Laboratory animal allergens

Allergic sensitivity to laboratory animals can pose a significant occupational hazard to anyone with regular animal contact. Reactions to mice and rats are most common although all furred animals produce allergens that can lead to sensitization and disease. Most of the relevant allergens of laboratory animals have been defined and characterized, which has revealed that these allergens are typically small, acidic glycoproteins and that many of them are members of a superfamily of extracellular proteins called lipocalins. In addition to understanding their molecular characteristics, the identification of these allergens has also made it possible to measure their distribution in laboratory environments and to relate exposure levels to sensitization and symptoms. These studies have shown that the major laboratory animal allergens are carried on small particles that are both capable of remaining airborne for extended periods and penetrating into the lower airways of exposed workers. These advances in the understanding of these important occupational allergens will allow for the development of better methods of diagnosis and avoidance for affected workers and others who may be at risk for future difficulties.     

Carbonylation of milk powder proteins as a consequence of processing conditions

During industrial treatments, milk proteins could be oxidatively modified, thus leading to the formation of modified/oxidised amino acid residues. The apparition of such modified residues may contribute to the formation of new immunologically reactive structures. Some of these adducts could, in an advanced stage, lead to cross-linked protein species whose proteolytic susceptibility would be drastically decreased. Such protein species, that are resistant to digestion, could also constitute major food allergens. Therefore, these oxidative protein modifications tend to increase the natural allergenicity of milk proteins. For these reasons, monitoring milk protein oxidative modifications could be very useful regarding both product quality and allergenicity issues. In the present paper, we highlight, using different analytical approaches, the preferential carbonylation of beta-lactoglobulin (beta-Lg) during industrial treatments of milk. This result is particularly interesting since native beta-Lg represents one of the major milk allergens.     

Allergen exposure and control

Allergens produced by the house dust mites (family Pyroglyphidae) are probably the single most important allergens associated with asthma world wide. If exposure to these allergens in houses could be sufficiently reduced, then asthma symptoms may be markedly reduced and even prevented from being initiated. Only about half of the many attempts to reduce mite allergens in houses have shown any clinical benefit. One reason may be that exposure was not reduced enough-however exposure to mite allergens has never been measured in any trial. This review summarises previous allergen control trials and then provides an outline of allergen exposure, including the nature of exposure, the analytical methods available and the recognised risks of allergen exposure. This provides a perspective to evaluate the individual methods used to kill mites and to reduce exposure to the allergens. The object is to provide a framework to improve and develop allergen avoidance as an effective component of asthma management.The author is supported by a project grant from the National Heath and Medical Research Council of Australia.     

Study of allergens from vegetables, fruits and berries

Different methods of obtaining allergens from vegetables, fruit and berries have been evaluated. Two technologies for obtaining food allergens have been considered; of these, Coca's method has proved to be more technological, economical and less labor-consuming. The newly developed technology has made it possible to obtain stable, safe and specifically active allergens which may be used for diagnosing food allergy.     

Immunological identification and characterization of individual food allergens

Food allergies of type-I-allergy are immunoglobulin E (IgE) mediated and caused by certain proteins or glycoproteins, which are called food allergens. An analytical marker of allergens is the IgE-reactivity to these substances. Normally food allergens are minor components in allergenic source material, which consist of a huge number of chemical different substances. Thus allergen extraction, separation and immunological detection methods are described which identify and characterize individual food allergens by a minimum of manipulation. Favoured separation methods of allergenic extracts are electrophoretic ones allowing the combination of highly resolved protein separations with immunological detection methods subsumed by the term immunoblotting. These techniques are a useful basis to characterize allergens by chemical methods. Once the primary protein structure of a food allergen is established, the way is cleared for the identification of epitopes. Epitopes are immunological detectable parts of a protein or glycoprotein generating the interface between chemical structure and immune-system. The nature of epitopes may differ, for instance, can be conformational, continuous, or built up by glycoconjugates, which determine the stability of food allergens, especially in the case of food processing. Progress in identification and characterization of food allergens will improve diagnostics and therapy of food allergy.     

Hypoallergenic derivatives of major grass pollen allergens for allergy vaccination

Grass pollen-induced hay-fever and allergic asthma represent a major health problem in industrialized countries. Whereas the symptoms of these allergic conditions can be controlled by pharmacotherapy, specific immunotherapy vaccination is the only causative approach towards the treatment of these type 1 allergies. Specific immunotherapy is based on administration of increasing amounts of the disease-causing allergens in the form of allergen-containing extracts. However, the extracts used for immunotherapy consist of allergenic and non-allergenic components and may induce severe anaphylactic side-effects upon therapeutic administration. With recent developments in molecular biology of pollen allergens it has become feasible to produce modified hypoallergenic derivatives of recombinant allergens with abrogated or greatly reduced likelihood of anaphylactic side-effects as compared to extract-based treatments. We have demonstrated this concept through reducing the anaphylactic potential of major rye grass pollen allergens by introducing a few point mutations which leave the overall structural fold of the molecule unaltered. These modified forms are expected to make allergen-specific immunotherapy more widely used in the future.     

Computational analysis of the relationship between allergenicity and digestibility of allergenic proteins in simulated gastric fluid

Background  

Safety assessment of genetically modified (GM) food, with regard to allergenic potential of transgene-encoded xenoproteins, typically involves several different methods, evaluation by digestibility being one thereof. However, there are still debates about whether the allergenicity of food allergens is related to their resistance to digestion by the gastric fluid. The disagreements may in part stem from classification of allergens only by their sources, which we believe is inadequate, and the difficulties in achieving identical experimental conditions for studying digestion by simulated gastric fluid (SGF) so that results can be compared. Here, we reclassify allergenic food allergens into alimentary canal-sensitized (ACS) and non-alimentary canal-sensitized (NACS) allergens and use a computational model that simulates gastric fluid digestion to analyze the digestibilities of these two types.     

Major house dust mite allergens Dermatophagoides pteronyssinus 1 and Dermatophagoides farinae 1 degrade and inactivate lung surfactant proteins A and D

Lung surfactant proteins (SP) A and D are calcium-dependent carbohydrate-binding proteins. In addition to playing multiple roles in innate immune defense such as bacterial aggregation and modulation of leukocyte function, SP-A and SP-D have also been implicated in the allergic response. They interact with a wide range of inhaled allergens, competing with their binding to cell-sequestered IgE resulting in inhibition of mast cell degranulation, and exogenous administration of SP-A and SP-D diminishes allergic hypersensitivity in vivo. House dust mite allergens are a major cause of allergic asthma in the western world, and here we confirm the interaction of SP-A and SP-D with two major mite allergens, Dermatophagoides pteronyssinus 1 and Dermatophagoides farinae 1, and show that the cysteine protease activity of these allergens results in the degradation of SP-A and SP-D under physiological conditions, with multiple sites of cleavage. A recombinant fragment of SP-D that is effective in diminishing allergic hypersensitivity in mouse models of dust mite allergy was more susceptible to degradation than the native full-length protein. Degradation was enhanced in the absence of calcium, with different sites of cleavage, indicating that the calcium associated with SP-A and SP-D influences accessibility to the allergens. Degradation of SP-A and SP-D was associated with diminished binding to carbohydrates and to D. pteronyssinus 1 itself and diminished capacity to agglutinate bacteria. Thus, the degradation and consequent inactivation of SP-A and SP-D may be a novel mechanism to account for the potent allergenicity of these common dust mite allergens.     

Lipocalins as allergens

The term allergy refers to clinical conditions caused by an inappropriate immune response to innocuous proteins in genetically predisposed persons. Allergens of animal origin are responsible for a significant proportion of allergies. In recent years, it has become evident that practically all respiratory animal allergens characterized at the molecular level belong to the lipocalin family of proteins. The current list comprises the major allergens of horse, cow, dog, mouse, rat and cockroach as well as beta-lactoglobulin of cow's milk. While the molecular structure of all these allergens is known, far less information is available regarding their immunological characteristics. Knowing the way the immune system recognizes these allergens and reacts to them might, however, be the key for discovering the common denominator of the allergenicity of lipocalins. The human body contains numerous endogenous lipocalins, and the immune system has to adapt to their presence. We have proposed that under these conditions the immune response against the lipocalin allergens which are structurally related to endogenous lipocalins might be the pathway to allergy in genetically predisposed persons. The same might well apply also to other allergens with homologous endogenous counterparts.     

植物过敏性蛋白质及其生物学功能

在引起I型超敏反应的变应原中 ,植物的花粉、果实和汁液可以分别作为吸入性变应原 (inhalentallergen)、食入性变应原 (ingestentallergen)、接触性变应原 (contactentallergen)使过敏者患上鼻炎、哮喘、枯草热等疾病。而其中引起这些超敏反应的植物类蛋白质本身在植物体内亦行使着特定的生物学功能。对这些植物类过敏性蛋白质的研究不仅在植物学本身研究中具有一定意义 ,同时在变态反应性疾病的免疫治疗中亦具有重要的应用价值。目前 ,这类涉及植物学、免疫学和变态反应学的研究逐渐形成了一个新的交叉研究领域。     

Early Exposure to Respiratory Allergens by Placental Transfer and Breastfeeding

The relationship between allergen exposure and the onset of or protection from allergic diseases remains unclear. Many factors could be related to immunological responses, such as the age when the exposure occurs, type of allergen, timing, dose, and allergen route. In this study, we investigated whether exposure to respiratory allergens could occur in pregnancy or early life. In particular, we assessed whether Der p 1 and Blo t 5, as well as specific antibodies against these allergens, could be detected in 90 paired cord blood and colostrum samples. Der p 1 was detected in 58.6% of colostrum and 29% of cord blood samples, whereas Blot 5 was positive in 41.3% and 9.6% of the samples, respectively. Similar to specific IgA, which could be detected in all samples for both mites, specific IgG was found in a high number of colostrum samples, 93.5% and 94.8% for Dp and Bt, respectively. Although allergens were not detected in all cord blood samples, a high percentage of them (≥95%) were positive for specific IgM to both mites in cord blood samples, suggesting that neonates can be exposed and sensitized to airborne allergens during pregnancy. Many studies have attempted to correlate allergen exposure or its prevention in early infancy with the onset of or protection from allergic diseases. However, conflicting and inconsistent data do not show a clear correlation with or suggest a way to prevent allergen sensitization. Nevertheless, these unconvincing results could be better understood if the relationship with many aspects of allergen exposure after pregnancy could be clarified. Thus, it is necessary to address basic issues related to allergen exposure, including the development of reproducible, standardized and reliable methods, and to determine how and where the exposure occurs.     

Production of Recombinant Peanut Allergen Ara h 2 using <Emphasis Type="Italic">Lactococcus lactis</Emphasis>

Background  

Natural allergen sources can supply large quantities of authentic allergen mixtures for use as immunotherapeutics. However, such extracts are complex, difficult to define, vary from batch to batch, which may lead to unpredictable efficacy and/or unacceptable levels of side effects. The use of recombinant expression systems for allergen production can alleviate some of these issues. Several allergens have been tested in high-level expression systems and in most cases show immunereactivity comparable to their natural counterparts. The gram positive lactic acid bacterium Lactococcus lactis is an attractive microorganism for use in the production of protein therapeutics. L. lactis is considered food grade, free of endotoxins, and is able to secrete the heterologous product together with few other native proteins. Hypersensitivity to peanut represents a serious allergic problem. Some of the major allergens in peanut have been described. However, for therapeutic usage more information about the individual allergenic components is needed. In this paper we report recombinant production of the Ara h 2 peanut allergen using L. lactis.     

Physical Properties of Intact Proteins May Predict Allergenicity or Lack Thereof

Background

Predicting the allergenicity of proteins is challenging. We considered the possibility that the properties of the intact protein that may alter the likelihood of being taken up by antigen presenting cells, may be useful adjuncts in predicting allergens and non-allergens in silico. It has been shown that negatively charged acidic proteins are preferentially processed by dendritic cells.

Methodology

Datasets (aeroallergen, food-allergen and non-allergen) for in-silico study were obtained from public databases. Isoelectric point (pI), net charge, and electrostatic potential (EP) were calculated from the protein sequence (for pI and net charge) or predicted structure (for EP).

Result

Allergens and non allergens differed significantly in pI, net charge, and EP (p<0.0001). Cluster analysis based on these parameters resulted in well defined clusters. Non-allergens were characterized by neutral to basic pI (mean±SE, 7.6±0.16) and positive charge. In contrast allergens were acidic (5.7±0.15) and negatively charged. Surface electrostatic potentials calculated from predicted structures were mostly negative for allergens and mostly positive for non-allergens. The classification accuracy for non-allergens was superior to that for allergens. Thus neutral to basic pI, positive charge, and positive electrostatic potentials characterize non-allergens, and seem rare in allergens (p<0.0001). It may be possible to predict reduced likelihood of allergenicity in such proteins, but this needs to be prospectively validated.     

An immunoglobulin-like fold in a major plant allergen: the solution structure of Phl p 2 from timothy grass pollen.

BACKGROUND: Grass pollen allergens are the most important and widespread elicitors of pollen allergy. One of the major plant allergens which millions of people worldwide are sensitized to is Phl p 2, a small protein from timothy grass pollen. Phl p 2 is representative of the large family of cross-reacting plant allergens classified as group 2/3. Recombinant Phl p 2 has been demonstrated by immunological cross-reactivity studies to be immunologically equivalent to the natural protein. RESULTS: We have solved the solution structure of recombinant Phl p 2 by means of nuclear magnetic resonance techniques. The three-dimensional structure of Phl p 2 consists of an all-beta fold with nine antiparallel beta strands that form a beta sandwich. The topology is that of an immunoglobulin-like fold with the addition of a C-terminal strand, as found in the C2 domain superfamily. Lack of functional and sequence similarity with these two families, however, suggests an independent evolution of Phl p 2 and other homologous plant allergens. CONCLUSIONS: Because of the high homology with other plant allergens of groups 1 and 2/3, the structure of Phl p 2 can be used to rationalize some of the immunological properties of the whole family. On the basis of the structure, we suggest possible sites of interaction with IgE antibodies. Knowledge of the Phl p 2 structure may assist the rational structure-based design of synthetic vaccines against grass pollen allergy.     

Genetic modification and plant food allergens: risks and benefits

Plant genetic engineering has the potential to both introduce new allergenic proteins into foods and remove established allergens. A number of allergenic plant proteins have been characterized, showing that many are related to proteins which have potentially valuable properties for use in nutritional enhancement, food processing and crop protection. It is therefore important to monitor the allergenic potential of proteins used for plant genetic engineering and major biotechnology companies have established systems for this. Current technology allows gene expression to be down-regulated using antisense or co-suppression and future developments may allow targeted gene mutation or gene replacement. However, the application of this technology may be limited at least in the short term by the presence of multiple allergens and their contribution to food processing or other properties. Furthermore, the long-term stability of these systems needs to be established as reversion could have serious consequences.     

BRONCHIAL ASTHMA IN ADULTS—Causes and Treatment

Food and inhalant allergens were of about equal importance in producing bronchial asthma in 907 adults between the ages of 15 and 55 years.Drug and especially bacterial allergens were infrequent causes.Psychogenic factors may activate or exaggerate causes mentioned but were not the sole cause of any case in this series.Food allergens are best studied and controlled by the standardized cereal-free elimination diet and at times other elimination diets. Test-negative diets usually fail because of the fallibility of both negative and positive skin tests.Inhalant allergy is controlled by the elimination of causative allergens by varying degrees of environmental control and often by pollen and dust filters in the window. Desensitization is necessary against those allergens that cannot be removed from the environment. Dilutions in the millions and billions may be required.Morphine and other opiates, Demerol, chloral, paraldehyde and large or moderate doses of barbiturates are contraindicated in bronchial asthma, as emphasized by deaths resulting from the use of them.Bronchoscopy is indicated for bronchial obstruction due to inspissated mucous plugs, but the danger of the procedure, especially with much sedation, must be recognized.When secondary infection is probable, the use of sulfadiazine, penicillin, aureomycin and chloramphenicol must be considered.     

BRONCHIAL ASTHMA IN ADULTS-Causes and Treatment

Food and inhalant allergens were of about equal importance in producing bronchial asthma in 907 adults between the ages of 15 and 55 years.Drug and especially bacterial allergens were infrequent causes.Psychogenic factors may activate or exaggerate causes mentioned but were not the sole cause of any case in this series. Food allergens are best studied and controlled by the standardized cereal-free elimination diet and at times other elimination diets. Test-negative diets usually fail because of the fallibility of both negative and positive skin tests. Inhalant allergy is controlled by the elimination of causative allergens by varying degrees of environmental control and often by pollen and dust filters in the window. Desensitization is necessary against those allergens that cannot be removed from the environment. Dilutions in the millions and billions may be required. Morphine and other opiates, Demerol, chloral, paraldehyde and large or moderate doses of barbiturates are contraindicated in bronchial asthma, as emphasized by deaths resulting from the use of them. Bronchoscopy is indicated for bronchial obstruction due to inspissated mucous plugs, but the danger of the procedure, especially with much sedation, must be recognized. When secondary infection is probable, the use of sulfadiazine, penicillin, aureomycin and chloramphenicol must be considered.     

Effects of fungicide treatment and vacuuming on pyroglyphid mites and their allergens in mattress dust

House-dust mites (Pyroglyphidae) are an important source of indoor airborne allergens. Several methods may be applied to reduce the population growth of these mites and thus the quantity of allergen formed. One such method is to interfere with the mites' food chain. Fungi are a key factor in this food chain: they serve as an indirect food source. In this study we investigated the results of the repeated application of a fungicide (natamycin) on mattresses. As controls we treated some mattresses with a placebo, while others were left untreated. The application of natamycin appeared to hamper mite development. Additional vacuuming reduced the quantity of mite allergens present. In the usual household situation repeated treatment will be necessary to obtain a long-term reduction.     

Fungal allergens and peptide epitopes

Fungal allergens represent a major cause of atopic disorders. Immunochemical and molecular characterization of fungal allergens has been hampered by the lack of pure proteins and to inherent variation among fungal proteins and in their poor yields. With the advent of molecular biology techniques, a number of allergens have been cloned, sequenced, and expressed from a variety of fungal species. The knowledge of the primary, secondary, and tertiary structures of these allergens, the immunodominant regions of these proteins, and their interaction with T and B-cell epitopes, results in better understanding of the molecular mechanisms of allergy and may provide avenues of immunologic intervention to treat patients. The present review deals with the current understanding of fungal allergen epitopes.     

Oral immunotherapy against a pollen allergy using a seed-based peptide vaccine

Peptide immunotherapy using dominant T-cell epitopes is safer and more effective than conventional immunotherapy for the treatment of immunoglobulin E (IgE)-mediated allergic diseases. When allergenic T-cell epitope peptides are expressed in the edible part of transgenic plants, successful mucosal immune tolerance to these allergens may be attainable by the consumption of these plants. In this study, we generated transgenic rice seed that accumulated high concentrations (about 60 microg per grain) of polypeptide consisting of seven dominant human T-cell epitopes derived from the Japanese cedar pollen allergens, Cry j 1 and Cry j 2, in the endosperm. Oral administration of these transgenic rice seeds to B10.S mice before or after they were immunized with Cry j 1 holoprotein reduced not only their T-cell proliferative response to Cry j 1, but also their serum IgE levels, proving the efficacy of oral immunotherapy for the treatment of pollinosis.