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H Breiteneder

Publications and source records attributed to H Breiteneder.

At least 91 records · Page 5Linked to original sources
PubMed1996

Dissection of immunoglobulin E and T lymphocyte reactivity of isoforms of the major birch pollen allergen Bet v 1: potential use of hypoallergenic isoforms for immunotherapy.

We dissected the T cell activation potency and the immunoglobulin (Ig) E-binding properties (allergenicity) of nine isoforms of Bet v 1 (Bet v 1a-Bet v 1l), the major birch pollen allergen. Immunoblot experiments showed that Bet v 1 isoforms differ in their ability to bind IgE from birch pollen-allergic patients. All patients tested displayed similar IgE-binding patterns toward each particular isoform. Based on these experiments, we grouped Bet v 1 isoforms in three classes: molecules with high IgE-binding activity (isoforms a, e, and j), intermediate IgE-binding (isoforms b, c, and f), and low/no IgE-binding activity (isoforms d, g, and 1). Bet v 1a, a recombinant isoform selected from a cDNA expression library using IgE immunoscreening exhibited the highest IgE-binding activity. Isoforms a, b, d, e, and 1 were chosen as representatives from the three classes for experimentation. The potency of each isoallergen to activate T lymphocytes from birch pollen-allergic patients was assayed using peripheral blood mononuclear cells, allergen-specific T cell lines, and peptide-mapped allergen-specific T cell clones. Among the patients, some displayed a broad range of T cell-recognition patterns for Bet v 1 isoforms whereas others seemed to be restricted to particular isoforms. In spite of this variability, the highest scores for T cell proliferative responses were observed with isoform d (low IgE binder), followed by b, 1, e, and a. In vivo (skin prick) tests showed that the potency of isoforms d and 1 to induce typical urticarial type 1 reactions in Bet v 1-allergic individuals was significantly lower than for isoforms a, b, and e. Taken together, our results indicate that hypoallergenic Bet v 1 isoforms are potent activators of allergen-specific T lymphocytes, and Bet v 1 isoforms with high in vitro IgE-binding activity and in vivo allergenicity can display low T cell antigenicity. Based on these findings, we propose a novel approach for immunotherapy of type I allergies: a treatment with high doses of hypoallergenic isoforms or recombinant variants of atopic allergens. We proceed on the assumption that this measure would modulate the quality of the T helper cell response to allergens in vivo. The therapy form would additionally implicate a reduced risk of anaphylactic side effects.

Allergens↗
PubMed1996

Diversity of human T cell receptor sequences of T cell clones with specificity for Bet v 1 peptide/MHC II complexes.

T cell clones (TCC) were raised from the peripheral blood of patients suffering from tree pollen allergy. All TCC were restricted by HLA-DR molecules. In order to investigate possible intervention targets in Type I allergic diseases, we have examined T cell receptor (TCR) alpha- and beta-chain nucleotide sequences of several allergen-reactive human CD4+ TCC specific for four frequently found epitopes of Bet v 1, the major birch pollen allergen. In general, TCC specific for the 4 epitopes investigated, used diverse TCRAV and TCRBV gene segments. Moreover, the junctional regions encoding the third complementarity determining regions (CDR3) of the TCR showed striking heterogeneities in length and amino acid composition. A more restricted use of two J gene segments (TCRBJ1S4 and 2S7) was only observed in the beta-chain of TCR used by TCC specific for epitope 1. In addition, all TCC specific for epitope 4 showed an arginine residue in the N-terminal region of their TCRBV CDR3 loops despite their sequence diversities. In view of the striking heterogeneities found, therapeutical strategies aimed at the clonal deletion of allergen-specific T cell clones, providing help for IgE synthesis, may not be feasible. Moreover, our results cast a doubt on the theory, that the CDR3 exclusively provides the primary contact with the peptide bound in the major histocompatibility (MHC) groove, and suggest additional interaction with MHC class II.

Allergens↗
PubMed1996

A randomized, placebo-controlled, double-blind crossover study of the effect of pentoxifylline on ocular fundus pulsations.

PURPOSE: To estimate the short-term effects of pentoxifylline on ocular blood flow in healthy volunteers. METHODS: In ten healthy subjects, either 200 or 400 mg of pentoxifylline or placebo was administered intravenously over 90 minutes in a randomized, double-blind, placebo-controlled, three-way crossover study design. Noninvasive measurements of blood pressure, pulse rate, flow variables in the radial artery, ocular fundus pulsations, and whole blood viscosity and filterability were performed at baseline and at 30-minute intervals until four hours after the start of drug infusion. Ocular fundus pulsation amplitude, which has been shown to estimate the pulsatile component of the ocular blood flow, was recorded with a laser interferometer in the macula, the optic disk, and a peripheral region. RESULTS: Fundus pulsation amplitude significantly increased after infusion of pentoxifylline, with maximum effect 150 to 180 minutes after the start of the infusion. This effect was dose-dependent and more pronounced in the macula (+17%, P < .001 vs baseline, after 200 mg; and +27%, P < .001 vs baseline, after 400 mg of pentoxifylline) and the peripheral region (+14%, P < .001; and +26%, P < .001) than in the optic disk (+11%, P < .002; and +13%, P < .001). Viscosity but not filterability of whole blood dose-dependently decreased. Peak systolic flow velocity in the radial artery decreased after infusion of 400 mg of pentoxifylline. Blood pressure and pulse rate were unchanged during the observation period. CONCLUSIONS: These findings demonstrate that pentoxifylline increases pulsatile ocular blood flow in healthy volunteers. Our results support the possibility that pentoxifylline could be used therapeutically in several eye diseases, such as diabetic retinopathy.

Adult↗
PubMed1996

Sequence comparisons of the CDR3 hyper-variable loops of human T cell receptors specific for three major T cell epitopes of the birch pollen allergen Bet v 1.

We have analysed the T cell receptor (TCR) alpha and beta chain sequences of 16 human CD4+ T cell clones (TCCs) specific for three important epitopes of the major birch pollen allergen Bet v 1. The TCCs were raised from the peripheral blood of eight patients with birch pollen allergy, showing allergic rhino-conjunctivitis and allergic asthma. The TCCs from these individuals were specific for Bet v 1-derived peptides: amino acids (aa)77-92 (epitope 1), aa93-108 (epitope 2) and aa113-126 (epitope 3). The DNA sequence analysis of the TCRAV and BV regions revealed heterogeneous repertoires for recognition of the peptides. Multiple combinations of AV/AJ and BV/BJ were used. However, some inter-individual restriction was evident. A limited selection of AVS and the normally infrequently used BV1S4 was obvious in TCCs specific for epitope 1. The TCRBV13 was more frequent in TCCs recognizing epitope 3. A very narrow distribution in length could be seen in the CDR3 sequences of the beta chain of TCRs with specificity for epitopes 1 and 2. Inter-individual positional micro-restriction was observed for the aa motif LR in the tCDR3 (epitope 1), for the aa residue M in the alphaCDR3 and for the aa residue G in the betaCDR3 (epitope 3). Our results illustrate clearly that each antigenic peptide derived from a single allergen, is capable of selecting different characteristics in the responding repertoire of TCRs, thus increasing the complexity of allergen-recognition by T lymphocytes. Therefore, our findings limit the potential use of TCR targeted therapeutical strategies in Type I allergy.

Allergens↗
PubMed1996

Effects of desmopressin on circulating P-selectin.

Von Willebrand factor (VWF) and P-selectin share an identical intracellular storage compartment and transport to the cell surface. We induced degranulation of the Weibel-Palade bodies and measured circulating (c)P-selectin in plasma to test whether soluble P-selectin is present in the Weibel-Palade bodies, or if P-selectin bound to the cell membrane of endothelial cells (EC) is rapidly proteolytically cleaved in vivo. A dose of 0.3 microgram/kg of desmopressin (DDAVP) or placebo was infused over 30 min to eight healthy men in a double-blind cross-over study. Plasma levels of cP- selectin and VWF-Ag were followed for 24 h. Despite a twofold increase in VWF-Ag levels immediately after a 2 h after infusion of DDAVP, no significant change in plasma concentrations of cP-selectin were observed. In summary, degranulation of the Weibel-Palade bodies is an unlikely source of cP-selectin. Thus cP-selectin in healthy subjects is conceivably attributable to other mechanisms, such as minor degrees of platelet activation or transcription induction of an alternatively spliced P-selectin.

Adult↗
PubMed1996

High-level expression of tree pollen isoallergens in Escherichia coli.

cDNAs coding for the major allergen of alder (Alnus glutinosa) pollen Aln g 1, for nine isoforms of Bet v 1, the major birch (Betula verrucosa) pollen allergen, and for four isoforms of Cor a 1, the major allergen of hazel (Corylus avellana) pollen, were inserted into the plasmid pMW175 or pMW 172 and expressed in Escherichia coli as recombinant non-fusion proteins. These constructs produced between 20 and 160 mg protein/l. The recombinant tree pollen isoallergens were tested in immunoblots for their antibody binding properties. For this purpose, we used two monoclonal antibodies (BIP 1 and BIP 4) raised against natural Bet v 1, a polyclonal rabbit anti-recombinant Bet v 1a, as well as serum IgE from allergic patients. Our results show that this expression system is suitable for the production of milligram amounts of tree pollen isoallergens which can be used for the characterization of allergenic epitopes recognized by T and B cells.

Allergens↗
PubMed1995

Molecular characterization of Api g 1, the major allergen of celery (Apium graveolens), and its immunological and structural relationships to a group of 17-kDa tree pollen allergens.

Individuals suffering from immediate hypersensitivity (type-I allergy) to a particular pollen frequently display intolerance to several foods of plant origin. In this respect, individuals sensitized to birch pollen and/or mugwort pollen frequently display type-I allergic symptoms after ingestion of celery. In this study, we expressed the major allergenic protein of celery, Api g 1, which is responsible for the birch-celery syndrome, in the form of a non-fusion protein. The open reading frame of the cDNA of Api g 1 codes for a protein of 153 amino acids with a molecular mass of 16.2 kDa and 40% identity (60% similarity) to the major allergen of birch pollen, Bet v 1. Furthermore, Api g 1 exhibited similar characteristics to (a) two proteins in parsley induced by fungal infection, (b) the major tree pollen allergens and (c) pathogenesis-related and stress-induced proteins in other plant species. The reactivity of recombinant Api g 1 with IgE antibodies present in sera from celery intolerant patients was comparable to that of the natural celery allergen. Cross-reactivity with Bet v 1 was proven by cross-inhibition experiments, which provides further support for the existence of the birch-celery syndrome and for the suggestion that allergies to some vegetable foods are epiphenomena to allergies caused by inhalation of tree pollen.

Allergens↗
PubMed1995

Cloning and sequencing of Mal d 1, the major allergen from apple (Malus domestica), and its immunological relationship to Bet v 1, the major birch pollen allergen.

The number of tree-pollen-allergic patients showing allergic reactions to apples, various vegetables and nuts is increasing. In this paper the molecular characterization of the major apple allergen, Mal d 1, is reported. The cDNA coding for Mal d 1 was cloned and sequenced. Its open reading frame codes for a protein of 159 amino acids with a predicted molecular mass of 17.7 kDa and a predicted pI of 5.9. Sequence comparison to Bet v 1, the major birch pollen allergen, revealed 64.5% identity on the amino acid level and 55.6% identity on the nucleic acid level. Recombinant Mal d 1 was expressed in the plasmid pMW 175 in E. coli BL 21 (DE3) and its immunological properties were tested. Crossreactivity with Bet v 1 was shown by inhibition assays.

Allergens↗
PubMed1995

Isoforms of Bet v 1, the major birch pollen allergen, analyzed by liquid chromatography, mass spectrometry, and cDNA cloning.

Bet v 1, the major allergen of birch pollen, displays a considerable degree of heterogeneity. Several charge variants have been detected by two-dimensional IgE immunoblots and isoelectric focusing techniques. This heterogeneity has been attributed to glycosylation (or other post-translational modifications) or to isogenes coding for Bet v 1 isoforms and/or allelic variants. However, until now, only limited structural data for Bet v 1 have been published. Recently, we described the expression, purification, and immunological properties of recombinant Bet v 1 (rBet v 1) produced in Escherichia coli as a non-fusion protein (Ferreira, F. D., Hoffmann-Sommergruber, K., Breiteneder, H., Pettenburger, K., Ebner, C., Sommergruber, W., Steiner, R., Bohle, B., Sperr, W. R., Valent, P., Kungl, A. J., Breitenbach, M., Kraft, D., and Scheiner, O. (1993) J. Biol. Chem. 268, 19574-19580). Here, we present a more detailed structural characterization of Bet v 1 by both cDNA cloning and mass spectrometry. Thirteen different cDNA clones coding for Bet v 1 isoforms were obtained by polymerase chain reaction amplification of birch pollen cDNA with a sequence-specific 5'-terminal primer and a nonspecific 3'-terminal primer or by immunological screening of a birch pollen cDNA library. These isoforms are referred to as Bet v 1b to Bet v 1n, whereas the previously isolated Bet v 1 cDNA (Breiteneder, H., Pettenburger, K., Bito, A., Valenta, R., Kraft, D., Rumpold, H., Scheiner, O., and Breitenbach, M. (1989) EMBO J. 8, 1935-1938) is now referred to as Bet v 1a. High performance liquid chromatography and plasma desorption mass spectrometry of proteolytic fragments of purified natural Bet v 1 (nBet v 1) and rBet v 1a were used to (i) confirm the primary structure of all Bet v 1 isoforms and (ii) to investigate any possible postsynthetic modifications on rBet v 1a or on the natural mixture of isoallergens obtained from birch pollen. Except for the cleavage of initiating methionine, no postsynthetic modifications were found in either nBet v 1 or rBet v 1a.

Allergens↗
PubMed1995

Diversity of TCRAV and TCRBV sequences used by human T-cell clones specific for a minimal epitope of Bet v 1, the major birch pollen allergen.

T-cell clones (TCC) were raised from the peripheral blood of patients suffering from tree pollen allergy. All TCC were restricted by HLA-DR molecules. In order to investigate possible intervention targets in Type I allergic diseases, we examined T-cell receptor (TCR) alpha and beta chain nucleotide sequences of five allergen-reactive human CD4+ TCC specific for a C-terminal epitope (BV 144) of Bet v 1, the major birch pollen allergen. Proliferation assays using synthetic peptides revealed the 10-mer LRAVESYLLA as minimal epitope for three TCC; two TCC also displayed reactivity with the nonapeptide LRAVESYLL. Two TCC expressed TCRBV2S3, all other BV144-specific TCC used diverse TCRAV and TCRBV gene segments. Moreover, the junctional regions encoding the third complementary determining regions (CDR3) of the TCR showed a striking heterogeneity in length and amino acid composition. Nevertheless, all TCC showed an arginine residue in the N-terminal region of their TCRBV CDR3 loops. Therefore, therapeutical strategies aimed at the clonal deletion of allergen-specific T-cell clones, providing help for IgE synthesis, will not be feasible. Our results cast a doubt on the theory that the CDR3 exclusively provides the primary contact with the peptide bound in the major histocompatibility (MHC) groove, and suggest additional interaction with MHC class II.

Allergens↗
PubMed1995

Identification of allergens in fruits and vegetables: IgE cross-reactivities with the important birch pollen allergens Bet v 1 and Bet v 2 (birch profilin).

BACKGROUND: In this study serum samples collected from 20 patients with birch pollen allergy were investigated. All patients had experienced allergic symptoms after contact with or ingestion of particular fresh fruits and vegetables known as birch pollen-related foods. METHODS: Serum samples were tested by means of immunoblotting for IgE reactivities with proteins in extracts of birch pollen, apple, pear, celery, carrot, and potato. Anti-Bet v 1 and anti-Bet v 2 antibodies were used to investigate cross-reactivity. Inhibition studies were performed by preincubation of sera with recombinant Bet v 1 and Bet v 2. RESULTS: IgE binding to proteins, corresponding to the major birch pollen allergen Bet v 1 and to Bet v 2 (birch pollen profilin) could be observed. An allergen homologous to Bet v 1 could be detected in apple, pear, and celery when a Bet v 1-specific monoclonal antibody was used. Testing a polyclonal rabbit anti-Bet v 2 antibody with extracts of the respective plants revealed the presence of profilins in every source tested. Inhibition with recombinant Bet v 1 and Bet v 2 led to complete blocking or marked reduction of IgE binding to proteins of comparable molecular weights in the respective food extracts, indicating IgE cross-reactivity. CONCLUSION: Our results indicate that many plant-derived food agents contain proteins with high homology to the birch pollen allergens Bet v 1 and Bet v 2 and must therefore be considered as potentially threatening for patients with tree pollen allergy.

Allergens↗
PubMed1995

T-cell epitopes of Phl p 1, major pollen allergen of timothy grass (Phleum pratense): evidence for crossreacting and non-crossreacting T-cell epitopes within grass group I allergens.

BACKGROUND: The use of peptides representing T-cell epitopes of allergens is a modern concept for improvement of specific immunotherapy. A prerequisite for this approach is the identification of T-cell epitopes of atopic allergens. METHODS: T-cell lines and 40 T-cell clones (TCC) specific for Phl p 1, the group I allergen of timothy grass (Phleum pratense), were established from the peripheral blood of nine patients allergic to grass pollen and mapped for epitope specificity by using overlapping dodecapeptides. Phenotype and cytokine production profile of TCC were investigated. Representative TCC were analyzed for HLA-restriction, T-cell receptor V beta gene usage, and crossreactivity with grass pollen extracts from Dactylis glomerata, Poa pratensis, Lolium perenne, Secale cereale, and selected amino acid sequence-derived peptides. RESULTS: Patients displayed IgE binding to all grass species investigated. Forty TCC were established. Fifteen T-cell epitopes could be identified on Phl p 1. Of 40 TCC, 39 displayed the helper cell (Th) phenotype; one clone was CD8+. Specific stimulation induced a Th2-like type of cytokine production in 20 of 39 TCC. Crossreactivity studies revealed crossreacting and non-crossreacting T-cell epitopes. CONCLUSION: Phl p 1, a major grass pollen allergen, harbors multiple T-cell epitopes. Species-specific and crossreacting T-cell epitopes exist among group I allergens of grasses. Epitope recognition patterns could not be correlated with particular HLA haplotypes. A restricted T-cell receptor V beta gene usage was not observed.

Allergens↗
PubMed1995

Comparison of non-invasive methods for the assessment of haemodynamic drug effects in healthy male and female volunteers: sex differences in cardiovascular responsiveness.

1. The study was performed to determine the sensitivity and short-term and day-to-day variability of a novel technique based on laser interferometry of ocular fundus pulsations and of non-invasive methods for the quantification of haemodynamic drug effects. An additional aim was to assess sex differences in haemodynamic responsiveness to cardiovascular drugs in male and female healthy volunteers. 2. Ten males and nine females (age range 20-33 years) were studied in a double-blind, randomized, cross-over trial. Simultaneous measurements from systemic haemodynamics, laser interferometry of ocular fundus pulsations, systolic time intervals from mechanocardiography, a/b ratio from oxymetric fingerplethysmography and Doppler sonography of the radial artery were used to describe the haemodynamic effects of cumulative, stepwise increasing intravenous doses of phenylephrine, isoprenaline, sodium nitroprusside and of placebo. 3. Laser interferometry detected the isoprenaline-effects at the lowest dose level of 0.1 micrograms min-1 with a high signal-to-noise ratio. The reproducibility of measurements under baseline was high, no changes were observed after systemically effective doses of phenylephrine or sodium nitroprusside. Systolic time intervals were sensitive and specific for isoprenaline-induced effects, PEP and QS2c-measurements had high reproducibility. Fingerplethysmography proved a sensitive measurement for the detection of the vasodilating effects of sodium nitroprusside, but was not specific, and showed low reproducibility. Measurements from Doppler sonography had lower reproducibility and sensitivity compared with the other applied methods. 4. There was a significant sex difference for several of the haemodynamic parameters under baseline conditions; however, the responsiveness to the drugs under study was not different, when drug effects were expressed as %-change from the baseline. 5. Laser interferometry is a valuable non-invasive, highly sensitive and specific approach for the detection of pulse pressure changes. A battery of non-invasive tests appears useful for the characterization of cardiovascular drugs. Gender differences may not pose a relevant problem for the study of acute haemodynamic effects of cardiovascular drugs.

Adult↗
PubMed1994

Four recombinant isoforms of Cor a 1, the major allergen of hazel pollen, show different reactivities with allergen-specific T-lymphocyte clones.

Purified preparations of allergenic proteins from plants, and in particular from pollens, consist of multiple closely related isoforms. These isoforms are highly similar in their amino acid sequences, yet they display different properties with respect to antibody binding. In this study we report of differential potencies of cross-reacting tree pollen allergens and cloned isoforms of these allergens to activate allergen-specific T-lymphocyte clones (T-cell clones; TCC). Six TCC with specificity for Bet v 1, a representative tree pollen major allergen, were established from peripheral blood of five birch-pollen-allergic donors. All TCC displayed the helper-cell phenotype. Five TCC reacted with distinct epitopes present on natural (n) and on recombinant (r) Bet v 1. One TCC could not be stimulated with r Bet v 1, in spite of strong reactivity with purified natural Bet v 1. The TCC were tested in proliferation assays using purified n Bet v 1, n Cor a 1 (the homologous major allergen of hazel pollen), r Bet v 1, four recombinant isoforms of Cor a 1 and peptides representing corresponding T-cell stimulating regions (isoepitopes) on these proteins. The clones showed different patterns of reactivity in response to stimulation with the five recombinant molecules and the corresponding peptides. Certain exchanges of amino acids within stimulating peptides correlated with a lack of proliferation of the TCC tested. These findings are important with respect to the use of broadly cross-reactive recombinant allergens or allergen-derived peptides for immunotherapy of type I allergy.

Allergens↗
PubMed1994

Environmental stress-mediated differential 3' end formation of chloroplast RNA-binding protein transcripts.

We report the characterization of transcripts from the halophyte, Mesembryanthemum crystallinum, encoding a protein with high homology to chloroplast RNA-binding proteins (cRBP). In this plant chloroplast-related functions are largely protected against salt stress. cRBP transcripts are derived from a single gene, Mc32crbp, although three size classes of polyadenylated mRNAs are detected. Transcription rate and steady state amounts of mRNA are developmentally regulated and light controlled with strong transcriptional activity as functional chloroplasts are established, and with lower maintenance activity thereafter. Upon salt stress, the rate of transcription decreases, although transcript levels increase. Accompanying stress, a change in the distribution of transcript size classes is observed as the longest transcript with an untranslated 3' end of 381 nucleotides increases relative to transcripts with shorter 3' ends. The long transcript is characterized by the presence of five sequence elements in the 3'-untranslated region that are present in cRBP mRNAs from a variety of plants, although not all elements are found in each mRNA. The results may indicate a mechanism by which mRNA levels of constitutively light-regulated genes may be modulated without enhanced transcription in response to environmental cues.

Adaptation, Physiological↗