Patients With Atopic Dermatitis Sensitized to Pet Dander Mount IgE and T-Cell Responses to Mammalian Cystatins, Including the Human Self-Protein.

BACKGROUND: Immediate and delayed-type hypersensitivity reactions to pet-borne allergens are common in atopic diseases. In atopic dermatitis (AD), controversy surrounds the contribution to the disease of cross-reactivity to self-proteins. Human cystatin A and the cat allergen Fel d 3 belong to the cystatins, an evolutionary conserved protein family. The objective of the present study was to assess crossreactivity between mammalian cystatins and to analyze T-cell responses to cystatin in AD patients sensitized to pet dander. METHODS: cDNA coding for dog cystatin was cloned from dog skin. Sera from 245 patients with IgE-mediated sensitization to cat and dog dander were tested for IgE binding to recombinantly expressed feline, canine, and human cystatin. Of these, 141 were also diagnosed with AD. RESULTS: Cystatin-specific IgE was detected in 36 patients (14.7%), of whom 19 were considerably affected by AD. Within the AD patients, 9 had measurable IgE against all 3 cystatins. Cystatin-sensitized AD patients did not differ from non-cystatin-sensitized patients in terms of disease severity, age, or total IgE levels. T-cell cytokine measurements showed elevated IL-4 levels after stimulation with feline and human cystatin. CONCLUSIONS: The humoral response suggests that in addition to Fel d 3, the homologous protein from dog might play a role in allergy. Furthermore, human cystatin appears to be capable of driving a type 2 immune response in sensitized AD patients and may therefore be considered a so-called autoallergen, as proposed for other evolutionary conserved proteins.

Patients With Atopic Dermatitis Sensitized to Pet Dander Mount IgE and T-Cell Responses to Mammalian Cystatins, Including the Human Self-Protein

Background: Immediate and delayed-type hypersensitivity reactions to pet-borne allergens are common in atopic diseases. In atopic dermatitis (AD), controversy surrounds the contribution to the disease of cross-reactivity to self-proteins. Human cystatin A and the cat allergen Fel d 3 belong to the cystatins, an evolutionary conserved protein family. The objective of the present study was to assess crossreactivity between mammalian cystatins and to analyze T-cell responses to cystatin in AD patients sensitized to pet dander. Methods: cDNA coding for dog cystatin was cloned from dog skin. Sera from 245 patients with IgE-mediated sensitization to cat and dog dander were tested for IgE binding to recombinantly expressed feline, canine, and human cystatin. Of these, 141 were also diagnosed with AD. Results: Cystatin-specific IgE was detected in 36 patients (14.7%), of whom 19 were considerably affected by AD. Within the AD patients, 9 had measurable IgE against all 3 cystatins. Cystatin-sensitized AD patients did not differ from non–cystatin-sensitized patients in terms of disease severity, age, or total IgE levels. T-cell cytokine measurements showed elevated IL-4 levels after stimulation with feline and human cystatin. Conclusion: The humoral response suggests that in addition to Fel d 3, the homologous protein from dog might play a role in allergy. Furthermore, human cystatin appears to be capable of driving a type 2 immune response in sensitized AD patients and may therefore be considered a so-called autoallergen, as proposed for other evolutionary conserved proteins. (AU)

Antecedentes: Las reacciones de hipersensibilidad de tipo inmediato y retardado a los alérgenos que están en las mascotas son comunes en las enfermedades atópicas. En este estudio, en pacientes con dermatitis atopica (DA), se analiza la reactividad cruzada con las autoproteínas y su contribución a la enfermedad. Tanto la cistatina A humana como el alérgeno felino Fel d 3 pertenecen a la familia de las cistatinas, una familia de proteínas conservadas evolutivamente. El objetivo del presente estudio fue evaluar la reactividad cruzada entre las cistatinas de mamíferos y analizar la respuestas de las células T a la cistatina en pacientes con DA sensibilizados a la caspa de las mascotas. Métodos: El ADNc que codifica la cistatina de perro se clonó a partir de piel de perro. Se analizaron sueros de 245 pacientes con sensibilización por IgE a la caspa de gato y perro para determinar la unión de IgE a cistatina felina, canina y humana expresada de forma recombinante, respectivamente. De estos 245 pacientes, 141 fueron diagnosticados de DA. Resultados: Se detectó IgE específica frente a cistatina en el 14,7% (36) de los pacientes, de los cuales 19 padecían DA. Dentro de los pacientes con DA, 9 tenían IgE medible contra las tres cistatinas. Los pacientes con DA sensibilizados frente a cistatina no difirieron de los pacientes no sensibilizados con cistatina en términos de gravedad de la enfermedad, edad o niveles totales de IgE. El análisis de citocinas de células T reveló niveles elevados de IL-4 después de la estimulación con cistatina felina y humana. Conclusión: La respuesta humoral sugiere que, además de Fel d 3, la proteína homóloga de perro también podría desempeñar un papel en la alergia. Además, la cistatina humana parece ser capaz de promover una respuesta inmune de tipo 2 en pacientes con DA sensibilizados y, por lo tanto, puede considerarse un autoalérgeno, como se ha propuesto para otras proteínas conservadas evolutivamente. (AU)

Anaphylaxis to bovine serum albumin tissue adhesive in a non-meat-allergic patient

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Identification of a polygalacturonase (Cup s 2) as the major CCD-bearing allergen in Cupressus sempervirens pollen.

As IgE glyco-epitopes, also referred to as cross-reactive carbohydrate determinants (CCDs), can share significant structural homologies between different plants, they are prone to extensive cross-reactivity among allergen pollen extracts. Here, cypress pollen allergens, especially a polygalacturonase (PG), were further characterized using double one-dimensional electrophoresis (D1-DE). The presence of specific IgE directed against CCDs was investigated by bromelain IgE inhibition and concanavalin A binding assays using sera of cypress pollen-sensitized patients. Our results showed that IgE reactivity to CCDs in Cupressus sempervirens pollen extracts is mainly related to bromelain-type epitopes of a newly identified cypress PG. This glycoprotein has been further characterized through an immunoproteomic approach and officially indexed as Cup s 2 by the WHO/IUIS allergen nomenclature. Cup s 2 could thus be associated with the increased prevalence of IgE reactivity to cypress pollen extracts because of CCD interference.

Two galactose-α-1,3-galactose carrying peptidases from pork kidney mediate anaphylactogenic responses in delayed meat allergy.

BACKGROUND: Serum IgE antibodies directed at galactose-α-1,3-galactose (α-Gal) are associated with a novel form of delayed anaphylaxis occurring upon consumption of red meat or innards. Pork kidney is known as the most potent trigger of this syndrome, but the culprit allergens have not yet been identified. The aim of this study was the identification and characterization of pork kidney proteins mediating delayed anaphylactic reactions through specific IgE to α-Gal. METHODS: A cohort of 59 patients with specific IgE to α-Gal was screened by immunoblot for IgE-reactive proteins in pork kidney. Proteins were identified by peptide mass fingerprinting. Isolated proteins were assayed in ELISA and ELISA inhibition, basophil activation and skin prick test. RESULTS: Several IgE-binding proteins of high molecular weight (100- >200 kDa) were detected in pork kidney extracts by immunoblot using patient sera and an anti-α-Gal antibody. Two major IgE-binding proteins were identified as porcine angiotensin-I-converting enzyme (ACE I) and aminopeptidase N (AP-N). Reactivity of patient sera and anti-α-Gal antibody to both proteins was abolished by carbohydrate oxidation. The α-Gal IgE epitopes were resistant to heat denaturation. Pork kidney extract, isolated ACE I, and AP-N were able to activate patient basophils and elicit positive responses in skin prick tests. CONCLUSION: Two cell-membrane proteins carrying α-Gal epitopes were identified in pork kidney. For the first time, isolated meat proteins were shown to induce basophil activation in patients with delayed anaphylaxis to red meat providing further confirmation for the clinical relevance of these α-Gal-carrying proteins.

Evaluation of two new recombinant guinea-pig lipocalins, Cav p 2 and Cav p 3, in the diagnosis of guinea-pig allergy.

BACKGROUND: Guinea-pigs, classical laboratory animals now often kept as pets, regularly elicit important allergic reactions. Two guinea-pig allergens have been described previously to some extent; however, biomolecular and immunological data are lacking. OBJECTIVE: To identify major guinea-pig allergens, produce them as recombinant proteins and define their allergenic potential and clinical importance in allergic patients. METHODS: Protein extracts were prepared from various guinea-pig tissues and allergens were purified by ion exchange chromatography. The N-termini of two immunoglobulin E (IgE)-reactive proteins were determined and degenerate primers were designed for cDNA amplification by RT-PCR. Allergenic properties of recombinant proteins were assayed by immunoblotting, ELISA and mediator release assays. Specific IgE were quantified in the sera of 26 guinea-pig-allergic patients. RESULTS: Major IgE-reactive proteins were detected in submaxillary and harderian gland extracts. Two proteins were identified and the cDNAs were cloned. The 17 kDa protein expressed in the harderian gland corresponds to the previously described Cav p 2. The 19 kDa protein, Cav p 3, is a new allergen expressed in the submaxillary gland. Recombinant Cav p 2 and Cav p 3 were recognized by IgE antibodies from 65% and 54% of guinea-pig-allergic patients, respectively. Both proteins demonstrated equivalent allergenic activity in the mediator release assays. CONCLUSION AND CLINICAL RELEVANCE: Two major guinea-pig allergens, Cav p 2 and Cav p 3, have been characterized and produced as recombinant proteins. Combined to guinea-pig serum albumin, the new allergens proved to be valuable in the component-resolved diagnosis of guinea-pig allergy.

Occupational inhalant allergy to pork followed by food allergy to pork and chicken: sensitization to hemoglobin and serum albumin.

BACKGROUND: Animal-derived proteins are implicated in primary food allergies, but also in inhalant allergies with secondary food allergy symptoms. The objective of this study was to define the allergen(s) implicated in a case of food allergy to chicken meat, which developed in a person previously sensitized to pork after occupational exposure. METHODS: A 42-year-old female with a history of occupational inhalant allergy to pork reported rhinitis, asthma, dysphonia and conjunctivitis 30 min after ingestion of chicken. Skin tests were positive to chicken meat. Protein extracts were prepared from chicken meat. Allergens were characterized by IgE immunoblotting, N-terminal sequencing and ELISA. RESULTS: The patient showed specific IgE binding to chicken meat proteins at 12, 14, 26, 55 and 65 kDa. N-terminal amino acid sequencing identified the 12- and 14-kDa proteins as the alpha- and beta-chain of hemoglobin. ELISA and immunoblot showed specific IgE binding to hemoglobin purified from chicken blood. IgE antibodies to chicken serum albumin were detected by ELISA. Inhibition studies with chicken and porcine hemoglobin as well as with serum albumins demonstrated cross-reactive IgE antibodies. CONCLUSIONS: We report a case of confirmed occupational inhalant allergy due to pork followed by food allergy to pork and 3 years later by food allergy to chicken. Porcine and chicken hemoglobin were found to be cross-reactive allergens. Cross-reactivity between porcine and chicken serum albumin was possibly linked to a prior sensitization to cat serum albumin.