bluuebunny/crossref_metadata_2025 huggingface dataset:
Isoforms of the Major Allergen of Birch Pollen Induce Different Immune Responses after Genetic Immunization
Authors: Hartl Arnulf , Kiesslich Jan , Weiss Richard , Bernhaupt Andrea , Mostböck Sven , Scheiblhofer Sandra , Flöckner Hannes , Sippl Manfred , Ebner Christof , Ferreira Fátima , Thalhamer Josef
Published: April 2003

Background: Recent publications indicate that immunization with plasmid DNA encoding allergens might represent a potential approach in allergen–specific immunotherapy. Objective: In the present study we have compared the immune responses induced by plasmid DNA encoding for two isoforms of Bet v 1, the major allergen of birch pollen. Methods: BALB/c mice were injected intradermally with plasmid DNA encoding for the genes of Bet v 1a (pCMV–Beta) and Bet v 1d (pCMV–Betd). In addition, the effect of immunostimulatory DNA sequences was investigated by appending and/or coinjecting CpG motifs. Antibody responses and IFN–γ and IL–4 levels were measured by ELISA. Allergen–specific proliferation was determined by incorporation of [3H]–thymidine. Results: The two isoforms induced a similar humoral response. The lack of any IgE production and the ratio of IgG1 to IgG2a clearly indicated a Th–1–type response. The antisera against both isoforms were highly cross–reactive, which was supported by the energy plot indicating similar folding of the two protein isoforms. However, determination of IFN–γ and IL–4 in the serum elicited a strikingly different cytokine profile during the course of the immune response. In contrast to pCMV–Beta, pCMV–Betd caused no significant allergen–specific proliferation and induced only marginal levels of the key cytokines. Conclusions: Based on the assumption that the induction of a strong Th–1 type response is a prerequisite for successful treatment of allergy, our results favor the use of isoform Bet v 1a in combination with CpG motifs for a novel type of allergen immunotherapy based on plasmid DNA immunization. Additionally, the data also confirm the assumption that the antigen itself can have a marked influence on the immune response after genetic immunization.
Involvement of Carbohydrate Epitopes in the IgE Response of Celery–Allergic Patients
Authors: Fötisch K. , Altmann F. , Haustein D. , Vieths S.
Published: April 2003

Background: This study was performed to get further insights into antibody responses to cross–reactive carbohydrate determinants (CCD), including initial experiments to prove the biological activity of anti–CCD IgE. Earlier studies have shown that IgE specific for CCD occurs in about 25% of celery–allergic patients. The clinical significance of these antibody specificities is doubtful. Methods: Patient sera were selected on the basis of a positive case history of celery allergy and multiple binding to high molecular weight celery allergens on immunoblots. Specific IgE to native and heated celery tuber was determined by the enzyme allergosorbent test (EAST). N–glycans were purified after extensive digestion of specific glycoproteins, such as pineapple stem bromelain, bovine fibrin, and human IgG, and used as antigens in an IgE ELISA as well as in EAST and immunoblotting inhibition experiments. Dose–related histamine release was performed with BSA neoglycoproteins containing 3–4 units of the purified glycopeptides. Results: Seven celery–allergic patients were identified who clearly presented IgE against the N–glycan purified from bromelain which is a common structure within the plant kingdom. Chemical defucosylation showed that α1,3–fucose is a key structure for IgE binding. In patients with anti–CCD IgE, the maximal inhibition of celery EAST by the bromelain glycan ranged from 22 to 100%. Inhibition of celery immunoblots by preincubation of patient serum with this glycan led to a quenching of multiple bands at masses >40 kD. After linking the bromelain glycopeptide to BSA, a strong dose–related histamine release was obtained in a celery–allergic patient, occurring at lower concentrations than with the recombinant major protein allergen from celery, Api g 1. Conclusions: Our results demonstrate that IgE specific for CCD is common in celery–allergic patients, and can represent the major proportion of IgE against this food. α1,3–fucose was confirmed to be an essential part of the IgE epitope. Immunoblotting inhibition indicated the presence of this carbohydrate determinant on multiple glycoproteins in celery extract. Although histamine release was only performed in 1 patient, our data show that proteins carrying multiple glycan units can be biologically active in patients sensitized to CCD.
Evaluation of Specific IgE to the Recombinant Group 2 Mite Allergens Lep d 2 and Tyr p 2 in the Pharmacia CAP System
Authors: Johansson Eva , Eriksson Tove Lisa Jenny , Olsson Susanna , Kronqvist Marianne , Whitley Paul , Johansson S.G.O. , Gafvelin Guro , van Hage-Hamsten Marianne
Published: April 2003

Background: Several recombinant allergens have been shown to be potentially useful for diagnosis of IgE–mediated allergy, but only a few recombinant allergens are at present commercially available in serological assays for detection of specific IgE. The aim of this study was to evaluate the IgE binding to the recombinant major dust mite allergens rLep d 2 and rTyr p 2 and compare it with the IgE binding to the commercial mite extracts Lepidoglyphus destructor and Tyrophagus putrescentiae in the Pharmacia RAST CAP System. Methods: The recombinant allergens rLep d 2 and rTyr p 2 were immobilised on ImmunoCAPs, and sera from 461 Swedish farmers who are frequently exposed to mites were analysed for specific IgE antibodies. Immunoblotting was performed to evaluate discrepancies between the results obtained with the recombinant and the commercial CAP assays. Results: The IgE values of each recombinant assay significantly correlated with the IgE values of the corresponding commercial CAP assay. The sensitivity of the rLep d 2 assay was 73.3% and that of the rTyr p 2 assay, 60.5% of that provided by the commercial L. destructor and T. putrescentiae assays. Two subjects out of 416, who tested negative in the commercial L. destructor assay, were positive to rLep d 2. The corresponding figures for rTyr p 2 and the T. putrescentiae extract were 5/418. The possibility that these subjects were sensitised to L. destructor and T. putrescentiae could not be excluded. Conclusions: The data suggest that it may be possible to use rLep d 2 and rTyr p 2 on ImmunoCAPs to detect and quantify IgE antibodies to these, the major allergens of L. destructor and T. putrescentiae. It appears likely that the addition of just a few more recombinant L. destructor and T. putrescentiae allergens in the CAP assay will be sufficient for in vitro diagnosis of IgE mediated allergy to L. destructor and T. putrescentiae.
Lactic Dehydrogenase Virus Infection Inhibits Allergic Eosinophil Reaction and IL–5 Gene Expression in vivo
Authors: Morimoto Masahiro , Iwata Hiroyuki , Hayashi Toshiharu
Published: April 2003

The effects of lactic dehydrogenase virus (LDV) infection on allergic eosinophil reaction and IL–5 gene expression were studied. LDV infection suppressed antigen–induced eosinophil recruitment into the peritoneal cavity in sensitized mice. The elevation of IL–5 gene expression in the spleen and mesenteric lymph nodes 6 h after ovalbumin challenge was significantly suppressed in LDV–infected mice compared with uninfected (control) mice. The expression of the interferon–γ and IL–2 genes in the spleen, but not in mesenteric lymph nodes, was significantly suppressed in LDV–infected mice compared with control mice. The present results suggest, that suppression of IL–5 gene expression by LDV infection may not be mediated by a mutual inhibitory mechanism between Th1 and Th2 cells.
Immunoglobulin Structure and Function as Revealed by Electron Microscopy
Authors: Roux Kenneth H.
Published: April 2003

Electron–microscopic (EM) analysis preceded crystallographic analysis [1, 2]of Igs by over a decade and was for a time the only direct way of analyzing their 3–D molecular structure. Once the X–ray structures were deduced, the role of EM gradually shifted from gross structural analysis to the addressing of more sophisticated structural and functional questions. EM remains a vital adjunct to the many physicochemical, biochemical, and serological tools brought to bear on these remarkable molecules as we try to relate form to function. In this review I will highlight some of the many contributions that have been made possible by virtue of being able to ‘see’ Ig molecules and immune complexes.
Murine Allergic Respiratory Responses to the Major House Dust Mite Allergen Der p 1
Authors: Clarke Amanda H. , Thomas Wayne R. , Rolland Jennifer M. , Dow Christopher , O’Brien Richard M.
Published: April 2003

Background: Although many studies have examined chronic asthma, limited data exist on acute immunopathogenic events induced by allergens. The aim of the study was to investigate the acute cellular, serologic and histopathologic events in airway inflammation produced by intranasal challenge of mice sensitised to the major house dust mite allergen Der p 1. Methods: C57BL/6 mice were immunised subcutaneously with Der p 1 in alum. Mice were bled and challenged intranasally with Der p 1 on day 14 and killed on day 17. Lungs were fixed in situ, processed and stained with haematoxylin and eosin. The degree of inflammation and eosinophil infiltration was quantified by image analysis. Specific IgE was determined by passive cutaneous anaphylaxis. Cells from spleen and draining lymph nodes were cultured for 24 h with Der p 1, and IL–3/GM–CSF released into supernatants was measured by bioassay. Results: Intranasal challenge of sensitised mice induced eosinophilic influx into the large and small airways and the alveolar regions of the lung, mucus plugging and in severe cases numerous Charcot–Leyden crystals. The quantitation of the inflammation induced by different sensitisation and challenge doses showed that optimal inflammation could be produced using only 1 μg of allergen for both sensitisation and challenge. The degree of inflammation was not related to the titre of IgE antibody and was indeed produced in its absence. T cell reactivity of spleen cells to the allergen was decreased suggesting cell migration or inactivation. Conclusions: Mice sensitised and challenged intranasally with as little as 1 μg of Der p 1 produced an extensive pulmonary eosinophilic inflammation which shared many of the features of the inflammation found in asthma. The small amount of allergens required and the use of intranasal challenge should provide a useful model.
Induction of IgE Responses Using a Recombinant Mosquito Salivary Allergen rAed a 2 without Adjuvant in Mice
Authors: Wang Hongsheng , Mao Xiaojuan , Simons F. Estelle R. , Peng Zhikang
Published: April 2003

Background: Reactions to mosquito bites are a global problem. Several salivary proteins from Aedes (Ae.) aegypti, the most common mosquito species, have been cloned and expressed. Plasmid DNA vaccination has been shown to be effective in the downregulation of IgE responses. To investigate the in vivo antigenicity of these recombinant proteins and to study the mechanisms underlying plasmid DNA vaccination, a mouse model sensitized with a recombinant antigen has been developed. Methods: BALB/c and C57BL/6 mice were injected intradermally with a 37–kD recombinant Ae. aegypti salivary allergen (rAed a 2) in the absence of adjuvant twice weekly for 8 weeks and then challenged twice with rAed a 2 at weeks 10 and 12. Serum rAed a 2–specific IgE, IgG1 and IgG2a were measured by ELISA. Intradermal tests were performed every 4 weeks. The binding capacity of rAed a 2–specific IgE to the native Aed a 2 was examined by immunoblotting. Results: In both strains, sensitization with rAed a 2 induced a significant increase in IgE and IgG1, but not IgG2a. In all sensitized mice, a positive immediate skin reaction was apparent, while delayed–type hypersensitivity reactions were not observed. BALB/c mice produced significantly higher levels of IgE and IgG1 and larger wheals than C57BL/6 mice. The IgE antibodies elicited by rAed a 2 bound to not only rAed a 2 but also its natural form in mosquito saliva. Conclusion: (1) Repeated injections of rAed a 2 without adjuvant induce predominant Th2–type responses in mice. (2) BALB/c mice are better responders for IgE production than C57BL/6 mice. (3) rAed a 2 has identical allergenicity to its natural form.
Role of Heme Oxygenase in Pulmonary Response to Antigen Challenge in Sensitized Rats in vivo
Authors: Jia Yu Xia , Sekizawa Kiyohisa , Okinaga Shoji , Lie Ria , Sasaki Hidetada
Published: April 2003

Heme oxygenase (HO) protects against oxidant–mediated lung injury. However, it is uncertain whether changes in HO activity modulate antigen–induced airway inflammation. We studied the effects of pretreatment with either hemoglobin, a HO inducer, or tin protoporphyrin (SnPP)–9, a specific HO inhibitor, on increases in pulmonary insufflation pressure (PIP) and plasma extravasation induced by intravenously injected ovalbumin (OA) antigen in rats sensitized to OA in vivo with Evans blue dye as a marker. Pretreatment with hemoglobin (300 mg/kg) significantly increased (p<0.01) and that with SnPP–9 (50 μmol/kg) significantly decreased (p<0.01) HO activity of the lung, but they failed to alter OA antigen (300 μg/kg)–induced increases in PIP. In contrast, hemoglobin pretreatment significantly decreased (p<0.01) and SnPP–9 pretreatment significantly increased (p<0.05) the leakage of dye induced by OA antigen in the trachea, main bronchi, and segmental bronchi. OA antigen–induced increases in plasma extravasation were also inhibited by superoxide dismutase (12,000 U/kg). These findings suggest the oxygen radicals are involved in increases in plasma extravasation induced by antigen challenge and HO protects against antigen–induced airway inflammation.
Hymenoptera Venom Allergy: Time Course of Specific IgE Concentrations during the First Weeks after a Sting
Authors: Rieger-Ziegler Verena , Rieger Edgar , Kränke Birger , Aberer Werner
Published: April 2003

Detection of IgE antibodies specific to honeybee or Vespula venoms is an important criterium firstly for the diagnosis of sensitization and secondly for the indication for a specific immunotherapy. Some authors recommend to postpone blood analysis after an insect sting for a certain time because circulating IgE antibodies might be consumed by the allergic reaction, which would result in a false–negative test outcome. We investigated IgE concentrations during the first weeks after an insect sting in 31 patients with an unequivocal history of an anaphylactic reaction after a honeybee (n = 13) or Vespula (n = 18) sting. Blood samples for analysis of specific IgE concentrations (CAP system, Pharmacia Diagnostics, Sweden) were collected within 2 weeks and 5±2 weeks after the insect sting. 12/13 patients with honeybee venom and 14/18 patients with Vespula venom sensitization had CAP classes 1 or higher within the first 2 weeks. Those 5 patients with CAP class 0 within the first 2 weeks had detectable IgE concentrations a few weeks later. We conclude that testing for specific IgE to hymenoptera venoms is in most cases useful even during the first 2 weeks after the hymenoptera sting. This allows early decisions on further diagnostic procedures and the therapeutic way to choose. Patients with no detectable IgE should, however, be retested after a few weeks.
Concepts of the Pathogenesis of Allergic Disease: Possible Roles of Epstein–Barr Virus Infection and Interleukin–2 Production
Authors: Okudaira Hirokazu , Mori Akio
Published: April 2003

The prevalence of atopic diseases in so–called developed countries has risen over the past several decades, which is too short a period for genetic changes to have taken place. Furthermore, the increase has occurred irrespective of race or geographical differences in the prevalence of atopic diseases. These facts strongly suggest that the increase is due not to genetic factor(s) but to environmental factor(s). In this article, previous investigations into the pathogenesis of allergic disease are reviewed. The roles of immune cells, cytokines, oncoproteins and viral infections are examined.
Systemic Immunological Changes Induced by Administration of Grass Pollen Allergens via the Oral Mucosa during Sublingual Immunotherapy
Authors: Fanta C. , Bohle B. , Hirt W. , Siemann U. , Horak F. , Kraft D. , Ebner H. , Ebner C.
Published: April 2003

Twenty–four patients suffering from grass pollen allergy underwent sublingual immunotherapy (SLIT) with standardized grass pollen extract for 1 year. In order to investigate immunological changes induced by the administration of allergens via the oral mucosa, the SLIT–spit method was applied. The cumulative dose of approximately 80 μg of major allergen (grass group 5 allergen), was relatively low. During the time of treatment, we could observe a significant increase in the levels of specific IgG and IgG4 antibodies. However, the titers of allergen–specific IgE antibodies showed a significant increase in the course of SLIT as well. Analyzing lymphoproliferative responses, a significant decrease in reactivity in response to stimulation with complete grass pollen extract (p = 0.001) and to recombinant Phl p 1 (a major allergen of timothy grass, p<0.001) could be observed, indicating the induction of immunological tolerance. Proliferative responses to a control antigen (tetanus toxoid) were not influenced by the treatment. At different time points during SLIT, allergen (Phl p 1)–specific T cell clones (TCC) were established from the peripheral blood of the patients. Cytokine production by allergen–stimulated T cells did not reveal any changes consistent with immune deviation, i.e. the ratio of Th1/Th2 TCC did not change during SLIT. In conclusion, we provide evidence that sublingual treatment leads to systemic changes in immunoreactivity to the administered allergen.
Immunogold Electron Microscopic Localization of the Cross–Reactive Two–EF–Hand Calcium–Binding Birch Pollen Allergen Bet v 4 in Dry and Rehydrated Birch Pollen
Authors: Grote Monika , Hayek Brigitte , Reichelt Rudolf , Kraft Dietrich , Valenta Rudolf
Published: April 2003

Background: Recently, a novel family of low–molecular–weight (8–9 kD), two–EF–hand calcium–binding proteins has been described as allergens in plant pollens. Approximately 10% of pollen–allergic patients have IgE antibodies which cross–react with the two–EF–hand allergens in tree, grass and weed pollens. The aim of the present study was to localize Bet v 4, the two–EF–hand allergen from birch, in mature, dry pollen and to study the release of this allergen after hydration of the pollen by immunogold electron microscopy. Methods: Using completely anhydrous fixation techniques in combination with immunogold electron microscopy, we localized Bet v 4 and, for control purposes, the major birch pollen allergen Bet v 1, in dry birch pollen as well as in pollen grains after different periods of hydration. Parallel with these morphological studies, we monitored the release of Bet v 4 and Bet v 1 into aqueous supernatants of hydrated birch pollen grains by immunoblotting. Results: Bet v 4 was found in the electron–dense cytosol, in particular between the vesicles and cisternae of the endoplasmic reticulum, inside mitochondria and in the vegetative as well as in the generative nucleus. Bet v 1 was localized in similar cellular compartments except for the mitochondria. After 30 s to 1 min of hydration, Bet v 4 migrated into the pollen exine and into the aqueous supernatants. Bet v 1 also moved out of the pollen grain, though not as quickly as Bet v 4. Conclusion: Bet v 4 represents an intracellular pollen protein which, following hydration of pollen grains, rapidly migrates to the pollen surface (exine) and is washed out. This behavior explains how Bet v 4, being primarily an intracellular pollen protein, becomes available to sensitize patients.
Heparin, Derived from the Mast Cells of Human Lungs Is Responsible for the Generation of Kinins in Allergic Reactions due to the Activation of the Contact System
Authors: Noga Oliver , Brunnée Tilo , Schäper Christoph , Kunkel Gert
Published: April 2003

Background: In a recent study mast cell heparin proteoglycan (HepPG) of a cell line derived from a mouse mastocytoma was isolated. Glycosaminoglycans proved to be an initiating surface for starting contact activation and could explain kinin generation present in allergic reactions. It is the aim of the present study to prove that HepPG or glycosaminoglycan derived from human mast cells is also capable of acting as a physiologic macromolecule and to induce contact activation. Methods: HepPG molecules were isolated by anionic column chromotography. Their ability to accelerate reciprocal activation of factor XII was investigated by spectrophotometry. The anticoagulant effect was demonstrated by an increase in partial thromboplastin time. HPLC was performed to correlate these effects with molecular weight (MW). Results: The isolated heparin showed high contact–activating and anticoagulant potency. Both actions were suppressed by incubation with heparinase I. The maximum contact activation peak appeared at a lower MW than the anticoagulant effect. Conclusion: These in vitro results explain the results of in vivo allergen challenge studies where a high degree of kinin generation occurs. Heparin derived from human mast cells therefore seems to represent the physiological macromolecule capable of activating the contact system and could be a missing link between cellular and humoral responses in allergic reactions.
Genetic and Environmental Factors Contributing to the Onset of Allergic Disorders
Authors: Parronchi P. , Brugnolo F. , Sampognaro S. , Maggi E.
Published: April 2003

Evidence has been accumulated to suggest that allergen-reactive Th2 cells play a triggering role in the activation and/or recruitment of IgE antibody-producing B cells, mast cells and eosinophils, the cellular triad involved in allergic inflammation. Recently, chemokines and chemokine receptors involved in such Th2-type response have been also defined. Th2 cells represent the polarized arm of the effector-specific responses that contribute to the protection against gastrointestinal nematodes and act as regulatory cells for chronic and/or excessive Th1-mediated responses. Th2 cells are generated from precursor naive Th cells when they encounter the specific antigen in an IL-4-containing microenvironment. The question of how these Th2 cells are selected in atopic patients is also unclear. Both the nature of the T cell receptor signalling provided by the allergen peptide ligand and a disregulation of IL-4 production likely concur to determine the Th2 profile of allergen-specific Th cells, but the genetic unbalanced IL-4 production is certainly overwhelming. Some gene products selectively expressed in Th2 cells or selectively controlling the expression of IL-4 have recently been described. These findings allow to suggest that the upregulation of genes controlling IL-4 expression and/or abnormalities of regulatory mechanisms of Th2 development and/or function may be responsible for Th2 responses against allergens in atopic people. The increasing prevalence of allergy in developed countries suggests that environmental factors acting either before or after birth also contribute to regulate the development of Th2 cells and/or their function. The reduction of infectious diseases in early life due to increasing vaccinations, antimicrobial treatments as well as changed lifestyle are certainly important in influencing the individual outcome in the Th response to ubiquitous allergens. Moreover, the recent evidence that bacterial DNA or oligodeoxynucleotides containing unmethylated ‘CpG motifs’ promote the development of Th1 cells via the production of immunomodulatory cytokines (namely IL-12, IL-18 and IFNs) by professional antigen-presenting cells confirms previous epidemiological data. The new insight into the pathophysiology of T cell responses in atopic diseases provides exciting opportunities for the development of novel immunotherapeutic strategies.
Characterization of T Cell Subpopulations Involved in the Pathogenesis of Asthma and Allergic Diseases
Authors: Yssel Hans , Groux Hervé
Published: April 2003

Allergic asthma is a complex and heterogeneous disease which is characterized by intermittent reversible airway obstruction, chronic inflammation of the airways, bronchial hyperreactivity and an infiltration of lymphocytes and eosinophils into the airway submucosa. Animal models and clinical studies in humans have indicated an important role for T helper type 2 lymphocytes, producing IL-4, IL-5 and IL-13, in the pathogenesis of this disorder. However, although IL-4 and IL-13 have strong anti-inflammatory properties, the physiologic anti-inflammatory Th2 response does not seem to be operational in allergic asthma. Moreover, the induction of a Th1 response seems to aggravate, rather than ameliorate, its inflammatory character. This article will focus on the involvement of T lymphocyte subpopulations in the pathogenesis of allergic asthma and allergic diseases. In addition, a potential role of the subpopulation(s) of T regulatory cells in the induction and/or maintaince of the disease process will be discussed.
Sodium Salicylate-Induced Apoptosis of Human Peripheral Blood Eosinophils Is Independent of the Activation of c-Jun N-Terminal Kinase and p38 Mitogen-Activated Protein Kinase
Authors: Wong C.K. , Zhang J.P. , Lam C.W.K. , Ho C.Y. , Hjelm N.M.
Published: April 2003

Background: It has been shown that the inhibition of eosinophilic apoptosis is an important mechanism for the development of blood and tissue eosinophilia in allergic diseases. Considerable attention has recently been focused on the role played by different intracellular kinase cascades in the control of apoptosis. In the present study, we investigated the effect of sodium salicylate (NaSal), a nonsteroidal anti-inflammatory drug, on mitogen-activated protein kinases (MAPK) and apoptosis of human eosinophils. Methods: Human blood eosinophils were purified from buffy coat. NaSal-induced apoptosis of eosinophils was assessed by morphological changes and Annexin-V binding assay. Changes of MAPK activity upon treatment with NaSal were measured by kinase activity assay and Western blot. Results: NaSal could induce apoptosis of human blood eosinophils in a dose- and time-dependent manner. It could also activate c-Jun N-terminal kinase (JNK) and p38 MAPK but not extracellular signal-regulated protein kinase (ERK) activity within 1 h. Pretreatment of eosinophils with p38 MAPK and JNK anti-sense (AS) phosphorothioate oligodeoxynucleotides (ODN) or specific p38 MAPK inhibitor SB 203580 did not have any significant effect on NaSal-induced apoptosis. However, ERK AS ODNs could trigger the apoptosis of normal eosinophils. Conclusion: There is no direct relationship between the activation of JNK and p38 MAPK pathways and NaSal-induced apoptosis in human peripheral blood eosinophils.
Once–Daily Theophylline Reduces Serum Eosinophil Cationic Protein and Eosinophil Levels in Induced Sputum of Asthmatics
Authors: Aizawa H. , Iwanaga T. , Inoue H. , Takata S. , Matsumoto K. , Takahashi N. , Yoshida M. , Hara N.
Published: April 2003

Background: Because eosinophilic airway inflammation is a characteristic feature of bronchial asthma, the treatment of airway inflammation is important in the management of asthma. Theophylline has been reported to reduce airway inflammation, in addition to its well–known bronchodilating effect. Objective: In order to evaluate the effects of theophylline on airway inflammation, we investigated 48 subjects with mild and moderate asthma. Methods: The patients were randomly divided into two groups, with or without theophylline treatment (control n = 24; theophylline, n = 24). We examined the level of serum eosinophil cationic protein (ECP), induced sputum samples, and peak expiratory flow (PEF) and obtained spirograms before and after 4 weeks of treatment with once–daily theophylline (200–600 mg/day) of subjects with mild or moderate asthma. Results: Theophylline significantly increased morning and evening PEF and significantly decreased the diurnal variation of PEF. After treatment with theophylline, both serum ECP and the percentage of eosinophils in induced sputum were significantly decreased. In contrast, peripheral blood eosinophil count was unchanged after treatment with theophylline. Conclusions: These findings suggest that theophylline reduces airway inflammation and the severity of asthma, presumably via suppression of both eosinophil activity and subsequent eosinophil infiltration of the airways.
Increased Eosinophil Cation Protein Level in Sensitized Nonasthmatics Is Linked to Subsequent Hyperresponsiveness to Methacholine
Authors: Rasmussen F. , Lambrechtsen J. , Siersted H.C. , Hansen H.S. , Hansen N.C.G.
Published: April 2003

Background: Increased levels of eosinophil cation protein (ECP) in sensitized subjects may reflect early stages of an ongoing inflammatory process and therefore precede asthma and bronchial hyperreactivity. Aim: To study whether nonasthmatic subjects with sensitization to allergens and increased ECP levels are at a higher risk for subsequent increased bronchial reactivity compared with sensitized nonasthmatics with normal ECP levels. Methods: A prospective study of 240 schoolchildren with a mean age of 13.9 years (range: 12.6–15.9) who were followed up after 6.3 years. Bronchial reactivity was assessed by methacholine provocation testing. Sensitization was defined by one or more positive reactions (>3 mm wheal) to 10 common aeroallergens by skin prick testing. Increased ECP was defined as values above 20 μg/l. This separated the subjects into four categories: group 1: healthy controls without sensitization (n = 147); group 2: sensitized subjects with a serum ECP below 20 μg/l (n = 55); group 3: sensitized subjects with an ECP level at or above 20 μg/l (n = 16), and group 4: all asthmatics (n = 22). Results: Bronchial reactivity was similar in subjects of groups 2 and 3 at baseline (p = 0.8). Six years later, subjects from group 3 were more responsive to methacholine compared with subjects from group 2 (median: 12.7 versus 20.5 μmol; p < 0.05). In a logistic regression with hyperresponsiveness to methacholine at follow-up as dependent variable, the odds ratios (OR) for the groups were, with group 1 as reference: group 2: OR = 2.2 (0.8–6.6: p = 0.2), group 3: 5.9 (1.6–21.7: p < 0.01). Conclusion: Subjects with sensitization and increased ECP levels are subsequently more airway-responsive to methacholine compared with sensitized subjects with normal ECP levels. This supports the hypothesis that sensitization is linked to increased bronchial reactivity through a process in which markers of inflammation are involved.
Differential Regulation of Activation-Induced Cell Death in Individual Human T Cell Clones
Authors: Janssen Ottmar , Stocker Andreas , Sanzenbacher Ralf , Oberg Hans-Heinrich , Siddiqi Mushtaq A. , Kabelitz Dieter
Published: April 2003

Background: Restimulation of T lymphocytes via the TCR/CD3 complex can result in CD95/CD95L-dependent activation-induced cell death (AICD). Although the correlation of AICD sensitivity to the T helper 1 phenotype was confirmed in different studies, the underlying mechanism is still debated. Thus, it has been suggested that in Th2 cells, AICD resistance is controlled by a TCR-induced upregulation of the CD95-associated inhibitory phosphatase, FAP-1. We and others demonstrated that AICD resistance is associated with a reduced surface expression of CD95L upon restimulation. Methods: Utilizing RT-PCR, Western blotting and flow cytometry, we analyzed time-dependent changes in levels of CD95L mRNA, cytosolic protein and surface expression in five long-term human T cell clones and polarized helper populations. Results: We confirm that the inducible CD95L surface expression is lower or absent in all tested AICD-resistant clones as compared to sensitive cells. It is of interest that striking differences with respect to the activation-dependent inducibility of CD95L mRNA expression in individual resistant clones were observed. In addition, alterations in the expression of the inhibitory phosphatase FAP-1 or TCR-dependent changes in CD95 sensitivity in AICD-resistant clones could be ruled out as a mechanism for AICD resistance of human T cell clones. Conclusions: (1) The data presented strongly support the previous notion that AICD resistance of human T cell clones is mainly regulated by a differential expression of CD95L. (2) Differential expression of CD95L on individual resistant clones results from a lack of mRNA induction in one set and from a markedly decreased surface expression of translated protein in another set of clones.
Identification of Hev b1 in Natural Latex Mattresses
Authors: Chardin H. , Chen Z. , Raulf-Heimsoth M. , Mayer C. , Sénéchal H. , Desvaux F.X. , Peltre G.
Published: April 2003

Background: Type 1 hypersensitivity to natural rubber latex proteins is a well-recognized health problem. Recent data have shown that allergens can be extracted from natural latex mattresses. As Hev b 1 (rubber elongation factor) and Hev b 6.02 (hevein) were described as major allergens, the present work was carried out to evaluate their presence in latex mattresses as well as in latex gloves. Methods: Extracted proteins from latex mattresses and gloves were separated by SDS-PAGE or two-dimensional gel electrophoresis, transferred onto nitrocellulose and detected with monoclonal antibodies specific for Hev b 1 and Hev b 6.02. Results: The results showed that various forms of Hev b 1, as well as degradation products of Hev b 1 were detected in latex mattresses and gloves, whereas Hev b 6.02 was not detected either in mattresses or in gloves. In a standardized latex extract, Hev b 1 and Hev b 6.01 (prohevein) were identified by the monoclonal antibodies. Conclusion: The fact that only Hev b 1 was detected by immunoblot in latex articles indicates that Hev b 1 may be the last protein to be washed out of latex products and that the Hev b 1 content may be used as a criterion for the estimation of the allergenicity of the latex products.
Superantigen-Induced T Cell Death by Apoptosis: Analysis on a Single Cell Level and Effect of IFN-γ and IL-4 Treatment
Authors: Weber Anne K. , Wahn Ulrich , Renz Harald
Published: April 2003

Background: A role of bacterial superantigens in several chronic inflammatory diseases has previously been proposed. Many of these diseases are associated with an imbalance of the T helper cell subsets and their cytokine production. Methods: Peripheral blood mononuclear cells from healthy donors were incubated with various concentrations of staphylococcal enterotoxin B (SEB) with or without IL-4 or IFN-γ. After different time points cell activation, proliferation, Fas expression, cytokine release and cell death via apoptosis were detected. Results: SEB treatment resulted in sequential T cell activation, proliferation, Fas expression, cytokine release, subsequently followed by cell death via apoptosis in a dose- and time-dependent manner. This biphasic effect occurred preferentially in SEB-responsive cells represented by the expression of Vβ3 and Vβ12 on T cells. A strong relationship between T cell activation and apoptosis was observed. The amplitude between these events increased with the dose of SEB. The highest rate of apoptotic T cells was observed at a dose of 1,000 ng/ml SEB. Addition of IFN-γ to SEB-treated cells significantly reduced the rate of apoptotic cells, whereas IL-4 prevented apoptosis only in SEB-untreated cells. Conclusion: These results support the concept that the dose of superantigen exposure determines the rate of T cell proliferation and subsequent cell death. This T cell immune response is modulated by the presence and the type of cytokines.
Mediation of the Immunomodulatory Effect of β-Estradiol on Inflammatory Responses by Inhibition of Recruitment and Activation of Inflammatory Cells and Their Gene Expression of TNF-α and IFN-γ
Authors: Salem Mohamed Labib , Hossain Mohammad Sohrab , Nomoto Kikuo
Published: April 2003

Background: Estrogen has long been reported to show immunomodulatory effects on immune responses, yet, its specific anti-inflammatory mechanism is not clear. Methods: In this study, we analyzed the effects of β-estradiol (E2), at its contraceptive dose, on both T cell-independent and T cell-dependent inflammations, and the associated immune mechanism, in female mice. The T cell-independent inflammation was locally induced either with an intradermal injection of olive oil in the footpad, or by an intraperitoneal injection of proteose peptone (PP). The T cell-dependent inflammation was induced by an intraperitoneal injection of the purified protein derivatives (PPD). Results: While E2 inhibited olive oil-induced inflammation as monitored by the decrease in footpad swelling, it did not affect the gene expression of monocyte chemoattractant protein-1 and IL-6 by cells at the inflammatory locus. E2 also inhibited PP-induced inflammation as monitored by the decrease in the number of inflammatory peritoneal exudate cells (PEC) coinciding with a marked decrease in the number of macrophages and granulocytes (Gr. 1+). While E2 did not affect the gene expression of monocyte chemoattractant protein-1 and IL-6 by PP-elicited PEC, it decreased both gene expression and production of TNF-α. E2 also decreased the number of cells expressing the lymphocyte function-activated protein-1 in PP-elicited PEC, but not for CD62L. In purified protein derivative-induced T cell-dependent inflammation, E2 decreased the total cellularity of PEC and the relative numbers of CD3+ and CD4+ T cells, and the number of cells expressing the lymphocyte activation markers CD40, CD44, CD69 and IL-2Rα in PEC. Furthermore, while E2 did not affect the gene expression of the early T lymphocyte activation protein-1 by PEC, it decreased the gene expression of INF-γ. Conclusion: Collectively, the results suggest that E2-mediated inhibition of inflammatory responses may be due to a combination of suppression of homing and activation of inflammatory cells and their production of TNF-α and IFN-γ.
Allergy and the Gut
Authors: Bischoff Stephan C. , Mayer Julika H. , Manns Michael P.
Published: April 2003

There have frequently been doubts as to the relevance of food allergy, in particular as far as the involvement of the intestinal tract is concerned. Several studies, however, have confirmed the existence of allergic reactions in the gut, with an estimated prevalence of about 1–2% in adults. Clinical symptoms are unspecific and include nausea, vomiting, abdominal pain, cramping and diarrhea. Intestinal mast cells, as well as intestinal eosinophils, have been shown to be involved in the pathogenesis of food-allergy-related enteropathy. In addition to classical IgE-dependent degranulation, further agonists have been demonstrated for mast cell activation, for example IL-4. The methods used to confirm the diagnosis of instestinal allergy are still insufficient. Until now, blinded oral challenge procedures with food antigens have been accepted as the ‘gold standard’ in diagnosing food allergy, although these tests have practical problems. Therefore, new test systems have been developed, such as endoscopic provocation tests, that may improve diagnostic procedures. Elimination diet still presents the main basis of therapy. Aspects to be focused on in the future are the role fo IgE-independent allergic mechanisms in intestinal allergy, the impact of cross-reactivity with other allergens and the relationship to other inflammatory bowel diseases such as Crohn’s disease, ulcerative colitis, celiac disease and irritable bowel syndrome.
Purification and Immunobiochemical Characterization of Folding Variants of the Recombinant Major Wasp Allergen Ves v 5 (Antigen 5)
Authors: Suck Roland , Weber Bernhard , Kahlert Helga , Hagen Susan , Cromwell Oliver , Fiebig Helmut
Published: April 2003

Background: Antigen 5 is one of three major allergens in wasp venoms, but unlike phospholipase A1 and hyaluronidase, both of which are enzymes, its biological function is unknown. The cDNA coding for this allergen has been isolated and used for recombinant expression. Thorough analysis of the expression product is essential in order to evaluate the usefulness for in vivo or in vitro application. Objective: In this study, folding variants of the recombinant major allergen Ves v 5 from Vespula vulgaris were immunologically and biochemically investigated in order to determine their possible applicability for diagnostic or therapeutic purposes. Method: The cDNA encoding Ves v 5 was cloned into the expression vector pSE420 which generates recombinant products lacking a tag sequence. After expression, inclusion bodies were purified, subsequently denatured and dialyzed against different solutions. The structural properties of soluble proteins were analyzed by size exclusion chromatography, non-reducing SDS-PAGE, native PAGE, N-terminal sequencing, proteolytic digestion and ion exchange chromatography. Immunological investigations were performed by using different monoclonal antibodies (mAbs) specific for Ves v 5 and IgE from patients allergic to wasp venom allergens. Results: After dialysis, soluble monomeric recombinant Ves v 5 was more than 95% pure in each case. Using different dialysis solutions, clearly distinguishable folding variants were obtained. In one case, the recombinant allergen was comparable with the natural counterpart in respect of migration in non-reducing SDS-PAGE, native PAGE and IgE reactivity. This variant reacted with two different Ves v 5-specific mAbs and produced a stable fragment after proteolytic digestion. Elution from a cation exchange chromatography column was achieved with 320 mM NaCl. In two other cases, folding variants exhibited a different migration behavior in SDS-PAGE and native PAGE compared with the natural allergen. Also, the mAb 1E11 recognized none of these variants since it presumably detected a conformational epitope. Moreover, the IgE reactivity was clearly reduced and proteolytic digestion effected almost complete degradation. These variants eluted from the cation exchange column with 400 mM NaCl. Conclusion: Defined folding strategies resulted in both soluble misfolded variants with reduced IgE reactivity, potentially suitable for immunotherapy, and natural-like folded variants for diagnosis.
Regulation of Specific Immune Responses by Chemical and Structural Modifications of Allergens
Authors: Akdis Cezmi A. , Blaser Kurt
Published: April 2003

Specific immunotherapy (SIT) is an efficient treatment of allergic diseases to defined allergens. Despite being used in clinical practice since early in this century, more rational and safer regimens are required, because SIT is faced with the risk of anaphylaxis and standardization problems of allergen-extract-based treatments. A better understanding of the pathogenesis of allergy and of the mechanisms of SIT has led to various approaches to overcome these problems. Knowledge of the influence of IgE-facilitated antigen presentation on allergen-specific Th2 responses increased the efforts to generate non-IgE-binding allergens. The current principal approach to allergen modification is to modify B cell epitopes in order to prevent IgE binding and effector cell cross-linking while preserving T cell epitopes to retain the capacity of inducing tolerance. In this way, the modified allergen will be directed to T cells by a phagocytosis/pinocytosis-mediated antigen uptake mechanism, bypassing IgE cross-linking and IgE-dependent antigen presentation. Accordingly, a differential regulation of allergen-specific T cell cytokine patterns and IgE:IgG production was demonstrated by modifications of the three-dimensional structure of allergens because of linearity in T cell epitopes and conformation dependence in B cell epitopes. In this context, chemically modified allergen extracts with low IgE-binding capacity have been developed to reduce anaphylactic side effects since the early 1980s. The progress of recombinant techniques for producing allergens and allergen derivatives has led to a dramatic improvement in the ability of developing novel vaccines for the treatment of allergy. This has enabled mutation or deletion of decisive amino acids in B cell epitopes and fractionation or oligomerization of allergens by genetic engineering as fruitful approaches to generate hypoallergenic vaccines. Moreover, non-IgE-binding short T cell epitope peptides and single-amino-acid-altered peptide ligands represent potential candidates for future SIT.
Antigen Recognition by Human γδ T Lymphocytes
Authors: Kabelitz Dieter , Glatzel Andrea , Wesch Daniela
Published: April 2003

Mature T lymphocytes carrying the conventional αβ T cell receptor (TCR) recognize peptide antigens in the context of MHC class I (CD8+) and MHC class II (CD4+) antigens, respectively. In striking contrast, human γδ T lymphocytes recognize unconventional antigens via their heterodimeric TCR in a non-MHC-restricted fashion. In this brief review, we discuss recent progress in the identification of ligands that are specifically recognized by human γδ T cells. It appears that γδ T cells have evolved to supplement the cellular immune response towards antigens that are not seen by αβ T lymphocytes.
The Immune Response in Adenoids and Tonsils
Authors: van Kempen M.J.P. , Rijkers G.T. , van Cauwenberge P.B.
Published: April 2003

The adenoid and tonsils are lymphoid tissues located in the pharynx that play an important role in host defense against invading antigens of the upper respiratory tract. Histologically, these structures consist of four well-defined microcompartments which all participate in the immune response: the cryptepithelium, the follicular germinal center with the mantle zone and interfollicular area. With the uptake of antigen by M-cells present in the cryptepithelium a process is initiated which ultimately results in the generation and dissemination of antigen-specific memory and mainly dimeric IgA-producing effector B-lymphocytes. This process requires successful cognate interactions between antigen-presenting cells and lymphocytes and mutually between lymphocytes, which depend not only on antigen-specific signals but also on the expression of various complementary adhesion and costimulatory molecules.
Inhibitory Effects of Human Neutrophil Functions by the 45-kD Glycoprotein Derived from the Parasitic Nematode Trichinella spiralis
Authors: Bruschi Fabrizio , Carulli Giovanni , Azzarà Antonio , Homan Wieger , Minnucci Sistina , Rizzuti-Gullaci Angela , Sbrana Silverio , Angiolini Claudia
Published: April 2003

Aim: We evaluated the effect of the 45-kD protein of Trichinella spiralis (gp45), purified by affinity chromatography, on random migration and chemotaxis, the oxidative metabolism of human neutrophils and on the CD11b upregulation induced by formyl-methionyl-leucyl-phenylalanine (f-MLP). Methods: Donor neutrophils incubated with different amounts of gp45 (0.5, 1, 1.5, 2 μg/ml) or buffer and the random migration and chemotaxis, evaluated by means of a special technique of image analysis, and the chemiluminescence response to f-MLP or phorbol myristate acetate (PMA) were analyzed. The effect on CD11b upregulation was assessed incubating cells with the protein, when activating them with f-MLP. Results: The results showed that gp45 inhibited both random and stimulated migrations, and reduced the response to f-MLP and PMA. Furthermore, gp45 significantly reduced the upregulation of the CD11b induced by f-MLP. Conclusion: The results show that gp45 inhibits PMN in different functions, suggesting an anti-inflammatory action.