bluuebunny/crossref_metadata_2025 huggingface dataset:
Increased mast cell tryptase in sudden infant death — anaphylaxis, hypoxia or artefact?
Authors: Edston E. , Gidlund E. , Wickman M. , Ribbing H. , Van Hage‐Hamsten M.
Published: March 2003

Background

Increased concentrations of mast cell tryptase in post mortem blood have frequently been observed in sudden infant deaths but the cause of this has not yet been clarified.

Objective

The aim was to evaluate factors (immunological, morphological and anamnestic data) behind the observed increase in mast cell tryptase in sudden infant deaths with elevated tryptase.

Methods

Mast cell tryptase and total immunoglobulin (Ig) E were measured in post mortem sera from 44 infants younger than 1.5 years. Radioallergosorbent tests were performed for possible allergens (mixture for relevant food allergens, Phadiatop and latex). IgG subclasses, IgM, and complement factors (C3, C4 and factor B) were measured with radial immunodiffusion. Mast cells, labelled with antibodies against mast cell tryptase, were counted in the lungs and heart. The circumstances of death and medical history of the deceased infant and family were obtained through police and hospital records.

Results

In 40% of the SIDS cases tryptase was elevated (>10 μg/L). Total IgE in serum was increased in 33% compared with clinical reference values but showed no association with mast cell tryptase. RAST tests were positive in three cases. In one of these cases both tryptase and total IgE were elevated. The only variable that was associated with high tryptase values was prone position at death (P ≤ 0.05 ). Allergy or asthma in the family were alleged in 50% of the cases, but was not associated with elevated tryptase or IgE. Children with elevated total IgE also displayed high concentrations of IgG1 and IgG2. Infants who died in the spring had significantly higher IgE than the others (P ≤ 0.05).

Conclusion

The results do not support the hypothesis that the elevated tryptase concentrations in sudden infant death are caused by allergy. The association between prone position at death and elevated tryptase could hypothetically be explained by mast cell degranulation due to, for example, a hypoxic stimulus in these infants.
Airway inflammation and altered alveolar macrophage phenotype pattern after repeated low‐dose allergen exposure of atopic asthmatic subjects
Authors: Lensmar C. , Prieto J. , Dahlén B. , Eklund A. , Grunewald J. , Roquet A.
Published: March 2003

Background

The alveolar macrophage (AM) constitutes an important link between pulmonary innate and adaptive immunity due to its antigen‐presenting capacity and ability to express different immunomodulating mediators. The role of AMs in the pathogenesis of allergic inflammation has yet to be fully determined.

Objective

To investigate clinical effects and any change in the AM phenotype pattern after inhalation of sub‐clinical doses of allergen by asthmatic patients.

Methods

Eight subjects with allergic asthma underwent repeated low‐dose allergen provocations equivalent to 10% of PD20. AMs recovered with bronchoalveolar lavage (BAL) were characterized by flow cytometric analysis of adhesion molecules, co‐stimulatory molecules and markers for AM population activation and heterogeneity.

Results

An allergic airway inflammation, sub‐clinical in six out of eight subjects, was obtained after low‐dose allergen provocations, as determined by increased airway methacholine reactivity, increased BAL fluid total cell and eosinophil counts and increased serum ECP levels. The AMs showed a post‐challenge altered phenotype pattern with a decreased expression of CD11a, CD16, CD71 and HLA class I and an increased expression of CD11b and CD14. The AMs were positive for CD83 and a weak post‐challenge increase in the CD83 expression was found.

Conclusion

Repeated low‐dose allergen exposure induces an allergic airway inflammation in asthmatic subjects. The inflammation is associated with an altered AM phenotype pattern, consistent with an influx of monocytes and a hypothetical increased accessory cell function in the airways, possibly contributing to the development and sustenance of airway inflammation in asthma.
Global Initiative for Asthma (GINA) and its objectives
Authors: Bousquet
Published: March 2003

Guidelines for the diagnosis and management of asthma have been available for the past 13 years and are updated regularly. However, asthma guidelines are rarely completely up‐to‐date because our knowledge about the pathophysiology and treatment of asthma is continually evolving. Guidelines are an increasingly familiar part of clinical practice and may have potential benefits and harms. The potential harms of guidelines can be minimized if they are rigorously developed using evidence‐based medicine. Guidelines should identify key decisions and their consequences and should review the consequences of alternative decisions. In addition, guidelines should be simple, user‐friendly and widely disseminated. Guidelines evolve over time and their recommendations should be supported by evidence from clinical trials. Evidence‐based medicine is the new paradigm but the results and reaction to the recent meta‐analysis on house‐dust mite control measures highlights the need for care when incorporating such information into guidelines. Newer therapeutic agents such as the leukotriene receptor antagonists are included in the most recent revision of the GINA guidelines, but their position in asthma therapy is not yet fully established. In countries where asthma guidelines have been implemented, there appears to have been a reduction in the prevalence of moderate persistent asthma but no decrease in severe asthma. This indicates that there is still room for improvement.
Influence of bronchial allergen challenge on histamine release by human basophils
Authors: Lie  , Van Der Veen  , Knol  , Mul  , Jansen  , Roos  , Van Der Zee
Published: March 2003

Background

Basophils can be primed by cytokines such as interleukin (IL) ‐3, IL‐5 or granulocyte macrophage‐colony stimulating factor (GM‐CSF). It has been described that the concentrations of these cytokines are enhanced at sites of allergic inflammation as well as systemic in allergic asthma.

Objective

To investigate the priming status of basophils as detected by thapsigargin‐induced histamine release during bronchial allergen challenge.

Methods

Ten subjects allergic to house dust mite were challenged via an aerosol delivery system. Spontaneous leucocyte histamine release as well as histamine release induced by various stimuli was measured in vitro at several time points. In addition, lung function parameters, serum IL‐5 and blood eosinophil counts were evaluated.

Results

We found no effect of bronchial allergen challenge upon spontaneous leucocyte histamine release, nor upon histamine release induced by anti‐immunoglobulin (Ig) E, house dust mite extract, C5a, fMLP, IL‐3, PMA+ thapsigargin or IL‐3+ thapsigargin. However, the priming status of basophils as measured by thapsigargin‐induced histamine release was enhanced at 24 h after bronchial allergen challenge. Analysis of the individual data showed a heterogeneous initial response (30 min, 6 h) followed by a predominant increase at 24 h after allergen challenge. This increase in the thapsigargin‐induced histamine release correlated with the increase in serum IL‐5 levels at 24 h after allergen challenge.

Conclusion

The priming status of human basophils as measured by thapsigargin‐induced histamine release is enhanced 24 h after allergen challenge.
Eosinophilic tracheobronchitis and airway cough hypersensitivity in chronic non‐productive cough
Authors: Fujimura  , Ogawa  , Yasui  , Matsuda
Published: March 2003

Background

We have shown that some patients presenting with chronic bronchodilator‐resistant non‐productive cough have global atopic tendency and airway cough hypersensitivity without non‐specific bronchial hyperresponsiveness, abbreviated as atopic cough. The cough is successfully treated with histamine H1‐antagonists and/or glucocorticoids.

Objective

This prospective study was conducted to elucidate the histological feature of atopic cough.

Methods

Tracheal and bronchial mucosa obtained by transbronchoscopic biopsy and bronchoalveolar lavage (BAL) cell component were studied with special emphasis on eosinophils in eight non‐smokers diagnosed with atopic cough, all of whom had increased sensitivity of cough response to inhaled capsaicin, normal lung function and bronchial responsiveness to methacholine and normal chest roentgenogram. Their cough completely resolved on histamine H1‐antagonists and/or glucocorticoids. Transbronchoscopic tracheal and bronchial biopsy and BAL were also performed in healthy non‐smokers as a control.

Results

A small number of eosinophils was detected in subepithelium of trachea in six of seven patients and in subepithelium of bronchi in seven of eight cough patients. The numbers of eosinophils in subepithelium of trachea and bronchi were significantly increased in the patients compared with control subjects. There was no BAL eosinophilia in any patients.

Conclusion

It is concluded that eosinophilic tracheobronchitis and cough hypersensitivity are pathological and physiological characteristics of atopic cough.
Colophony: an in vitro model for the induction of sensitization
Authors: Elms  , Allan  , Pengelly  , Fishwick  , Beckett  , Curran
Published: March 2003

Background

The potential of colophony fumes from soldering flux to induce asthma has been known since the 1970s, however, no direct in vitro or in vivo evidence has been reported. The present study investigated the potential of colophony to stimulate human phagocytic cells to produce reactive oxygen species.

Methods

The human cell line HL‐60 was differentiated to produce cells with a monocyte‐like and a neutrophil‐like phenotypes. A number of procedures were used to confirm the phenotype of these differentiated cells including morphology, esterase activity, flow cytometry and phagocytosis. The potential of colophony to stimulate human phagocytic cells to produce reactive oxygen species was monitored using flow cytoenzymology.

Results

We were able to show that intracellular peroxide levels were increased in both monocyte‐like and neutrophil‐like cells, but not in undifferentiated HL‐60 cells following the addition of colophony.

Conclusions

The resin acid epoxides and hydroperoxides which have been suggested to be sensitizers in contact allergy, are degraded during the soldering process. However, conditions for the oxidation of colophony may occur in vivo as a result of the colophony‐induced oxidative burst from neutrophils and monocytes. These oxidation products may then interact with body proteins to further initiate immune responses. Therefore for the preparation of low molecular weight chemical (LMWC)–protein conjugates, consideration must be taken to determine whether the LMWC is undergoing a reaction in vivo before it is interacting with body proteins.
Is fungal α‐amylase in bread an allergen?
Authors: Sander  , Raulf‐Heimsoth  , Van Kampen  , Baur
Published: March 2003

Background

The enzyme α‐amylase from Aspergillus oryzae used in bakeries to improve the bread quality has been identified as an inhalative allergen in baker's asthma. It is doubtful whether this enzyme can induce allergic sensitization in regular bread consumers.

Objective

To find out whether fungal α‐amylase in bread and rolls retains its antibody‐binding capacity and allergenicity after the baking procedure.

Methods

Rabbit antibodies directed to fungal α‐amylase were used for the development of a two‐site enzyme‐linked immunosorbent assay (ELISA). This assay was used to analyse different fractions of bread and rolls baked with the usual amounts of α‐amylase in comparison with control products without added enzyme. Competitive experiments between bakers' sera containing specific immunoglobulin (Ig) E to α‐amylase and the rabbit antibodies were performed. Additionally, specific IgE binding to fungal α‐amylase was inhibited by native or heated α‐amylase.

Results

With the highly specific two‐site ELISA for native α‐amylase in the crust of bread bottom and sides, 2.3–7 ng antigenic α‐amylase per gram crust were measured. No α‐amylase could be detected in the crumb fractions. Rabbit antibodies to native α‐amylase completely inhibited human IgE binding to α‐amylase allergen disks.

Conclusion

The results prove residual antibody‐binding capacity of α‐amylase in bread crusts and in the crust of some rolls. In comparison with the content of α‐amylase in dough, between 0.1 and 20% of the antibody‐binding capacity remained.
Exposure and allergic sensitization to cockroach allergen in East Germany
Authors: Hirsch  , Stappenbeck  , Neumeister  , Weiland  , Von Mutius  , Keil  , Leupold
Published: March 2003

Background

Some studies suggest that the prevalence of sensitization to cockroach allergens may be higher in the United States than in Europe, but there are no comparable data from population‐based studies.

Objectives

To determine the prevalence of allergic sensitization to German cockroach (GCR) in German schoolchildren and to assess its clinical relevance; and to determine the exposure to the major GCR allergen Bla g 2 in non‐selected homes and nurseries.

Methods

The prevalence of allergic sensitization to GCR and other allergens was determined by measurement of specific IgE and skin‐prick tests in a cross‐sectional study of 2993 children aged 5–11 years in Dresden, Germany. The prevalence of atopic disease was determined by questionnaire, and pulmonary function and bronchial hyperresponsiveness to hypertonic saline were measured. Bla g 2 exposure was determined on floors of 187 kitchens and 47 nurseries by a commercial sandwich ELISA.

Results

One hundred and twenty‐seven (4.2%) of the children had specific IgE (> 0.7 kU/L) against GCR. Among children with current wheeze, 8.4% were GCR‐sensitized. Compared to data from the United States, the prevalence of sensitization to cockroach was similar in children without asthma (3.9%), but less frequent in asthmatic children from Dresden (6.1%). After adjustment for positive reactions to other allergens (SX1 test) no significant impact of GCR sensitization on wheeze or other symptoms and diagnoses was found. Bla g 2 was detected in 29% of the kitchens and 43% of the nurseries. None of these sites had exposure levels above the proposed threshold for causing disease of 80 ng/g dust.

Conclusion

The data suggest that allergic sensitization to GCR is less frequent in asthmatics from Dresden, Gemany than in US cities. The data indicate that GCR sensitization is not an independent risk factor for asthma and other atopic diseases in 5–11‐year‐olds from this city.
Der p 1 and Der f 1, the highly related and major allergens from house dust mites, are differentially affected by a plant cystatin
Authors: Pernas  , Sanchez‐Ramos  , Sanchez‐Monge  , Lombardero  , Arteaga  , Castañera  , Salcedo
Published: March 2003

Background

Der p 1 and Der f 1 are highly related allergens from the two main house dust mite species, Dermatophagoides pteronyssinus and Dermatophagoides farinae, respectively. A link between the cysteine proteinase activity of Der p 1 and its allergenicity has recently been demonstrated.

Objective

To test the effects of several proteinase inhibitors, mainly a chestnut cystatin (CsC), on the enzymatic activity of Der p 1 and Der f 1, as well as to study the potential acaricide properties of the inhibitors.

Methods

Inhibition tests were performed using mite extracts, as well as isolated Der p 1 and Der f 1 as targets. Immunodetection after SDS‐PAGE and N‐terminal amino acid sequencing were used to demonstrate the inactivation of chestnut cystatin by Der p 1. Bioassays including different inhibitors in a semisynthetic diet were performed to evaluate a potential effect on larval survival.

Results

CsC inhibited the crude digestive proteinase activity of D. farinae, but it was not active towards the equivalent enzyme from D. pteronyssinus. This differential behaviour was fully explained by testing CsC against the two purified allergens, Der f 1 and Der p 1; the former was highly susceptible, whereas the latter was not affected. In contrast, other cysteine proteinase inhibitors, such as egg white cystatin and E‐64, inhibited the proteinase activity of both mite allergens. Der p 1 inactivated CsC by a specific proteolytic cleavage between Gly 6 and Val 7, thus given rise to a noninhibitory processed protein. Besides the in vitro activity, CsC also showed an in vivo effect on D. farinae, drastically increasing the larval mortality when added to a semisynthetic diet.

Conclusion

Der p 1 and Der f 1 behave very differently towards a plant cystatin, thus indicating substantial differences in their proteolytic activity. This fact could be significant with regards to the immunogenic properties of both allergens.
Comparison of genetically engineered hypoallergenic rBet v 1 derivatives with rBet v 1 wild‐type by skin prick and intradermal testing: results obtained in a French population
Authors: Pauli  , Purohit  , Oster  , De Blay  , Vrtala  , Niederberger  , Kraft  , Valenta
Published: March 2003

Background

Bet v 1, the major allergen in birch pollen, is recognized by more than 90% of patients allergic to birch in northern and central Europe. Immunotherapy is commonly performed with birch pollen extracts. Recently, hypoallergenic derivatives of Bet v 1 (rBet v 1 fragments, rBet v 1 dimer and trimer) were constructed and purified.

Objective

Our aim was to compare the allergenic activity of wild‐type rBet v 1 with recombinant Bet v 1 derivatives (rBet v 1 fragments, dimer and trimer) with potentially reduced anaphylactic activity by skin testing in a French population.

Methods

Among the 36 birch pollen allergic patients included in the study, 29 were tested by skin prick testing and 30 by intradermal injections with purified monosubstances: rBet v 1 fragments (F1: aa1–74 and F2: aa75–160), Bet v 1 dimer and trimer. Intradermal tests were performed by the end‐point intradermal titration method. Eight of the intradermally‐tested patients were previously hyposensitized. Tests were performed over a period of 6 months (before, during and after birch pollen season); Bet v 1‐specific IgE and IgG4 subclass responses were measured by immunoblotting and ELISA.

Results

All patients showed lower reactivity with the modified rBet v 1 allergens, both in skin prick and intradermal tests. In 25 and 23 out of 29 patients the lowest concentration of fragment 1 and 2, respectively, resulting in a positive prick test was 100‐fold higher than the lowest concentration of monomer resulting in a positive prick test. For dimer it was 100‐fold or more in 25 out of 29 patients, and for trimer it was 100‐fold or superior in 26 out of 29 patients. By intradermal testing, the end‐point concentration was 160‐fold higher for trimer than for monomer in 24 patients and 40‐fold higher in five patients. For the two fragments the end‐point concentration was 160‐fold higher in 20 out of 22 patients.

Conclusion

Genetically modified hypoallergenic derivatives of the major birch pollen allergen, Bet v 1 showed reduced capacity to induce immediate type skin reactions. They may represent candidate molecules for immunotherapy of birch pollen allergy with reduced risk of anaphylactic side‐effects.
Recombinant Hev b 1: large‐scale production and immunological characterization
Authors: Rihs  , Chen  , Schumacher  , Rozynek  , Cremer  , Lundberg  , Raulf‐Heimsoth  , Petersen  , Baur
Published: March 2003

Background

Hev b 1 represents one of the most important allergens in Hevea brasiliensis latex. It is difficult to get an appropriate amount of native Hev b 1 (nHev b 1) for research purposes.

Objective

The aim of this study was to produce sufficient amounts of Hev b 1 by recombinant methods to prove its suitability for latex allergy diagnostics.

Methods

We isolated total RNA of Hevea brasiliensis leaves and synthesized cDNA by RT PCR. Recombinant Hev b 1 (rHev b 1) as well as three fragments (amino acid residues 29–137, 48–137, 78–137) were subcloned and expressed as fusion proteins with Maltose‐binding protein (MBP) in Escherichia coli. The MBP‐rHev b 1 fusion protein was examined by RAST with the CAP method, histamine release test and immunoblots with human sera from spina bifida patients as well as from health care workers with latex allergy and monoclonal antibodies.

Results

Histamine release test and immunoblots revealed the high allergenicity of the MBP‐rHev b 1 construct. By the CAP method, 54 out of 58 serum samples (93%) from latex‐sensitized spina bifida patients previously showing immunoglobulin (Ig) E to nHev b 1 exhibited IgE‐binding to rHev b 1. Among 71 latex‐allergic health care workers tested, 16 (22.5%) had IgE antibodies to rHev b 1. The analysis of the fusion proteins carrying rHev b 1 fragments revealed that the loss of the N‐terminal 28 amino acid residues did not affect IgE‐binding. In contrast, the lack of the first 47 amino acid residues led to decreased IgE‐binding reactivity in two out of four sera tested, whereas the absence of the N‐terminal 77 residues abolished IgE‐binding in these two sera.

Conclusion

The MBP‐rHev b 1 fusion protein exhibits a corresponding IgE‐binding reactivity to nHev b 1 and may therefore substitute natural Hev b 1 for both in vitro diagnostics and research purposes.
Reduced IFNγ production in response to IL‐12 stimulation and/or reduced IL‐12 production in atopic patients
Authors: Matsui  , Kaneko  , Teramoto  , Fukao  , Inoue  , Kasahara  , Takemura  , Seishima  , Kondo
Published: March 2003

Background

Several studies have shown that interleukin (IL)‐4 and interferon‐gamma (IFNγ) are important for the regulation of IgE production and that IL‐12 induces IFNγ.

Objective

IFNγ production in response to IL‐12 stimulation and IL‐12 production were investigated in peripheral blood mononuclear cells (PBMCs) of atopic patients with various levels of serum IgE.

Methods

Cytokine production from PBMCs was measured following stimulation with a nonspecific stimulator (phytohemagglutinin: PHA), a specific stimulator (Der f 1) and IL‐12 in 17 healthy controls and 23 atopic patients with various serum IgE levels.

Results

The IFNγ production by IL‐12‐stimulated PBMCs in the atopic group was lower than that in the control group. Furthermore, the serum IgE level was negatively correlated with IFNγ production by PBMCs stimulated with IL‐12 (P < 0.001), and with IL‐12 production by PBMCs stimulated with Der f 1 (P < 0.001). Although the IFNγ concentrations by PHA‐stimulated PBMCs were correlated with those by IL‐12‐stimulated PBMCs, there were differences in several patients.

Conclusions

Our results indicated that atopic patients may have some abnormality in the IL‐12‐IFNγ loop. It was shown that the elevation in IgE levels in atopic patients may be due to reduced IFNγ production in response to IL‐12 stimulation and/or due to reduced IL‐12 production.
A moderate and unspecific release of cysteinyl leukotrienes by aspirin from peripheral blood leucocytes precludes its value for aspirin sensitivity testing in asthma
Authors: Pierzchalska M. , Mastalerz L. , Sanak M. , Zazula M. , Szczeklik A.
Published: March 2003

Aspirin‐induced asthma (AIA) is a clinical syndrome related to cysteinyl leukotriene overproduction in airways. The confirmation of the diagnosis requires inconvenient provocation tests with acetyl salicylic acid (ASA).

A study was performed to evaluate whether measurement in vitro of cysteinyl leukotrienes (cys‐LTs) release by isolated peripheral blood leucocytes, stimulated with ASA, can be of use for diagnosis of AIA.

A cellular allergen stimulation test, CAST, was adapted to measure leukotriene release from leucocytes of 32 aspirin‐tolerant and 26 aspirin‐intolerant asthmatics. The cells were stimulated with Lys‐ASA, N‐formyl‐methionyl‐leucyl‐phenylalanine (fMLP), or both fMLP and Lys‐ASA, in a buffer containing IL‐3, and results compared with human leukaemia cell line (Hl‐60) response to Lys‐ASA. Cys‐LTs were measured in cell supernatant fluids by ELISA.

ASA had a rather week stimulatory effect on cys‐LTs release in both groups of patients. Contrary to some previous studies, no significant differences were found between cys‐LTs release by leucocytes from AIA and ATA, or by differentiated Hl‐60 cells.

Measurement of cysteinyl‐leukotriene release by peripheral blood leucocytes pre‐treated with aspirin has no value for diagnosis of AIA.
Microflora‐associated characteristics in faeces from allergic and nonallergic infants
Authors: Böttcher M. F. , Nordin E. K. , Sandin A. , Midtvedt T. , Björkstén B.
Published: March 2003

Background 

The prevalence of allergic diseases has increased particularly over the past 30–40 years. A reduced microbial stimulation during infancy may result in a development of a disturbed balance between Th1‐ and Th2‐like immunity. The gut flora is, quantitatively, the most important source for such stimulation.

Objective 

The aim of the study was to compare the gut microbial flora in 25 allergic and 47 nonallergic 13‐month‐old infants (range 11–18), through analysing microflora‐associated biochemical markers in faeces.

Methods 

Microflora associated characteristics (MACs) were assessed by determining the concentrations of eight different short chain fatty acids and the conversion of cholesterol to coprostanol by gas chromatography. Faecal tryptic activity was analysed spectrophotometrically.

Results 

The allergic infants had lower levels of propionic, i‐butyric, butyric, i‐valeric and valeric acid. In contrast, they had higher levels of the rarely detected i‐caproic acid, which has been associated with the presence of Clostridium difficile. Furthermore, the allergic infants had higher relative distribution of acetic and i‐caproic acid. None of the other parameters differed between the groups.

Conclusion 

The results demonstrate differences in the MACs between allergic and nonallergic infants, indicating differences in the composition of the gut flora that may disturb the development of a normal Th1‐/Th2‐balance in allergic children.
Biologically active recombinant forms of a major house dust mite group 1 allergen Der f 1 with full activities of both cysteine protease and IgE binding
Authors: Yasuhara T. , Takai T. , Yuuki T. , Okudaira H. , Okumura Y.
Published: March 2003

Background Group 1 allergens from mite faeces, Der f 1 and Der p 1, are the most significant in‐door allergens. Therefore, they are the most important component in the standardization of house dust mite extract for diagnosis and allergen‐specific immunotherapy (AIT). Although their cDNAs have been cloned, efforts to prepare biologically active recombinant forms in expression systems using bacteria or yeast have failed.

Objective Our purpose is to establish an efficient system to prepare recombinant Der f 1(rDer f 1), identical in quality to native Der f 1.

Methods The preproforms of Der f 1 and a mutant N53Q, whose consensus motif for N‐glycosylation was disrupted, were expressed in yeast Pichia pastoris. Cysteine protease activity and IgE reactivity were analysed using synthetic substrates and by RAST‐EIA, respectively.

Results The proforms of the two rDer f 1 molecules were efficiently secreted into culture medium. Their prosequences were removed autocatalytically by dialysis against acidic buffer. Although the wild‐type rDer f 1 was more highly glycosylated than native Der f 1, N53Q had almost the same apparent molecular weight as native Der f 1 on SDS‐PAGE. Both the protease and IgE binding activities of the mature rDer f 1 molecules were the same as those of native Der f 1, whereas the proforms had no or markedly reduced activities.

Conclusion The efficient system to prepare active rDer f 1s established in this study is useful for diagnosis and standardized AIT for house dust mite allergy. Furthermore, the system would be a tool for analysis of IgE epitopes, determination of tertiary structure, allergen engineering for safer and more effective AIT, resolving the relation between the enzymatic activity and pathogenesis, and the development of therapeutic inhibitors.
Identification of some rabbit allergens as lipocalins
Authors: Baker J. , Berry A. , Boscato L. M. , Gordon S. , Walsh B. J. , Stuart M. C.
Published: March 2003

Rabbits are frequently used as laboratory animals or kept as domestic pets. Rabbit serum albumin and a 17‐kDa protein referred to as Ory c 1 have previously been reported as allergens. Several other allergenic proteins have been recognized by crossed immuno‐electrophoresis but have not been characterized.

The aim of this study was to characterize the allergenic proteins present in rabbit saliva, urine and fur on the basis of molecular size and, where possible, to determine their amino acid sequences.

Extracts from the male New Zealand white rabbit were used for developing specific direct RAST and RAST inhibition assays. Proteins in the extracts were separated by SDS‐PAGE and the individual allergens identified by immunoblotting with serum from rabbit‐allergic individuals. The N‐termini of four allergens were sequenced.

Saliva was the most potent extract. In total, 26 protein bands were recognized as allergens in the three extracts: 12 in saliva, seven in urine and seven in fur. Their molecular weights ranged from an 8‐kDa species in saliva to an 80‐kDa protein in urine. The N terminal sequences of an 18 kDa and a 21‐kDa species in saliva, were identified as lipocalins with sequence similarity to a recently described odourant binding protein.

This is the first evidence that allergens from the rabbit are members of the lipocalin superfamily of proteins, suggesting that similar mechanisms may be involved in eliciting the allergic response to rabbits. The 18 kDa allergen from saliva may be the previously named rabbit allergen, Ory c 1.
Diagnostic value of skin‐prick and patch tests and serum eosinophil cationic protein and cow's milk‐specific IgE in infants with cow's milk allergy
Authors: Saarinen K. M. , Suomalainen H. , Savilahti E.
Published: March 2003

The diagnosis of cow's milk allergy is based on a clinical response to an elimination‐challenge test with cow's milk.

We studied the usefulness of the skin‐prick and patch tests and measurement of cow's milk‐specific IgE and eosinophil cationic protein in serum as diagnostic tools for cow's milk allergy in a cohort of 6209 unselected infants followed from birth for the development of cow's milk allergy.

Of the 239 infants challenged with cow's milk, 118 showed a positive and 121 a negative response at a mean age of 6.9 months. A positive reaction to a skin‐prick test with cow's milk ( 3 mm) was seen in 72 (61%) and 29 (24%) infants with positive and negative challenges, elevated serum cow's milk‐specific IgE ( 0.7 kU/L) in 52 (45%) and 15 (13%) infants, a positive reaction to patch test with cow's milk protein fractions in 26 (26%) and eight (8%) infants, and elevated serum eosinophil cationic protein ( 20 μg/L) in 22 (21%) and seven (13%) infants, respectively. Parallel use of the four tests with the above‐mentioned cut‐off values correctly classified 73% of the infants with a sensitivity of 0.76 and a specificity of 0.67. An immediate reaction to cow's milk challenge correlated with skin prick test positivity and elevated serum milk‐specific IgE, and tended to correlate with patch test positivity.

No single test or parallel use of the four tests could predict the challenge outcome acceptably in this prospectively followed, unselected cohort of 6209 infants. A positive reaction to one or more tests needs to be confirmed by a challenge test and a negative response to all four tests does not rule out the possibility of cow's milk allergy.
Discontinuous IgE‐binding epitopes contain multiple continuous epitope regions: results of an epitope mapping on recombinant Hol l 5, a major allergen from velvet grass pollen
Authors: Schramm G. , Bufe A. , Petersen A. , Haas H. , Merget R. , Schlaak M. , Becker W.‐M.
Published: March 2003

The knowledge of IgE‐binding epitopes on allergen molecules is important for better understanding allergen–antibody interactions and, thus, for developing new strategies for immunotherapy.

Our purpose was to more precisely define the number and structure of IgE‐binding epitopes of a paradigmatic major grass pollen allergen.

We performed an IgE‐binding epitope mapping of rHol l 5, a group V pollen allergen of velvet grass (Holcus lanatus), with overlapping fragments (length between 15 and 186 amino acids), which were expressed in E. coli as MBP fusion proteins. Using sera of 65 grass pollen allergic patients, the fragments were analysed by immunoblotting for IgE reactivity. Specificity of antibody binding was confirmed by competitive blot inhibition assays.

At least four different continuous IgE‐binding epitopes were identified on small fragments (about 30 amino acids), and at least five different discontinuous IgE‐binding epitopes on larger fragments, which were destroyed by further fragmentation. The fragments were differentially recognized by individual patients' sera. By investigating IgE‐binding to one of the small fragments in more detail, we found further epitope regions on this fragment. It was noteworthy that IgE reactivity to small fragments was weak compared to large fragments or to the complete molecule. Competitive blot inhibition experiments showed that binding of IgE antibodies to the small fragments was specific but with lower avidity than to the complete rHol l 5.

rHol l 5 harbours multiple discontinuous as well as continuous IgE‐binding epitopes spread over the whole molecule, which were individually recognized by IgE antibodies from different patients. Low avidity of IgE antibodies to small fragments suggests that the continuous epitope regions do not represent the complete epitope and are most probably parts of discontinuous epitopes.
Inhibition of IgE‐induced activation of human mast cells by IL‐10
Authors: Royer B. , Varadaradjalou S. , Saas P. , Guillosson J. J. , Kantelip J. P. , Arock M.
Published: March 2003

Background IL‐10 exhibits anti‐inflammatory effects on activated rodent mast cells (MC) in vitro and inhibits allergen‐induced airway inflammation in vivo in murine models. The effects of IL‐10 on the allergic activation of human MC are presently unknown.

Objective In light of the well‐known heterogeneity of mast cell reactivity between animal species, one cannot readily predict the response of human MC to IL‐10. Moreover, the impact of IL‐10 on MC‐derived proinflammatory mediators is still unknown. Thus, the objective of this study was to investigate the effects of IL‐10 on the release of inflammatory mediators by IgE/anti‐IgE‐challenged human cord blood‐derived mast cells (CBMC), used as an in vitro model of MC phenotypically similar to human lung MC.

Materials and methods Highly purified human MC were obtained by a first step of long‐term culture of cord blood mononuclear cells in the presence of human recombinant stem cell factor (rhSCF) and of human recombinant IL‐6 (rhIL‐6), followed by a second step of purification by depletion of contaminating cells with an immunomagnetic method. The cells were treated with human IgE, then challenged with anti‐human IgE, in the presence or the absence of recombinant rhIL‐10 used at various concentrations. Histamine, tumour necrosis factor‐alpha (TNF‐α), IL‐5 and IL‐8 were measured in the various supernatants collected at different times after the beginning of the challenge.

Results IL‐10 inhibited the release of TNF‐α and of IL‐8, but not of IL‐5, by activated CBMC. Interestingly, IL‐10 also inhibited the release of histamine by activated CBMC, contrasting with data reported for rodent MC.

Conclusions These findings suggest that IL‐10 might have anti‐inflammatory effects on IgE/anti‐IgE‐challenged human MC by inhibiting their release of TNF‐α, IL‐8 and histamine. These data provide new insights into the control of human mast cell activation and might lead to a better knowledge of the cellular mechanisms controlling allergic reactions.
Comparison of once daily fluticasone propionate aqueous nasal spray with once daily budesonide reservoir powder device in patients with perennial rhinitis
Authors: Kivisaari E. , Baker R. C. , Price M. J.
Published: March 2003

Background Previous studies comparing the corticosteroids fluticasone propionate (FP) and budesonide (BUD) in both perennial and seasonal rhinitis have shown no consistent difference between treatments. However, the therapeutic outcomes may have been influenced by study design.

Objective To compare the effect of FP aqueous nasal spray (ANS; 200 µg/day) with BUD reservoir powder device (RPD; 200 µg/day) on rhinitis symptoms, productivity loss and device preference in patients with perennial rhinitis.

Methods After a 2‐week run‐in period, 440 patients were randomized to receive either FPANS, BUD RPD or matched placebo (ANS or RPD) for 8 weeks, followed by an open‐label 4‐week follow‐up treatment with FPANS. Patients completed diary card visual analogue scores for nasal symptoms, and questionnaires on satisfaction with the treatment and preferred choice of device.

Results During weeks 1–4, the visual analogue total nasal symptom scores (VATNS) in the FPANS group were significantly lower than scores in the BUD RPD group (mean difference =−17.8; 95% CI = −34.4, −1.3; P = 0.036). FPANS also significantly reduced the VATNS compared with the ANS placebo at all time‐points assessed (P ≤ 0.005). BUD RPD did not significantly differ from the RPD placebo at weeks 5–8 (P = 0.167), or the ANS placebo at any time‐point (P ≤ 0.151). Over the 8‐week treatment period FPANS was significantly more effective than BUD RPD at reducing sneezing (mean difference =−4.4; 95% CI = −8.6, −0.3; P = 0.036) and nasal itching (mean difference =−5.3; 95% CI = −9.9, −0.8; P = 0.022), and was significantly superior to the ANS placebo for all symptoms assessed at weeks 1–4 and 1–8 (P < 0.016). At the same time‐points BUD RPD was no better at alleviating nasal itching than the RPD placebo (P ≤ 0.306), and compared with the ANS placebo, significantly reduced only one symptom; nasal blockage (P ≤ 0.016). After 8 weeks of treatment, patients preferred the ANS device to the RPD (P < 0.001), and at 12 weeks a significantly greater number of patients were satisfied with FPANS treatment compared with BUD RPD (P = 0.0019) or the respective placebos (P = 0.0001).

Conclusion FPANS and BUD RPD are effective therapies with a good safety profile for the treatment of perennial rhinitis but, in this direct placebo‐controlled comparison, FPANS was more efficacious than BUD RPD, and the patients preferred the ANS device to the RPD.
Novel B and T cell epitopes of chicken ovomucoid (Gal d 1) induce T cell secretion of IL‐6, IL‐13, and IFN‐γ
Authors: Holen E. , Bolann B. , Elsayed S.
Published: March 2003

Background Chicken ovomucoid (OM, Gal d 1) has an important role in the pathogenesis of IgE‐mediated allergic reactions to hen's egg white.

Objectives The purpose of this study was to clarify the mechanisms of T cell recognition of ovomucoid using intact OM and chemically modified, characterized and homogeneous solid phase synthetic peptides covering the whole molecule.

Methods Eighteen overlapping peptides were prepared by solid phase F‐moc polyamide peptide synthesis (SPPS), characterized and high‐pressure liquid chromatography (HPLC) purified. The peptides, together with intact, denatured and oxidized OM, were used to stimulate patient‐derived cell cultures for mapping T cell epitopes. Proliferation responses, T cell phenotype and cytokine secretion using peripheral blood mononuclear cells (PBMC) from eight individuals and T cell lines (TCL) derived from six hen's egg‐allergic patients, were examined. In addition, intact, denatured, oxidized and deglycosylated OM, as well as the peptides solely or with their keyhole limpet haemocyanin (KLH) complexes, were tested. For locating IgE and IgG B cell epitopes, seven egg‐allergic patient sera and three OM‐polyclonal sera were used. Healthy non‐allergic individuals were included as controls.

Results Seven peptides were recognized by specific IgE, while OM‐specific TCL recognized 10 peptides. Six of the OM peptides were commonly recognized both by patient S‐IgE and blood‐derived TCL. Among those, one novel epitope, peptide OM 61–74, had the ability to bind IgE. Another peptide, OM 101–114, was recognized by IgE and IgG sera, but not by any of the TCLs. In contrast, the peptides OM 41–56, OM 71–84, OM 131–144 and OM 171–186 were exclusively T cell epitopes with no affinity to specific antibodies. Abundant TCL secretion of IFN‐γ, IL‐6, IL‐4, IL‐13, IL‐10 and TNF‐α in response to OM stimulation indicates the contribution of Th2 as well as Th1/Th0 CD4+ cell subsets. For allergic patients moderate amounts of IFN‐γ, IL‐13, and high amounts of IL‐6, were secreted in response to TCL stimulation by OM peptides. High amounts of IL‐6 were secreted in response to all molecular forms of OM (intact‐, modified‐OM and the peptides 71–84 and 51–64) when TCLs from two non‐allergic donors were used.

Conclusions One novel B cell epitope (OM 61–74) and 10 T cell epitopes have been identified. The most reactive epitopes of the OM molecule comprise the motifs 1–14 to 71–84, the overlapping peptide‐pairs OM 121–134 and OM 131–144 and peptides OM 161–174 and 171–186. Peptides OM 1–14 and 171–186 are the only ones capable of inducing IL‐4 secretion. Only one peptide (OM 11–24) induces IL‐10 secretion. Those peptides recognized as both T and B cell epitopes or only T cell epitopes, have the potential to induce T cell secretion of moderate to high amounts of IL‐13, IFN‐γ and particularly IL‐6.
Reducing the need for food allergen challenges in young children: a comparison of in vitro with in vivo tests
Authors: Hill D. J. , Hosking C. S. , Reyes‐Benito L. V.
Published: March 2003

Background Double‐blind placebo‐controlled food challenges (DBPCFC), the gold standard for the diagnosis of food hypersensitivity, are time‐consuming and not without risk. We have recently reported skin prick test (SPT) weal diameters to cow's milk, egg and peanut above which infants and young children referred for investigation of suspected food allergy showed an adverse reaction on food challenge. We have termed these the ‘100% diagnostic SPT levels’. In this study, we compare in vivo with in vitro measurement of IgE antibody levels to three common food allergens — cow's milk, egg and peanut — in infants and young children with suspected food allergy, in order to reduce the need for food challenges.

Methods SPT and Enzyme Allergo‐sorbent Test (EAST) (from 1992 to 1998) and CAP values (from 1999 to 2000) were performed in 820 children < 2 years of age with suspected allergy to cow's milk and/or egg and/or peanut. SPT levels previously shown to be diagnostic of challenge‐proven allergy to cow's milk, egg and peanut were used as the ‘100% diagnostic SPT levels’ and compared with EAST and CAP values associated with IgE food allergy according to the manufacturer's definition.

Results McNemar's test showed a significant difference between the ‘100% diagnostic SPT levels’ and positive EAST in identifying patients who did not require food challenge for cow's milk (P = 0.01), egg (P < 10−6) and peanut (P < 10−6), and a significant difference between the ‘100% diagnostic SPT levels’ and positive CAP (P < 10−6) for egg and peanut but not cow's milk. Twenty‐three per cent of food challenges which, based on the results of EAST and CAP, would have been necessary to confirm the diagnosis of food allergy were avoided by the use of the ‘100% diagnostic SPT levels’.

Conclusion The use of the ‘100% diagnostic SPT levels’ compared with in vitro measurement of IgE antibody to cow's milk, egg and peanut reduces the need for food challenge in young children with suspected food allergy.
Cetirizine inhibits skin reactions but not mediator release in immediate and developing late‐phase allergic cutaneous reactions. A double‐blind, placebo‐controlled study
Authors: Nielsen P. N. , Skov P. S. , Poulsen L. K. , Schmelz M. , Petersen L. J.
Published: March 2003

Background Recent reports have indicated cetirizine, a potent H1‐receptor antagonist, to possess a number of anti‐inflammatory effects, e.g. inhibition of mast cell degranulation and inhibition of leucocyte migration and activation.

Objective The aim of this study was to compare the effects of cetirizine on skin responses and mediator release in intact skin in immediate and developing late‐phase allergic reactions by microdialysis technique.

Methods Cetirizine 10 mg once daily or matching placebo were administered to 10 atopic subjects for 6 days followed by a 2‐week washout in a randomized, double‐blind, placebo‐controlled, cross‐over trial. Immediate skin test responses to allergen, codeine, and histamine and late‐phase reactions to allergen were assessed. The time course of extracellular levels of inflammatory mediators in intact skin were monitored by microdialysis techniques using 2 kDa and 3 MDa cut‐off fibers, respectively.

Results Cetirizine significantly reduced immediate weal and flare reactions to allergen, codeine, and histamine. Injection of allergen, but not buffer controls, induced a significant release of histamine, tryptase, prostaglandin D2, total protein, and eosinophilic cationic protein. No significant increase of leukotriene B4 and myeloperoxidase was observed. Cetirizine inhibited early total protein extravasation by 40%, but this did not reach a significant level. None of the inflammatory mediators were significantly inhibited by cetirizine. Cetirizine significantly reduced the late‐phase skin induration to allergen by approximately 30%.

Conclusion Cetirizine potently reduced skin responses in immediate allergic reactions without inhibition of early mediators. These data indicate cetirizine to be a potent H1‐receptor antagonist with no effect on mast cell activation. It did not inhibit any of the late‐phase mediators, but it reduced the late skin reaction. These data suggest that mediators other than those actually measured may play a significant role in the clinical late‐phase reaction.
Prevalence of respiratory and atopic disorders in Chinese schoolchildren
Authors: Wong G. W. K. , Hui D. S. C. , Chan H. H. , Fok T. F. , Leung R. , Zhong N. S. , Chen Y. Z. , Lai C. K. W.
Published: March 2003

Background Epidemiological surveys have shown that the prevalence of asthma in the Asian population is relatively low. Within the Chinese population, schoolchildren from Hong Kong were found to have the highest rate of asthma.

Objective To compare the prevalence of respiratory and atopic disorders, and to assess the role of atopy in the development of asthma, in Chinese schoolchildren from Hong Kong, Beijing and Guangzhou.

Methods Community‐based random samples of schoolchildren aged 9–11 years from three Chinese cities (Hong Kong, Beijing and Guangzhou) were recruited for study using the International Study of Asthma and Allergies in Childhood (ISAAC) Phase II protocol. Subjects were studied by parental questionnaires (n = 10902), skin‐prick tests (n = 3479) and skin examination (n = 3479).

Results The prevalence rates of current wheeze, speech limiting wheeze, rhinoconjunctivitis and flexural dermatitis were significantly more common in Hong Kong than in Beijing or Guangzhou. The atopy rate was also higher in Hong Kong (41.2%) than in Beijing (23.9%) or Guangzhou (30.8%). Atopy was strongly correlated with current wheeze (OR 7.74; 95% CI = 5.70–10.51). Subgroup analyses of children from Hong Kong revealed that children born in mainland China who had subsequently migrated to Hong Kong had a significantly lower rate of allergic symptoms and atopy than those children born in Hong Kong.

Conclusion Using a standardized written questionnaire along with a skin prick test and skin examination, we confirmed that the prevalence of asthma, allergic diseases and atopy was highest in schoolchildren from Hong Kong. Atopic sensitization is an important factor associated with asthma in Chinese children.