der p 2-specific IgG antibodies induced with carrier-bound Der p 2 peptides in mice and rabbits were tested for their capacity to inhibit IgE binding and basophil activation in allergic patients. Of five overlapping peptides (P1-P5) covering the Der p 2 sequence, two peptides (P2 and P4) were identified, which showed no relevant IgE reactivity, allergenic activity, and induced lower Der p 2-specific T-cell activation than Der p 2. However, when coupled to a carrier, P2 and P4 induced Der p 2-specific IgG antibodies in animals, which inhibited allergic patients' IgE binding to the allergen and allergen-induced basophil activation similar as antibodies induced with Der p 2. Carrier-bound Der p 2 peptides should allow avoiding IgE-mediated side-effects, and because of their low potential to activate allergen-specific T cells, they may reduce late-phase side-effects during SIT. Further, these peptides may be also useful for prophylactic vaccination.

201 Clinical Efficacy and Mucosal/Systemic Antibody Response Changes After Sublingual Immunotherapy in Mite-Allergic Children

BackgroundThis study aimed to evaluate the clinical efficacy and mucosal/systemic antibody response changes after sublingual immunotherapy (SLIT) using Dermatophagoides pteronyssinus (Dpt) allergens with or without bacterial extracts in mite-allergic Brazilian children.MethodsOne-hundred and 2 patients presenting allergic rhinitis with or without asthma were selected for a randomized double-blind, placebo-controlled trial and distributed into 3 groups: DPT (Dpt allergen extract, n = 34), DPT + MRB (Dpt allergen plus mixed respiratory bacterial extracts, n = 36), and Placebo (n = 32). Clinical evaluation and immunological analyses were carried out before and after 12 and 18 months of treatment, including rhinitis/asthma symptom and medication scores, skin prick test (SPT) to Dpt extract, and measurements of Dpt-, Der p 1-, Der p 2-specific IgE, IgG4, and IgG1 in serum and -specific IgA in saliva and nasal lavage fluid.ResultsClinical results showed a significant decline in rhinitis/asthma symptom scores in all groups, but medication use decreased only in active DPT group at 12 months. SPT results showed no significant changes and SLIT was generally safe, with no severe systemic reactions. SLIT using Dpt allergen alone induced increased serum IgG4 levels to Dpt, Der p 1 and Der p 2, and increased serum IgG1 and salivary IgA levels to Dpt and Der p 1. SLIT using DPT+MRB was able to decrease IgE levels, particularly to Der p 2, to increase salivary IgA levels to Der p 1, but had no changes on specific IgG4 and IgG1 levels.ConclusionsTherefore, SLIT seems to be effective in ameliorating clinical symptoms, but only active SLIT was able to modulate the mucosal and systemic antibody responses. These findings support the role of specific serum IgG4 and IgG1, in addition to salivary IgA, as protective or blocking antibodies as well as biomarkers of tolerance that may be useful for monitoring activation of tolerance-inducing mechanisms during allergen immunotherapy.

Increased intake of oily fish in pregnancy: effects on neonatal immune responses and on clinical outcomes in infants at 6 mo

Long-chain n-3 PUFAs found in oily fish may have a role in lowering the risk of allergic disease. The objective was to assess whether an increased intake of oily fish in pregnancy modifies neonatal immune responses and early markers of atopy. Women (n = 123) were randomly assigned to continue their habitual diet, which was low in oily fish, or to consume 2 portions of salmon per week (providing 3.45 g EPA plus DHA) from 20 wk gestation until delivery. In umbilical cord blood samples (n = 101), we measured n-3 fatty acids, IgE concentrations, and immunologic responses. Infants were clinically evaluated at age 6 mo (n = 86). Cord blood mononuclear cell (CBMC) production of interleukin (IL)-2, IL-4, IL-5, IL-10, and tumor necrosis factor-α in response to phytohemagglutinin (PHA) and of IL-2 in response to Dermatophagoides pteronyssinus allergen 1 (Derp1) was lower in the salmon group (all P ≤ 0.03). In the subgroup of CBMCs in which an allergic phenotype was confirmed in the mother or father, IL-10 production in response to Toll-like receptor 2, 3, and 4 agonists, ovalbumin, salmon parvalbumin, or Derp1 and prostaglandin E(2) production in response to lipopolysaccharide or PHA was lower in the salmon group (all P ≤ 0.045). Total IgE at birth and total IgE, incidence and severity of atopic dermatitis, and skin-prick-test positivity at 6 mo of age were not different between the 2 groups. Oily fish intervention in pregnancy modifies neonatal immune responses but may not affect markers of infant atopy assessed at 6 mo of age. This trial is registered at clinicaltrials.gov as NCT00801502.

Modulation of the Humoral Response to Dermatophagoides pteronyssinus Allergens in BALB/c Mice by Extract Modification and Adjuvant Use

Background: Currently, several strategies are being used in order to improve the safety and efficacy of allergen-specific immunotherapy; these strategies include the use of modified hypoallergenic extracts as well as different adjuvants with immunomodulatory properties in combination with native or modified extracts. The objectives of this study were to investigate the humoral response generated in mice to modified Dermatophagoides pteronyssinus extracts in the presence or absence of two different adjuvants. Methods: BALB/c mice were inoculated either with native, depigmented or depigmented-polymerised D. pteronyssinus without adjuvants or combined with aluminium hydroxide or oligodeoxinucleotides containing CpG motifs. IgE concentration, specific total IgG, IgG1 and IgG2a titres were measured in mice sera and cross-reactivity inhibition experiments were performed. IgG antigenic profiles were obtained by immunoblotting for all formulations. Results: Inoculation of depigmented-polymerised extract induced statistically significant lower IgE levels than the native extract even when adsorbed onto aluminium hydroxide. When this extract was inoculated in the presence of oligodeoxinucleotides containing CpG motifs, it elicited high IgG levels, a high IgG2a/lgG1 ratio and low IgE production. Furthermore, the antigenic profiles observed after extract inoculation showed punctual differences between the depigmented-polymerised extract and the native or depigmented extracts. Conclusions: Our results suggest that the depigmentation and polymerisation process modifies the native extract’s antigenic and immunogenic properties and converts the depigmented-polymerised extract into a better choice for allergen-specific immunotherapy.

Association of house dust mite-specific IgE with asthma control, medications and household pets

BackgroundEvidence is conflicting regarding the effectiveness of creating a low-allergen environment or reducing allergen exposure to control asthma exacerbations.ObjectiveThis study determined the association of house dust mite (HDM)-specific IgE levels with asthma symptom control, selected medications, family history of allergic disease, and exposure to second-hand smoke and household pets.MethodsSerum samples from 102 doctor-diagnosed allergic asthma patients and 100 non-atopic controls were subjected to enzyme-linked immunosorbent assay using the HDM species Dermatophagoides pteronyssinus (Dp), Dermatophagoides farinae (Df), and Blomia tropicalis (Bt) allergens. Point-biserial correlation coefficient, Pearson R correlation, and logistic regression analyses were used to determine association of HDM-specific IgE levels with the abovementioned variables.ResultsOf the 102 cases, 38.24%, 47.06%, and 33.33% were sensitized to Bt, Df, and Dp, respectively. Sensitized patients showed greater probability [Bt (OR = 1.21), Df (OR = 1.14), and Dp (OR = 1.35)] to manifest symptoms than those who were not. Obtained p-values [Bt (p = 0.73), Df (p = 0.83), and Dp (p = 0.59)], however, proved that HDM-specific IgE levels had no significant contribution in predicting or explaining occurrence of asthma symptoms. Bt- and Df-specific IgEs showed moderately weak but significant relationship with bambuterol HCl and expectorant, respectively. Patients currently on said medications registered higher HDM-specific IgE levels than those who were not. No significant correlation between IgE levels and family history of allergic disease or with exposure to second-hand smoke was seen. Dp-specific IgE levels of patients exposed to household pets were significantly lower compared to those without exposure.ConclusionThis study proves that sensitization to Bt, Df, and Dp allergens is not significantly associated with asthma symptoms and control. Although cases were shown to be sensitized to HDMs, their current medications were at least effective in controlling their asthma symptoms.

Transforming Growth Factor Beta: A Role in the Upper Airway and Rhinosinusitis—Dermatophagoides Pteronyssinus–Induced Apoptosis with Pulmonary Alveolar Cells

There is a link with the upper and lower airway and disruption of alveolar epithelial cells, which is a potential trigger for the reactivation of the epithelial-mesenchymal trophic unit (EMTU) and induced characteristic airway changes associated with allergic asthma. Dermatophagoides pteronyssinus is a common inhalant indoor allergen and is known for causing allergic rhinitis and airway inflammation. Transforming growth factor beta 1 (TGF-beta1) is a major participant in the airway remodeling of asthma, a component of cellular stress response pathways, and enhanced epithelial immunoreactivity is known to occur in allergic rhinitis. In this study, we show the ability of D. pteronyssinus allergens from dialyzed standardized immunotherapy extract to induce apoptosis and increase TGF-beta1 secretion in a confluent A549 cell line model. A549 cells were treated with either 600 AU/mL dialyzed D. pteronyssinus immunotherapy extract (eDp) or Ctl media (Ctl) for 24 hours. Cells and supernatants were collected, washed, and treated with Annexin V-FITC Apoptosis Detection Kit II (BD Pharmingen, La Jolla, CA) and then analyzed by flow cytometry. TGF-beta1 secretion was determined by ELISA using cell culture supernatants. The eDp group showed a fourfold increase in early apoptotic cells with a twofold increase in late apoptotic cells versus the Ctl group, along with a 1.65-fold increase of TGF-beta1. eDp induced viable A549 cells to undergo apoptosis determined by flow cytometry analysis with a significant increase in TGF-beta1 secretion compared with Ctl.

Mite component–specific IgE repertoire and phenotypes of allergic disease in childhood: The tropical perspective

Sensitization to perennial aeroallergens correlates with the risk of persistent asthma (AS) in children. In tropical Singapore, multiple codominant species of mites abound in the indoor environment, and preferential species-specific sensitization has been associated with different phenotypes of allergic disease. We investigated the pattern of mite component-specific IgE (mcsIgE) in children with different phenotypes of clinical allergic disease in an environment with multiple mite species exposure. A prospective evaluation of newly diagnosed patients with clinical diagnosis of allergic rhinitis (AR), atopic dermatitis (AD), or AS and sensitization to one or more aeroallergens were performed. Sera were tested for specific IgE against an extensive panel of Dermatophagoides pteronyssinus and Blomia tropicalis allergens. A total of 253 children were included, mean age 7.3 yr, 79% fulfilled criteria for AR, 46% AS, 71% AD, and 31% for all three. Sensitization to one or both mites was observed in 91% of children, 89% were sensitized to D. pteronyssinus, and 70% to B. tropicalis. The most common mite allergens recognized by these atopic children were Der p 1 (64%), Der p 2 (71%), Blo t 5 (45%), Blo t 7 (44%), and Blo t 21 (56%). Specific IgE responses to an increased number of distinct mite allergens correlated with the complexity of the allergic phenotype. In multivariate analysis, an increased risk for the multi-systemic phenotype (AR + AS + AD) was associated with sensitization to an increased repertoire of mite components (three or more) (OR 4.3, 95% CI 2.1-8.8, p = 0.001) and a positive parental history of AS (OR 2.4, 95% CI 1.2-2.9, p = 0.013). A highly pleiomorphic IgE response to the prevalent indoor mites is associated with the presence of a multi-systemic allergic phenotype in childhood in a tropical environment.

Efficacy and Safety of a Glutaraldehyde-Modified House Dust Mite Extract in Allergic Rhinitis

Modification of allergens by glutaraldehyde in extracts used for immunotherapy reduces the risk for side effects, but therapeutic efficacy of such extracts requires further evaluation. The aim of this study was to evaluate the efficacy and safety of immunotherapy with PURETHAL Mites (PM), a single-strength glutaraldehyde-modified aluminum hydroxide-adsorbed extract of house-dust mites (HDM). In a multicenter, randomized, placebo-controlled double-blind setting, HDM-allergic subjects (n = 140) were treated with modified allergen extract or placebo over a 1-year period. The primary outcome parameter was a combined symptom and medication score (clinical index score [CIS]). Secondary efficacy parameters were the result of a titrated conjunctival provocation test (CPT), rhinitis/rhinoconjunctivitis quality of life (RQL) score, and serum concentrations of IgE and IgG against specific HDM allergens and a documentation of adverse events (AE). We evaluated 140 patients (66 treatment and 74 placebo) for clinical efficacy. The allergoid treatment for 1 year resulted in significantly greater CIS improvement and higher RQL scores. The response threshold in the titrated CPT (p = 0.009) and the serum concentrations of IgG4 (p < 0.001) against Dermatophagoides pteronyssinus allergens after treatment were also significantly different between groups. In total, 88 patients (46 PM/42 placebo) out of a safety population of 145 reported 278 (158 PM/120 placebo) AE. Except for local reactions, no specific AE appeared to be associated with PURETHAL Mites (HAL-Allergy, Leiden, The Netherlands). The findings of this study indicate that allergen injection therapy with modified HDM extract is superior to placebo in allergic rhinitis therapy. The treatment was well tolerated and no serious drug-related AE were observed.

Indoor allergen levels in Guangzhou city, southern China

High levels of sensitization to house dust mites have been observed in Chinese allergic patients. This study has measured levels and distributions of mite and cockroach allergens in household dust in Guangzhou. Influences of home characteristics and seasonal changes on allergen levels were also investigated. Dust samples were collected from bedding and living room from households in Guangzhou. Major allergens from Dermatophagoides pteronyssinus, D. farinae, D. microceras, Blomia tropicalis and cockroach allergens were measured by ELISA. Home characteristics were obtained from a questionnaire. Four hundred and four dust samples were collected from 107 homes during October 2006 to November 2007. House dust mite allergen levels were detectable in 99% of the bedding samples. Der f 1 levels were significantly higher than Der p 1 levels. High levels of mite allergens (>10 μg/g) were observed in 88% of all the bedding samples. Cockroach allergens were detected in 93% of households and were higher in living room samples than in bedding samples. Blo t 5 and Der m 1 could not be detected in the dust samples. Having fabric furniture was a predictor of high allergen levels. Der f 1 levels were higher in summer time than in winter time. Cockroach allergens were higher in winter time than in summer time. In Guangzhou, Der f 1 is the predominant mite allergen in dust with very high levels in bedding. Cockroach allergens are also common.

Exposure to poultry dust and health effects in poultry workers: impact of mould and mite allergens

The aim of the study was to evaluate exposure to moulds and house dust mite Dermatophagoides pteronyssinus in poultry farms, and related health effects in poultry workers (PW). The study involved 41 PW and 45 control office workers. Working environment was evaluated for D. pteronyssinus allergen (Der p 1), moulds and endotoxin. In workers, eye, skin and respiratory symptoms, ventilatory lung function, atopy markers (skin prick test to inhalatory allergens, total IgE) and specific IgG to moulds were assessed. Der p 1 levels ranged <0.1-3.3 microg/g, exposure to fungi was 4.9 x 10(3)-6.8 x 10(4) cfu/m(3), with prevailing Aspergillus, Penicillium and Mucor species, and endotoxin levels ranged 230-284 EU/m(3). In comparison to control subjects, significantly higher prevalence of work-related nose, asthma, eye and skin symptoms, and slight decline in ventilatory lung function was found in PW. PW had significantly higher prevalence of IgG antibodies to moulds comparing to controls (63 vs. 36%, respectively, P = 0.01), especially to Alternaria and Aspergillus species. The prevalence of atopy markers in PW was lower than in population-based studies. Hazardous levels of Der p 1, endotoxin and moulds were determined in poultry houses. High prevalence of work-related symptoms and IgG antibodies to moulds was found in PW. Healthy worker effect is proposed as an explanation of low atopy markers prevalence among PW.

Skin Sensitization and Classroom Exposure to Dermatophagoides Pteronyssinus and Dermatophagoides Farinae Allergens in Andean Ecuadorian Students

Skin Sensitization and Classroom Exposure to Dermatophagoides Pteronyssinus and Dermatophagoides Farinae Allergens in Andean Ecuadorian Students

Interference of Dermatophagoides' Specific Immunoglobulins G in the Quantification of Mite's Specific Immunoglobulins E

In order to investigate the interference of specific IgG in the quantification of specific IgE, using the ImmunoCAP 250® system, we studied, in parallel, the interference by total adsorption of interferent and the classical method of adding interferent increasingly. Furthermore, to evaluate if the interference is affected by different solid phases, total extract of Dermatophagoides pteronyssinus and recombinant allergens of Dermatophagoides farinae were used. The results showed a statistical significant interference by IgG in the quantification of the specific IgE, but neither analytical nor clinical significant interference were observed. Therefore, this analytical system provides an accurate method for determination of the specific IgE concentration contributing to the allergic disease diagnosis quality.

Engineering of major house dust mite allergens Der p 1 and Der p 2 for allergen‐specific immunotherapy

Specifically designed recombinant allergens with reduced IgE reactivity are promising candidates for a more defined, effective, and safer specific immunotherapy (SIT). We sought to obtain hypoallergenic hybrid molecules which could potentially be applied to house dust mite (HDM) allergy treatment. Two hybrid molecules (QM1 and QM2) derived from the two major Dermatophagoides pteronyssinus allergens, Der p 1 and Der p 2, were engineered by PCR, produced in Escherichia coli, and purified. The overall IgE-binding capacity of the hybrids was compared with their single components by Western blot, specific IgE, skin prick test (SPT), and IgE-inhibition assays. T cell proliferation assay were performed to confirm their retention of T cell reactivity. Immune responses to the hybrid molecules were studied in BALB/c mice. The IgE reactivity of both hybrid proteins was strongly reduced as evaluated by in vitro methods. Furthermore, in vivo SPTs performed on 106 HDM-allergic patients showed that the hybrid proteins had a significantly lower potency to induce cutaneous reactions than the individual components. Hybrid molecules induced higher T cell proliferation responses than those produced by an equimolecular mixture of Der p 1 and Der p 2. Immunization of mice with the hybrid proteins induced Der p 1- and Der p 2-specific IgG, which inhibited the binding of allergic patients' IgE to these natural allergens. QM1 and QM2 hybrids exhibited less IgE-binding activity but preserved immunogenicity and fulfilled the basic requirements for hypoallergenic molecules suitable for a future SIT of HDM allergy.

Comparison and Evaluation of Several Dermatophagoides Pteronyssinus Allergen Extracts for Skin Prick Test

To evaluate the significance of several Dermatophagoides pteronyssinus allergen extracts for skin prick test (SPT) in patients allergic to Dermatophagoides pteronyssinus. Two hundred and nineteen patients enrolled in Peking Union Medical College Hospital underwent SPT and serum specific IgE assay to detect the Dermatophagoides pteronyssinus allergen. Three kinds of house dust mite allergen extracts were used for SPT, including the Dermatophagoides pteronyssinus extract prepared by our laboratory (group A), standardized Dermatophagoides pteronyssinus extract (group B), and mixed extracts of Dermatophagoides pteronyssinus and Dermatophagoides farinae (group C). Human serum specific IgE result was regarded as the reference standard for diagnosis of Dermatophagoides pteronyssinus allergy. The receiver operating characteristic (ROC) curve was used to evaluate the diagnostic performance of SPT with the extracts of three groups. SPT results showed that the median wheal diameter of group A, group B, and group C was 0.43, 0.35, and 0.28 cm, respectively, with significant difference among three groups (P<0.05). The difference was significant between group A and B (P<0.01) as well as group A and C (P<0.01), but not between group B and C (P>0.05). There was no local urticaria or systemic allergic reactions following the procedure of SPT. Local reaction was observed in 5 patients and delayed reaction was in 2 patients of group A. As for group B and C, local reaction occurred in 3 cases and delayed reaction in 2 cases in each group. The area under ROC curve of SPT with extract in group A, group B, and group C was 0.765, 0.801, and 0.782, respectively. Based on the detection results of serum specific IgE, the sensitivity of SPT in diagnosis of Dermatophagoides pteronyssinus allergy with extract of group A, group B, and group C was 92.4%, 87.0%, and 81.5%, and the specificity was 60.6%, 73.2%, and 74.8%, respectively. The Dermatophagoides pteronyssinus extract for SPT prepared by our laboratory offers good sensitivity and specificity comparable to commercially available allergen extracts, and it may be an appropriate candidate for clinical screening and diagnosis of Dermatophagoides pteronyssinus allergy.

Comparison of Three Methods of Protein Extraction from Dermatophagoides Pteronyssinus for Two-dimensional Electrophoresis

To explore an effective method of Dermatophagoides pteronyssinus protein extraction suitable for two-dimensional electrophoresis (2-DE) analysis. The extracts of Dermatophagoides pteronyssinus were prepared with Coca's solution, lysis buffer of 2-DE, and Trizol reagent, respectively. Bicinchoninic acid (BCA) assay was used to determine the total protein concentration of the samples. The efficiency of different protein extraction methods were evaluated with 2-DE analysis. The concentrations of extracted protein by methods of Coca's solution, lysis buffer, and Trizol reagent were 0.63 g/L, 0.90 g/L, and 0.80 g/L, respectively. The 2-DE analysis results showed that some protein spots in low molecular weight (LMW) range could be detected with the Coca's solution method. With the lysis buffer of 2-DE method, more protein spots in LMW range could be detected, while the medium molecular weight (MMW) protein spots were absent. Several MMW protein spots (174-178 kD and 133 kD) and more LMW protein spots were detected with Trizol reagent method. Among Coca's solution, lysis buffer of 2-DE, and Trizol reagent, the concentration of extracted protein of Dermatophagoides pteronyssinus by lysis buffer of 2-DE is the highest. However, most protein components of Dermatophagoides pteronyssinus purified mite bodies can be extracted by Trizol reagent, which may generally reflect the whole profile of Dermatophagoides pteronyssinus allergens.

Compatibility of imported fire ant whole body extract with cat, ragweed, Dermatophagoides pteronyssinus, and timothy grass allergens

Recommendations regarding the administration of imported fire ant whole body extract (IFA WBE) combined with aeroallergens or environmental allergens in a single immunotherapy injection are lacking. To evaluate the degradative effect of IFA WBE on cat, ragweed, Dermatophagoides pteronyssinus, and timothy grass allergens. Imported fire ant whole body extract was combined with extracts of cat, ragweed, D pteronyssinus, and timothy grass. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was performed on each sample after storage for 0, 1, 3, and 6 months at 4 degrees C. In addition, cat and ragweed combinations were evaluated by radial immunodiffusion (RID); D pteronyssinus by enzyme-linked immunosorbent assay (ELISA) inhibition; and timothy grass by ELISA inhibition and Western blot. Imported fire ant whole body extract combined with timothy grass demonstrated degradation of timothy grass allergens by SDS-PAGE, ELISA inhibition, and Western blot results. Cat and ragweed allergens were stable after mixing with IFA WBE, based on SDS-PAGE and RID analyses. Stability of D pteronyssinus allergens with IFA WBE was evident from SDS-PAGE and ELISA inhibition data. Imported fire ant whole body extract combined with timothy grass resulted in significant and rapid timothy protein degradation. Imported fire ant whole body extract mixed with cat, ragweed, or D pteronyssinus revealed aeroallergen stability, yielding the possibility of combining these extracts in a single immunotherapy injection. Compatibilities of IFA WBE with other common aeroallergens remain undetermined and thus are not recommended for single-injection immunotherapy formulations.

Proteolytically Active Allergens Cause Barrier Breakdown

Two major allergens--the house dust mite Dermatophagoides pteronyssinus (Der p 1) and cockroach allergens--are proteolytically active and stimulate the protease-activated receptor 2 (PAR-2). Jeong et al. (2008, this issue) exposed mouse and human epidermis to both allergens and correlated the observed delay in permeability barrier recovery to PAR-2 activation/signaling. This article exposes the secretive boundaries between barrier homeostasis and immunity.

Ascaris lumbricoides–Induced Interleukin‐10 Is Not Associated with Atopy in Schoolchildren in a Rural Area of the Tropics

In areas where intestinal helminth infections are endemic, infections by these parasites may protect against skin test-measured reactivity to allergens, and it has been suggested that interleukin (IL)-10 may mediate this effect. This study investigated whether IL-10 and populations of IL-10+ T cells may modulate atopy in children living in an area where intestinal helminth infections are endemic. Ecuadorian schoolchildren from rural communities were assessed for skin test-measured reactivity to Periplaneta americana allergen and Dermatophagoides pteronyssinus allergen. Blood samples were collected from 39 skin test-positive and 41 skin test-negative children, and peripheral-blood leukocytes were cultured in the presence of Ascaris lumbricoides antigen, to measure IL-10 protein and the frequency of T cell populations expressing intracellular IL-10. We also investigated whether these immunological factors affected the association between allergen-specific IgE and skin test-measured reactivity to aeroallergens. There was no evidence of association between the level of A. lumbricoides-induced IL-10 protein or IL-10+ T cells and skin test-measured reactivity to allergens. The association between allergen-specific IgE and skin test-measured reactivity was not affected by the level of IL-10 protein or the frequency of IL-10+ T cells. The results of this study do not support the notion that IL-10 plays a role in modulating atopy in children living in a tropical area where intestinal helminth infections are endemic.