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H Breiteneder

Publications and source records attributed to H Breiteneder.

At least 55 records · Page 3Linked to original sources
PubMed1999

IgE reactivity to Api g 1, a major celery allergen, in a Central European population is based on primary sensitization by Bet v 1.

BACKGROUND: Up to 70% of patients with tree pollen allergy display allergic symptoms when eating certain fruits and vegetables. Homologous proteins with allergenic features are present in a wide range of plant species and can cause allergic reactions. OBJECTIVE: The aim of this study was to evaluate recombinant Api g 1, a major celery allergen, for in vivo and in vitro diagnosis of celery allergy in populations from Davos, Switzerland, and Montpellier, France. METHODS: A group of patients with celery and birch pollen allergy from Davos was tested, and the results from those tests were compared with results from a group of patients allergic to celery from Montpellier. Skin prick tests were performed with a commercial celery extract, crude celery, and purified recombinant Api g 1. Quantitative and qualitative serology was done with natural and recombinant allergens by means of RASTs and immunoblotting. RESULTS: Recombinant Api g 1 allowed accurate in vivo diagnosis of celery allergy in all patients from the Swiss group. RAST results with celery extract were negative in 8 of 24 patients; results of immunoblotting with celery extract were negative in 4 of 24 patients, and results of immunoblotting with recombinant (r)Api g 1 were negative in 8 of 24 patients. In the French group 11 of 12 patients had a positive skin reaction with crude celery extract, but only 2 patients reacted with rApi g 1. RAST results for celery were positive in 8 of 12 patients. In immunoblotting experiments 8 patient sera displayed IgE directed against various celery allergens, whereas no patients sera had rApi g 1-specific IgE. CONCLUSION: Our results document that rApi g 1 allows accurate in vivo diagnosis only in areas where birch trees are common. In areas where no birch trees grow, primary sensitization takes place through different pollen allergens (eg, mugwort pollen). Moreover, it became evident that birch pollen and celery allergy are highly related in Central Europe, whereas in Southern Europe the mugwort-celery type is predominant.

Adult↗
PubMed1999

New Bet v 1 isoforms including a naturally occurring truncated form of the protein derived from Austrian birch pollen.

Bet v 1, the major pollen allergen from white birch, displays a considerable degree of heterogeneity. Until now, all molecular and immunological characterization studies of Bet v 1 isoforms have been performed with commercially available pollen of Swedish origin. In regard to clinical studies with Austrian birch pollen allergic individuals, knowledge about the isoform repertoire in Austrian birch pollen was necessary. cDNAs coding for Bet v 1 isoforms from Austrian birch pollen were cloned by PCR amplification and sequenced. Besides the Austrian variants of the Swedish isoforms Bet v 1a (62% of the clones), ALK167 (4%), and Bet v 1d/h, Bet v 1g, and Bet v 11 (24%), three sequences with a significantly lower homology to known isoforms and two Bet v 1a-homologous sequences with a 7 bp insertion coding for a truncated protein were detected. No Austrian variants of the majority of the Swedish isoforms were found. The isoforms coding for truncated proteins were expressed in Escherichia coli and tested by immunoblotting. They bound a polyclonal anti-Bet v 1 antibody but did not recognize birch pollen allergic patients' serum IgE and two Bet v 1-specific monoclonal antibodies. The similarity of the Bet v 1 isoform patterns of Swedish and Austrian birch pollen justifies the use of Bet v 1 derived from Swedish pollen for clinical studies with birch pollen allergic individuals from outside Northern Europe.

Allergens↗
PubMed1999

Identification of a highly promiscuous and an HLA allele-specific T-cell epitope in the birch major allergen Bet v 1: HLA restriction, epitope mapping and TCR sequence comparisons.

BACKGROUND: Allergen-specific CD4+ T cells play an important regulatory role in atopic allergy. OBJECTIVE: To investigate the human leucocyte antigen (HLA) restriction and T-cell receptor (TCR) usage of allergen-specific T-cell clones (TCCs) that react with defined epitopes of Bet v 1, the major birch pollen allergen. METHODS: Five Bet v 1-specific TCCs derived from two birch pollen-allergic individuals and specific for Bet v 1, were epitope-mapped with overlapping synthetic peptides. In addition, HLA-restriction and TCR CDR3 sequences were determined. RESULTS: Three TCCs reacted with a Bet v 1 peptide containing amino acid residues 21-33 (BP21), the other two TCCs reacted with a minimal peptide comprising residues 37-45 (BP37). Studies using neutralizing anti-HLA-monoclonal antibodies and HLA-typed APCs showed that the BP37-specific TCCs were restricted by a HLA-DQA1*0301/DQB1*0603 heterodimer. In contrast, BP21 was recognized in a highly promiscuous manner. TCCs recognizing this sequence were restricted by HLA-DPB1*0201, a HLA-DQA1*0201/DQB1*0201 heterodimer, or HLA-DRB3*0101. Reverse transcription-polymerase chain reaction with primers for all known TCRAV and TCRBV gene segments, followed by CDR3 region sequencing, revealed the usage of five different TCRAV and four different TCRBV gene segments by the TCCs, as well as diversity in the joining region. All BP21-specific TCCs contained a negatively charged residue in their CDR3alpha regions, the CDR3beta regions showed a high concentration of polar and OH-group bearing residues. BP37-specific TCCs shared the amino acid combination LY in the middle of their CDR3alpha regions, the CDR3beta regions showed high concentration of OH-group bearing or charged residues. CONCLUSIONS: This study shows the existence of a highly promiscuous T-cell epitope in Bet v 1. The presence of additional T-cell epitopes in Bet v 1 may, however, hamper the clinical applicability of the epitope. Likewise, the diversity in TCR usage by T cells recognizing the epitope does not support the development of TCR-directed immunotherapy for birch pollen allergy.

Alleles↗
PubMed1999

Molecular characterization of Dau c 1, the Bet v 1 homologous protein from carrot and its cross-reactivity with Bet v 1 and Api g 1.

BACKGROUND: Up to 70% of patients with birch pollen allergy exhibit the so-called oral allergy syndrome, an IgE-mediated food allergy. The most frequent and therefore best characterized pollen-fruit syndrome is apple allergy in patients suffering from tree pollen-induced pollinosis. The occurrence of adverse reactions to proteins present in vegetables such as celery and carrots in patients suffering from pollen allergy has also been reported. cDNAs for Bet v 1 homologous proteins have been cloned from celery, apple and cherry. Objective The aim of the study was to identify Bet v 1 homologues from carrot (Daucus carota), to isolate the respective cDNA, to compare the IgE-binding capacity of the natural protein to the recombinant allergen and determine the cross-reactivity to Api g 1 and Bet v 1. METHODS: Molecular characterization of the carrot allergen was performed using IgE-immunoblotting, cross-inhibition assays, N-terminal sequencing, PCR-based cDNA cloning and expression of the recombinant protein in Escherichia coli. RESULTS: A 16-kDa protein from carrot was identified as a major IgE-binding component and designated Dau c 1. Sequencing corresponding cDNAs revealed three extremely similar sequences (Dau c 1.1, 1.2 and 1.3) with an open reading frame of 462 bp coding for 154 amino acid residues. CONCLUSIONS: Purified recombinant Dau c 1.2 was tested in immunoblots displaying IgE-binding capacity comparable to its natural counterpart. Cross-inhibition assays verified the existence of common B-cell epitopes present on Dau c 1, Api g 1 as well as on Bet v 1.

Allergens↗
PubMed1999

Allergen mimotopes in food enhance type I allergic reactions in mice.

BIP1is a murine IgG antibody capable of enhancing the IgE binding to Bet v 1, the major birch pollen allergen. We have previously generated a mimotope of BIP1, designated Bet mim 1, from a constrained phage display peptide library. We demonstrated that oral immunization of BALB/c mice with the Bet mim 1 mimotope resulted in the induction of Bet v 1-specific IgG. The aim of this study was to test the influence of such an oral immunization with Bet mim 1 on a subsequent type I allergic response to Bet v 1. Phages displaying Bet mim 1 or control mimotopes, or PBS alone, were delivered to BALB/c mice by intragastric gavages prior to systemic sensitization with recombinant Bet v 1 and Al(OH)(3), an adjuvant inducing preferentially IgE antibody responses. Only mice fed with Bet mim 1-phages displayed substantially enhanced type I allergic skin reactivity to Bet v 1, as compared to mice pretreated with control mimotopes or PBS. A gastric digestion assay indicated that Bet v 1 and its homologue from apple, Mal d 1, were degraded within seconds under physiological conditions. In contrast, phage-displayed mimotopes were resistant to digestion. Our data indicate that allergen mimics in the diet that resist digestion, can induce allergen specific IgG able to enhance an allergic response. We therefore conclude that sensitization via the oral route may represent a mechanism for aggravating type I allergic reactions, probably leading to an earlier onset of symptoms even at lower allergen dosage.

Allergens↗
PubMed1998

Identification and cloning of prs a 1, a 32-kDa endochitinase and major allergen of avocado, and its expression in the yeast Pichia pastoris.

Avocado, the fruit of the tropical tree Persea americana, is a source of allergens that can elicit diverse IgE-mediated reactions including anaphylaxis in sensitized individuals. We characterized a 32-kDa major avocado allergen, Prs a 1, which is recognized by 15 out of 20 avocado- and/or latex-allergic patients. Natural Prs a 1 was purified, and its N-terminal and two tryptic peptide sequences were determined. We isolated the Prs a 1 encoding cDNA by PCR using degenerate primers and 5'-rapid amplification of cDNA ends. The Prs a 1 cDNA coded for an endochitinase of 326 amino acids with a leader peptide of 25 amino acids. We expressed Prs a 1 in the yeast Pichia pastoris at 50 mg/liter of culture medium. The recombinant Prs a 1 showed endochitinase activity, inhibited growth and branching of Fusarium oxysporum hyphae, and possessed IgE binding capacity. IgE cross-reactivity with latex proteins including a 20-kDa allergen, most likely prohevein, was demonstrated, providing an explanation for the commonly observed cross-sensitization between avocado and latex proteins. Sequence comparison showed that Prs a 1 and prohevein had 70% similarity in their chitin-binding domains. Characterization of chitinases as allergens has implications for engineering transgenic crops with increased levels of chitinases.

Allergens↗
PubMed1998

BRCA1-related breast cancer in Austrian breast and ovarian cancer families: specific BRCA1 mutations and pathological characteristics.

We identified 17 BRCA1 mutations in 86 Austrian breast and ovarian cancer families (20%) that were screened for mutations by denaturing high-performance liquid chromatography (DHPLC) and the protein truncation test (PTT). Eleven distinct mutations were detected, 4 of them (962del4, 2795del4, 3135del4 and L3376stop) not previously reported in families of non-Austrian origin. In addition, 6 rare missense mutations (allele frequency < 1%) with unknown biological effects were identified. Four mutations occurred more than once in the Austrian population: 2795del4 (3 times), Cys61Gly (3 times) 5382insC (2 times) and Q1806stop (2 times). Haplotype analysis of the 4 recurrent mutations suggested a common ancestor for each of these. Thirty-four breast cancer cases from 17 families with BRCA1 mutations were further analyzed. We observed a low median age of onset (39.5 years). Sixty-eight percent of all BRCA1 breast cancer cases had negative axillary lymph nodes. This group showed a significant prevalence of a negative estrogen and progesterone receptor status and stage I tumors compared with an age-related, node-negative control group. The prevalence of grade III tumors was marginally significant. Survival analysis either with a control group matched for age (within 5 years), grade, histologic subtype and estrogen receptor status, or with an age-related, node-negative comparison group, showed no statistical difference.

Adult↗
PubMed1998

cDNA cloning of the 43-kDa latex allergen Hev b 7 with sequence similarity to patatins and its expression in the yeast Pichia pastoris.

IgE-mediated hypersensitivity to latex proteins present in health care products, particularly in latex gloves, has become an important public health problem in recent years. We purified natural Hev b 7, a 43-kDa patatin-like allergen from the latex of Hevea brasiliensis and determined several internal peptide sequences. A heterologous hybridization probe of a patatin gene of potato, to which these peptides could be aligned best, was used to screen a latex cDNA library. The cDNA encoded an acidic protein of 388 amino acids with a molecular mass of 42.9 kDa. The deduced amino acid sequence had 39-42% identity to patatins from Solanum tuberosum. The purified recombinant Hev b 7 expressed in the yeast Pichia pastoris displayed, similarly to patatins from S. tuberosum, esterase activity. Both natural and recombinant Hev b 7 were recognized by IgE from sera of latex-sensitized allergic individuals. In contrast to patatins from S. tuberosum and Nicotiana tabacum, natural Hev b 7 lacked an N-terminal leader peptide for targeting to the endoplasmatic reticulum and was not glycosylated. These results establish the 43-kDa patatin-like protein as a latex allergen and raise the possibility of different cellular localization and function compared to S. tuberosum patatins.

Allergens↗
PubMed1998

Long-lived Th2 clones specific for seasonal and perennial allergens can be detected in blood and skin by their TCR-hypervariable regions.

We investigated the longevity of allergen-specific Th cells derived from patients suffering from either allergic rhinitis or atopic dermatitis. T cell clones (TCC) specific for seasonal and perennial allergens were raised. To determine whether these TCC were long-lived in vivo, PBMC and allergen-specific polyclonal T cell lines, collected and established inside a period of up to 4 years, were screened for the TCC of interest. For this purpose, a T cell tracing protocol was established in which oligonucleotides specific for the TCR beta-chain hypervariable junctional region were used as tools to identify each particular TCC. Seven pollen-specific TCC and two house dust mite-specific TCC, with a Th2-like cytokine production pattern in vitro, were demonstrated to be long-lived memory T cells in vivo. Specificity of the tracing protocol was ascertained by TCR sequence analysis. We conclude that allergen-specific TCC can persist for years, evidence for which can be monitored in blood, but also in the target organ of the allergic disorder. The data indicate that in vitro-characterized, allergen-specific, long-lived TCC may well reflect a repertoire of T lymphocytes of pathogenetic importance in vivo.

Adult↗
PubMed1998

A mimotope defined by phage display inhibits IgE binding to the plant panallergen profilin.

Birch pollen and mugwort pollen allergies are often associated with hypersensitivity to plant foods. This clinical and serological cross-reactivity is mediated by IgE antibodies reacting with homologous proteins in pollen and food. Cross-reacting homologs of the important birch pollen allergen Bet v 2 (profilin) could be detected in other pollen, fruits, nuts, and vegetables, such as celery tuber. We purified IgG/IgE antibodies from the serum of an exclusively profilin-allergic patient using affinity columns either coupled with protein extracts from mugwort pollen, birch pollen, or celery tuber. Constrained and unconstrained random nonapeptide libraries were pooled and screened with the anti-profilin antibody preparations to define cross-reactive ligands. Specific ligands were enriched by successive panning rounds using the profilin-specific antibodies in series. After the last panning round enriched phage clones were screened with purified profilin-specific antibodies and IgE-binding clones were sequenced. Five out of eight positive clones (62.5 %) displayed the same circular peptide CAISGGYPVC. This peptide was synthesized and examined for its ability to inhibit IgE binding to blotted mugwort pollen, birch pollen, or celery tuber profilin. Inhibition studies showed reduction of IgE binding to profilins in all three protein extracts. As the sequence of the mimotope did not show any homology to the known birch profilin sequence this peptide is considered to mimic a common conformational IgE epitope for these examined profilins.

Allergens↗
PubMed1998

Topical fundus pulsation measurements in age-related macular degeneration.

BACKGROUND: The purpose of the present study was to investigate regional fundus pulsations in age-related macular degeneration (AMD) patients with subretinal neovascular membranes. METHODS: Local fundus pulsation amplitudes (FPAs) were measured in 12 patients with AMD with classic neovascular membranes. Measurements were performed directly on the membrane and adjacent to the membrane. FPAs were assessed with a recently developed laser interferometric method. FPA measurements were performed in 12 healthy subjects at similar posterior pole locations. RESULTS: In AMD patients FPAs were consistently lower when measured directly on the neovascular membrane ("inside") than at measurement sites around the membrane ("outside"). The difference in FPA was 26 +/- 3% (mean +/- SEM, range 13-40%, P < 0.0001). In healthy subjects, however, FPAs were significantly higher at the measurement points corresponding to "inside" points (15 +/- 4%, P < 0.0006). CONCLUSIONS: We have shown that FPAs are reduced at classic neovascular membranes in patients with AMD. The mechanism behind this finding remains unclear. Hence, future studies have to ascertain whether this observation is associated with changes in fundus layers or with local choroidal perfusion abnormalities.

Aged↗
PubMed1998

Bet v 1, the major birch pollen allergen, and Mal d 1, the major apple allergen, cross-react at the level of allergen-specific T helper cells.

BACKGROUND: Food allergy to apple is frequent in individuals allergic to tree pollen. The major allergens of birch, Bet v 1, and apple, Mal d 1, have been cloned and sequenced and display a high degree of sequence identity, leading to IgE cross-reactivity. OBJECTIVE: We sought to investigate cross-reactivity between Bet v 1 and Mal d 1 at the level of allergen-specific T lymphocytes. METHODS: PBMCs of 13 patients allergic to birch pollen with oral allergy syndrome to apple were stimulated with rBet v 1 and rMal d 1, respectively, thereby establishing allergen-specific T-cell lines and T-cell clones. rMal d 1-specific T-cell cultures were tested for reactivity with rBet v 1, and rBet v 1-specific T cells were analyzed for reactivity with apple allergen. Cytokine production patterns in response to specific stimulation were evaluated. A selection of cross-reacting T-cell clones was mapped for epitope specificity by the use of overlapping Bet v 1- derived peptides. RESULTS: Nineteen Mal d 1-specific T-cell clones were produced, 79% of which cross-reacted with Bet v 1. Eight of 18 Bet v 1-specific T-cell clones cross-reacted with Mal d 1. Six peptides representing cross-reactive T-cell epitopes could be identified. The respective fragments from birch and apple displayed approximately 50% amino acid identity. Seventy percent of the cross-reactive T-cell clones revealed a T(H2)-like cytokine production pattern. CONCLUSION: The results indicate that cross-reactivity between apple and birch pollen leading to the clinical oral allergy syndrome occurs not only at the serologic, but also at the cellular level.

Adult↗
PubMed1998

Conserved sequence motifs of CDR3 loops of TCR specific for two major epitopes of the grass pollen allergen PhI p 1.

We analyzed the cDNA sequence data of complementarity determining regions (CDR3) of epitope mapped alphabeta TCR of T cell clones (TCC). The TCC were specific for the major timothy grass (Phleum pratense) pollen allergen Phl p 1 and were derived from the peripheral blood of seven unrelated grass pollen-allergic individuals. Each TCR recognized one of two immunodominant T cell epitopes, PP73 or PP103, of Phl p 1. Although a diversity of recombined V and J segments was observed, amino acid motifs as long as five residues were conserved among CDR3 loops of TCR from TCC of different atopic individuals specific for the same peptide. The conserved sequences could comprise as much as 60% of the CDR3. All amino acid residues of the motifs of the CDR3beta and most of the CDR3alpha of all TCR used in this study were encoded by randomly added nucleotides. This indicates that they were specifically selected for by the peptide bound to the MHC class II molecule. For one selected patient, a larger number of TCC, specific for PP103, was analyzed. The TCR repertoire was limited to three different TCR. The same MHC class II molecule, DRB1*1301, was identified to present PP103 to each of the three TCR.

Allergens↗
PubMed1998

Peptide mimotopes displayed by phage inhibit antibody binding to bet v 1, the major birch pollen allergen, and induce specific IgG response in mice.

The major birch pollen allergen Bet v 1 is one of the most extensively characterized allergens both on the molecular and the immunological level. To define conformational B cell epitopes on Bet v 1, we screened filamentous phage libraries expressing circular or linear nonapeptides to select ligands specific for anti-Bet v 1 murine monoclonal antibodies BIP1 and BIP4. The deduced amino acid sequence of the BIP1 ligand was CFPYCYPSESA, and of the BIP4-ligand, CRQTRTMPGC. Both sequences derived from the circular phage library. Alignments to the sequence of Bet v 1 showed no similarities, indicating that the antibodies most likely recognize discontinuous epitopes. Phages displaying these mimotopes were capable of inhibiting interactions of the anti-Bet v 1 monoclonals with Bet v 1 in a dose-dependent manner in ELISA. In contrast, sequence-identical synthetic peptides were ineffective in blocking the antibody-allergen interactions. This is in agreement with the conformational inhomogeneity of the peptides in solution as observed by nuclear magnetic resonance spectroscopy. Intragastric administration of phages expressing the BIP1 mimotope induced a Bet v 1-specific IgG response in Balb/c mice. We conclude that peptide mimotopes, when displayed on phages, may induce a protective IgG response preventing IgE-mediated allergic reactions, suggesting a possible clinical application.

Administration, Oral↗
PubMed1998

Modulation of IgE reactivity of allergens by site-directed mutagenesis: potential use of hypoallergenic variants for immunotherapy.

Specific immunotherapy is an efficient treatment for patients suffering from type I allergy. The mechanisms underlying successful immunotherapy are assumed to operate at the level of T helper cells, leading to a modulation of the immune response to allergens. During immunotherapy, increasing doses of allergens are given on a regular basis, and the beneficial effects for the patient depend on the concentration of allergen used. On the other hand, the risk of IgE-mediated anaphylactic side effects also increase with the amount of allergen applied per injection. Therefore, we have proposed the use of hypoallergenic (low IgE binding activity) forms of allergens for immunotherapy. We evaluated by site-directed mutagenesis the contributions of individual amino acid residues/positions for IgE binding to Bet v 1, the major allergen of birch pollen. We found that IgE binding to Bet v 1 depended on at least six amino acid residues/positions. Immunoblot analyses and inhibition experiments showed that the multiple-point Bet v 1 mutant exhibited extremely low reactivity with serum IgE from birch pollen-allergic patients. In vivo (skin prick) tests showed that the potency of the multiple-point mutant to induce typical urticarial type I reactions in pollen-allergic patients was significantly lower than for wild-type Bet v 1. Proliferation assays of allergen-specific T cell clones demonstrated that these six amino acid exchanges in the Bet v 1 sequence did not influence T cell recognition. Thus, the Bet v 1 six-point mutant displayed significantly reduced IgE binding activity, but conserved T cell activating capacity, which is necessary for immunomodulation. The approach described here may be generally applied to produce allergen variants to be used in a safe therapy form of immediate-type allergies.

Allergens↗