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Background: Cyclophilins are ubiquitous panallergens whose epidemiologic, diagnostic, and clinical relevance is largely unknown and whose sensitization is rarely examined in routine allergy practice., Objective: We investigated the epidemiologic, diagnostic, and clinical relevance of cyclophilins in seasonal allergic rhinitis and its comorbidities., Methods: We examined a random sample of 253 (25%) of 1263 Italian children with seasonal allergic rhinitis from the Panallergens in Pediatrics (PAN-PED) cohort with characterized disease phenotypes. Nested studies of sensitization prevalence, correlation, and allergen extract inhibition were performed in patients sensitized to birch pollen extract but lacking IgE to Bet v 1/2/4 (74/1263) or with highest serum level of IgE to Bet v 1 (26/1263); and in patients with sensitization to various extracts (ragweed, mugwort, pellitory, Plantago, and plane tree), but not to their respective major allergenic molecule, profilins, and polcalcins. IgE to cyclophilin was detected with recombinant Bet v 7, and extract inhibition tests were performed with the same rBet v 7., Results: IgE to rBet v 7 was detected in 43 (17%) of 253 patients. It was associated with asthma (P < .028) and oral allergy syndrome (P < .017) in univariate but not multivariate analysis adjusted for IgE to profilins (Phl p 12), PR-10s (Bet v 1), and lipid transfer proteins (Pru p 3). IgE to rBet v 7 was also highly prevalent (47/74, 63%) among patients with unexplained sensitization to birch pollen extract. In patients with unexplained sensitization to ragweed, mugwort, pellitory, Plantago and plane tree pollen, the levels of IgE to those extracts correlated with the levels of IgE to rBet v 7, and they were also significantly inhibited by rBet v 7 (inhibition range 45%-74%)., Conclusions: IgE sensitization to cyclophilin is frequent in pollen-allergic patients living in temperate areas and can produce "false" positive outcomes in skin prick and IgE tests to pollen extracts. Molecular diagnostic guidelines should include this panallergen family., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Ragweed pollen allergy is the most common seasonal allergy in western Romania. Prolonged exposure to ragweed pollen may induce sensitization to pan-allergens such as calcium-binding proteins (polcalcins) and progression to more severe symptoms. We aimed to detect IgE sensitization to recombinant Amb a 9 and Amb a 10 in a Romanian population, to assess their potential clinical relevance and cross-reactivity, as well as to investigate the relation with clinical symptoms. rAmb a 9 and rAmb a 10 produced in Escherichia coli were used to detect specific IgE in sera from 87 clinically characterized ragweed-allergic patients in ELISA, for basophil activation experiments and rabbit immunization. Rabbit rAmb a 9- and rAmb a 10-specific sera were used to detect possible cross-reactivity with rArt v 5 and reactivity towards ragweed and mugwort pollen extracts. The results showed an IgE reactivity of 25% to rAmb a 9 and 35% to rAmb a 10. rAmb a 10 induced basophil degranulation in three out of four patients tested. Moreover, polcalcin-negative patients reported significantly more skin symptoms, whereas polcalcin-positive patients tended to report more respiratory symptoms. Furthermore, both rabbit antisera showed low reactivity towards extracts and showed high reactivity to rArt v 5, suggesting strong cross-reactivity. Our study indicated that recombinant ragweed polcalcins might be considered for molecular diagnosis.

Background: Blocking the major cat allergen, Fel d 1, with mAbs was effective in preventing an acute cat allergic response., Objectives: This study sought to extend the allergen-specific antibody approach and demonstrate that a combination of mAbs targeting Bet v 1, the immunodominant and most abundant allergenic protein in birch pollen, can prevent the birch allergic response., Methods: Bet v 1-specific mAbs, REGN5713, REGN5714, and REGN5715, were isolated using the VelocImmune platform. Surface plasmon resonance, x-ray crystallography, and cryo-electron microscopy determined binding kinetics and structural data. Inhibition of IgE-binding, basophil activation, and mast cell degranulation were assessed via blocking ELISA, flow cytometry, and the passive cutaneous anaphylaxis mouse model., Results: REGN5713, REGN5714, and REGN5715 bind with high affinity and noncompetitively to Bet v 1. A cocktail of all 3 antibodies, REGN5713/14/15, blocks IgE binding to Bet v 1 and inhibits Bet v 1- and birch pollen extract-induced basophil activation ex vivo and mast cell degranulation in vivo. Crystal structures of the complex of Bet v 1 with immunoglobulin antigen-binding fragments of REGN5713 or REGN5715 show distinct interaction sites on Bet v 1. Cryo-electron microscopy reveals a planar and roughly symmetrical complex formed by REGN5713/14/15 bound to Bet v 1., Conclusions: These data confirm the immunodominance of Bet v 1 in birch allergy and demonstrate blockade of the birch allergic response with REGN5713/14/15. Structural analyses show simultaneous binding of REGN5713, REGN5714, and REGN5715 with substantial areas of Bet v 1 exposed, suggesting that targeting specific epitopes is sufficient to block the allergic response., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Japanese Cedar (JC) pollinosis is the most common seasonal allergic rhinitis in Japan. Throughout the JC pollen season, patients suffer from the allergic symptoms, resulting in a reduction of quality of life. Allergy immunotherapy (AIT) is an established treatment option for a wide range of allergens that unlike symptomatic treatments (e.g. antihistamines) may provide sustained immune tolerance. However, AIT, especially subcutaneous immunotherapy (SCIT) has a fatal anaphylaxis risk due to the use of crude allergen extracts. Consequently, development of allergen derivatives with substantially reduced anaphylactic potential is desirable. An allergen derivative that showed reduced IgE-binding and anaphylactic potential was developed through conjugation of native Cry j 1 (n Cry j 1), a major JC allergen, to the polysaccharide pullulan followed by chemical but non-covalent denaturation. The resulting Cry j 1 allergen derivative, Dn p-Cry j 1, showed reduced IgE-binding and IgE-mediated effector cell activation in vitro using an ELISA competition assay and a mast cell activation model (EXiLE). Reduced anaphylactic potential of Dn p-Cry j 1 in vivo was demonstrated using the rat passive cutaneous anaphylaxis (PCA) assay. The difference in anaphylactic potential of Dn p-Cry j 1 compared to n Cry j 1 in wild-type rats was of the same magnitude as the difference seen in the anaphylaxis reactions obtained with n Cry j 1 in wild-type rats and mast-cell deficient rats, indicating a dramatic reduction in anaphylactic potential of Dn p-Cry j 1. These results indicate that Dn p-Cry j 1 is a promising candidate for next-generation JC AIT., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The previous slide-glass type system could simultaneously detect reactive and highly reactive oxygen species, i.e., superoxide radicals (O 2 -· ) and hypochlorite ions (OCl - ) elicited from leucocytes in sample blood, but had some drawbacks, i.e., signal noise from air-flow stirring, potential biohazard risks, etc. because of open samples placed on a slide glass. We overcame these drawbacks by adopting a fluidic-chip container in a new system, which resulted in higher sensitivity and more stable measurements. Using the new system, we conducted a pilot study on nominally healthy volunteers to find whether or not the monitored activities of leukocytes can distinguish more or less unhealthy conditions from healthy ones. At first, healthy volunteers of both genders and of various ages showed that the fluctuation magnitudes (%) of O 2 -· and OCl - were nearly similar to each other and to that of the neutrophil count fluctuation. These parameters sometimes exceeded the healthy fluctuation range. By comparing these large fluctuations with the data of an inflammation marker C-reactive protein (CRP), the neutrophil count fluctuation and the timings/symptoms of abnormalities found in questionnaire, we could gain information suggesting the factors causing the large fluctuations. The new system could detect bodily abnormalities earlier than CRP or self-aware symptoms., (© 2021. The Author(s).)

Background : The cross-reactive allergen between mugwort ( Artemisia vulgaris ) and kidney bean ( Phaseolus vulgaris ) has not yet been identified. Methods : A total of 24 patients were included in this study. The sera of patients were analyzed for the concentrations of specific IgE antibodies. The allergenicity and cross-reactivity were investigated by Western blotting and immunoblot inhibitory experiments. Results : The immunoblotting indicated the binding of patients' IgE to crude mugwort extract at ~26 kDa protein (15 cases), ~60 kDa (15 cases), and 10-15 kDa proteins (12 cases). The results of the immunoblot-inhibition assay showed that kidney bean seed extract inhibited specific IgE binding to mugwort at 10-15 kDa, ~26 kDa, and ~60 kDa in 4 (16.7%), 1 (4.2%) and 2 (8.3%) cases, respectively. On the other hand, mugwort extract was demonstrated to inhibit specific IgE binding to kidney bean seed at 10-15 kDa, 15-20 kDa, ~30 kDa, and 60 kDa in 1 (4.2%), 3 (12.5%), 4 (16.7%), and 3 (12.5%) cases, respectively. Conclusion : The 26-30 kDa, 10-15 kDa, and 60 kDa proteins are potential causative agents of the cross-reactivity between mugwort and kidney beans. The findings of this study improved the current understanding on the allergenicity of kidney beans and would provide insights into the refinement of treatment strategy for anaphylaxis.

Objective: At least 3 years of sublingual immunotherapy (SLIT) is required to achieve long-term clinical tolerance for allergens. However, immunological changes with more than 3 years of SLIT have not yet been elucidated in detail. The present study investigated whether the numbers of regulatory T (Treg) cells and regulatory B (Breg) cells increased with 4 years of SLIT and if these increases correlated with clinical effects for pollinosis., Methods: Seven Japanese cedar pollinosis patients received SLIT in 2014 or 2015 and continued treatment until May 2019. In May 2017 and May 2019, peripheral blood mononuclear cells (PBMCs) were collected from the patients, and analyzed by flow cytometer., Results: (1) The visual analogue scale (VAS) was significantly higher in 2019 than in 2017. (2) The percentages of Foxp3 + Treg cells, type 1 regulatory T (Tr1) cells, and Breg cells in PBMCs were significantly higher in 2019 than in 2017. (3) The percentage of Foxp3 + Treg cells in PBMCs positively correlated with VAS, whereas those of Tr1 cells and Breg cells did not., Conclusions: These results suggest that 4 years of SLIT is needed to achieve sustained increases in Foxp3 + Treg cells, which are closely associated with the efficacy of SLIT.

Background: In the pathogenesis of intermittent allergic rhinitis (IAR), the inflammatory reaction is of importance. CD48, belonging to the CD2 family, participates in mast cell-stimulating cross-talk, facilitates the formation of the mast cell/eosinophil effector unit, and is expressed by eosinophils., Objectives: To assess the serum level of soluble form of CD48 (sCD48) in patients with IAR during and out of the pollen season and correlate with the disease severity and with eosinophil-related parameters., Materials and Methods: Sixty-three patients (female: 79%; mean age: 30.58) were included to the study. Forty-five patients were assessed during the pollen season and other 42 patients during out of the pollen season. Twenty-four patients (female: 37.50%; mean age: 27.90) were evaluated twice, during the pollen season and out of the pollen season. sCD48, ECP, eotaxin-1/CCL11 serum levels together with complete blood count, and fractional exhaled nitric oxide bronchial and nasal fraction (FeNO) were performed. The severity of symptoms was assessed using the Total Nasal Symptom Score (TNSS), and neutrophil-to-lymphocyte (NLR) and eosinophil-to-lymphocyte (ELR) ratios were calculated., Results: sCD48 serum level, FeNO nasal and bronchial fractions, and TNSS were significantly higher in the IAR group in the pollen season compared with out of the pollen season. Differences in ECP, eotaxin-1/CCL11 serum levels, and NLR and ELR were not significant between season and out of the season. No correlations were found between sCD48 and eosinophil-related parameters., Conclusions: sCD48 may be a biomarker to the exacerbation phase in patients with IAR. One can assume that CD48 participates in the pathogenesis of IAR., (© 2020 S. Karger AG, Basel.)

Background: Allergic rhinitis is a common disorder that affects 10% to 40% of the population worldwide. Allergen immunotherapy (AIT) represents the only therapy that has the potential to resolve clinical symptoms of allergic rhinitis. However, up to 30% of patients do not respond to AIT. Biomarkers predicting the clinical efficacy of AIT as early as possible would significantly improve the patient selection and reduce unnecessary societal costs., Methods: Artemisia pollen allergic patients who received at least 1-year AIT were enrolled. Clinical responses before and after 1-year AIT were evaluated to determine AIT responders. Artemisia specific IgE and IgG4 levels were measured by using ImmunoCAP and enzyme-linked immunosorbent assay (ELISA) separately. Stepwise regression analysis was performed to identify which rhinitis-relevant parameters explained the most variability in AIT results. Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics was applied to identify the potential candidate biomarkers in the sera of responders and non-responders collected before and after 1-year therapy. The diagnostic performance of the potential biomarkers was then assessed using enzyme-linked immunosorbent assay (ELISA) in 30 responders and 15 non-responders., Results: Artemisia specific IgE and IgG4 levels were elevated only in the responders. Regression analysis of allergic rhinitis-relevant parameters provided a robust model that included two most significant variables (sneeze and nasal congestion). Thirteen candidate biomarkers were identified for predicting AIT outcomes. Based on their association with allergy and protein fold change (more than 1.1 or less than 0.9), four proteins were identified to be potential biomarkers for predicting effective AIT. However, further ELISA revealed that only leukotriene A 4 hydrolase (LTA 4 H) was consistent with the proteomics data. The LTA 4 H level in responders increased significantly (P < 0.001) after 1-year therapy, while that of non-responders remained unchanged. Assessment of LTA 4 H generated area under curve (AUC) value of 0.844 (95% confidence interval: 0.727 to 0.962; P < 0.05) in distinguishing responders from the non-responders, suggesting that serum LTA 4 H might be a potential biomarker for predicting the efficiency of AIT., Conclusion: Serum LTA 4 H may be a potential biomarker for early prediction of an effective AIT., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Ma, Cao, Yu, Shi, Yan, Liu, Pan, Sun, Wei, Wang, Wei, Wang and Yin.)

Background: Pollen exposure induces local and systemic allergic immune responses in sensitized individuals, but nonsensitized individuals also are exposed to pollen. The kinetics of symptom expression under natural pollen exposure have never been systematically studied, especially in subjects without allergy., Objective: We monitored the humoral immune response under natural pollen exposure to potentially uncover nasal biomarkers for in-season symptom severity and identify protective factors., Methods: We compared humoral immune response kinetics in a panel study of subjects with seasonal allergic rhinitis (SAR) and subjects without allergy and tested for cross-sectional and interseasonal differences in levels of serum and nasal, total, and Betula verrucosa 1-specific immunoglobulin isotypes; immunoglobulin free light chains; cytokines; and chemokines. Nonsupervised principal component analysis was performed for all nasal immune variables, and single immune variables were correlated with in-season symptom severity by Spearman test., Results: Symptoms followed airborne pollen concentrations in subjects with SAR, with a time lag between 0 and 13 days depending on the pollen type. Of the 7 subjects with nonallergy, 4 also exhibited in-season symptoms whereas 3 did not. Cumulative symptoms in those without allergy were lower than in those with SAR but followed the pollen exposure with similar kinetics. Nasal eotaxin-2, CCL22/MDC, and monocyte chemoattactant protein-1 (MCP-1) levels were higher in subjects with SAR, whereas IL-8 levels were higher in subjects without allergy. Principal component analysis and Spearman correlations identified nasal levels of IL-8, IL-33, and Betula verrucosa 1-specific IgG 4 (sIgG 4 ) and Betula verrucosa 1-specific IgE (sIgE) antibodies as predictive for seasonal symptom severity., Conclusions: Nasal pollen-specific IgA and IgG isotypes are potentially protective within the humoral compartment. Nasal levels of IL-8, IL-33, sIgG 4 and sIgE could be predictive biomarkers for pollen-specific symptom expression, irrespective of atopy., (Copyright © 2020. Published by Elsevier Inc.)

Background: Severe allergy to fruits mediated by a 7 kDa allergen belonging to the gibberellin-regulated protein (GRP) family is known to be associated with Cupressaceae pollinosis., Objective: To identify and characterize Cupressaceae pollen allergens involved in GRP-related fruit allergy., Methods: Pru p 7-related proteins from pollen of Cupressus sempervirens, Juniperus ashei and Cryptomeria japonica were identified using a rabbit anti-Pru p 7 antiserum, purified chromatographically and sequenced by mass spectrometry and bioinformatic comparisons. The C sempervirens protein was produced as a recombinant allergen in Pichia pastoris. IgE antibody binding to pollen GRP proteins was analysed in a peach allergic (n = 54) and a cypress pollen allergic (n = 88) patient population from southern France using ImmunoCAP., Results: In each of the three Cupressaceae species studied, a 7 kDa pollen protein related to Pru p 7 was identified and found to comprise an amino acid sequence of 63 residues in length, 92%-98% identical to each other and 67%-68% identical to Pru p 7. The C sempervirens, J ashei and C japonica GRP allergens have been officially recognized by the WHO/IUIS Allergen Nomenclature Sub-Committee and named Cup s 7, Jun a 7 and Cry j 7, respectively. Recombinant Cup s 7 showed IgE antibody binding capacity comparable to that of the purified natural allergen. Among 51 peach allergic subjects sensitized to Pru p 7, substantially higher levels of IgE to Cup s 7 than to Pru p 7 were found. Further, the pollen protein was able to completely outcompete IgE binding to Pru p 7, while the reverse competition effect was modest, consistent with primary sensitization by the pollen allergen., Conclusion and Clinical Relevance: Pru p 7-related pollen allergens from three Cupressaceae species have been characterized and may become useful for the identification of pollinosis patients at risk of developing severe fruit allergy., (© 2020 John Wiley & Sons Ltd.)

Background: Bermuda grass is a prevalent allergen that flourishes in tropical climates. Its exposure is traditionally believed to be low in Ontario due to the colder environment. However, high sensitization rates have been observed in Kingston, Ontario., Objective: We sought to investigate whether its allergens can provoke allergic rhinitis (AR) symptoms in sensitized participants from south-eastern Ontario and determine if nasal allergen challenge (NAC) model is appropriate to study Bermuda grass-induced AR., Methods: Twenty-one participants sensitized to Bermuda grass and 12 nonallergic participants completed a titrated NAC with increasing allergen concentrations at a screening visit. Total nasal symptom score (TNSS) and peak nasal inspiratory flow were collected before allergen exposure and 10 minutes after delivery of each concentration. Twelve participants with a Bermuda grass allergy who met the qualifying criteria (TNSS ≥ 8 and peak nasal inspiratory flow fall ≥ 50%) and 11 nonallergic controls returned for single-dose NAC visit., Results: At titrated NAC, 19 of 21 sensitized participants met the criteria of positive allergic response when challenged. During single-dose NAC, participants with allergy had significantly greater TNSS between 15 minutes and 3 hours after NAC than controls. Likewise, allergic participants had a significantly increased number of nasal lavage eosinophils at both 1 and 6 hours after NAC. Bermuda grass-specific immunoglobulin E was significantly increased in Bermuda grass allergic participants at NAC than screening visit., Conclusion: Although Bermuda grass is a non-native allergen in Ontario, it can induce AR symptoms in sensitized participants, and the NAC model is appropriate to study Bermuda grass-induced AR., (Copyright © 2020 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Background: There is growing interest both in testing IgE in nasal secretions (NS) and in molecular diagnosis of seasonal allergic rhinitis (SAR). Yet, the reliability of nasal IgE detection with the newest molecular assays has never been assessed in a large cohort of pollen allergic patients., Objective: To investigate with microarray technology and compare the repertoires of specific IgE (sIgE) antibodies in NS and sera of a large population of children and adults with SAR., Methods: Nasal secretions were collected with an absorbent device (Merocel 2000 ® , Medtronic) and a minimal dilution procedure from 90 children and 71 adults with SAR. Total IgE (tIgE) (ImmunoCAP, Thermo Fisher Scientific (TFS)) and sIgE antibodies against 112 allergen molecules (ISAC-112, TFS) were measured in NS and serum., Results: Nasal sIgE was detectable in 68.3% of the patients. The detected nasal sIgE antibodies recognized airborne (88%), vegetable (10%), and animal food or other (<1%) allergen molecules. The prevalence and average levels of sIgE in NS and serum were highly interrelated at population level. A positive nasal sIgE antibody to a given molecule predicted the detection of the same antibody in the patient's serum with a specificity of 99.7% and a sensitivity of 40%., Conclusions: The concentration of sIgE is much lower in nasal secretions than in the serum. sIgE assays with very high analytical sensitivity and sampling methods with minimal dilution will be therefore needed to validate nasal secretions as alternative to serum in testing the sIgE repertoire., (© 2019 The Authors. Pediatric Allergy and Immunology published by John Wiley & Sons Ltd.)

Olive pollen is one of the most important causes of respiratory allergy, with Ole e 1 being the most clinically relevant sensitizing allergen. Peptide-based vaccines represent promising therapeutic approaches, but the use of adjuvants is required to strengthen the weak immunogenicity of small peptides. We propose the use of dendrimeric scaffolds conjugated to the T cell immunodominant epitope of Ole e 1 (OE 109-130 ) for the development of novel vaccines against olive pollen allergy. Four dendrimeric scaffolds containing an ester/ether with nine mannoses, an ester succinimidyl linker with nine N -acetyl-glucosamine units or nine ethylene glycol units conjugated to OE 109-130 peptide were designed, and their cytotoxicity, internalization pattern, and immunomodulatory properties were analyzed in vitro. None of the dendrimers exhibited cytotoxicity in humanized rat basophil (RBL-2H3), human bronchial epithelial Calu-3, and human mast LAD2 cell lines. Confocal images indicated that mannosylated glycodendropeptides exhibited lower colocalization with a lysosomal marker. Moreover, mannosylated glycodendropeptides showed higher transport tendency through the epithelial barrier formed by Calu-3 cells cultured at the air-liquid interface. Finally, mannosylated glycodendropeptides promoted Treg and IL10 + Treg proliferation and IL-10 secretion by peripheral blood mononuclear cells from allergic patients. Mannosylated dendrimers conjugated with OE 109-130 peptide from Ole e 1 have been identified as suitable candidates for the development of novel vaccines of olive pollen allergy.

Background: An increased prevalence of allergies and an increased incidence of breast cancer have been observed. The hypothesis that atopy may have a protective effect against the risk of different types of breast cancer was evaluated., Methods: In this study, 11,101 patients (11,101 women with a mean age of 55.2±14.7 years) with different types of breast cancer were tested for allergies. Allergies were confirmed based on the retrospective analysis of allergy diagnostic procedures in patients who had been previously diagnosed with breast cancer. The retrospective prevalence rates of active allergic diseases, including allergic rhinitis, bronchial asthma and atopic dermatitis, were assessed. All patients were also analyzed for bronchial asthma and allergic rhinitis according to the relevant guidelines. A group of healthy control patients was used for the comparisons., Results: The women with breast cancer had a significantly lower incidence of IgE-mediated allergic diseases than the controls. The odds ratios (ORs) for allergic rhinitis, atopic dermatitis, and bronchial asthma were 0.61 (95% CI: 0.57-0.73), 0.17 (95% CI: 0.11-0.44), and 0.73 (95% CI: 0.65-0.83), respectively. The mean serum concentrations of total IgE were significantly lower in the study population of women with breast cancer than in the patients of the control group (39.2 ± 26.2 kU/L vs. 108.5 ± 38.5 kU/L; p = 0.002)., Conclusion: Our results suggest that the overall incidence of allergies, especially allergic rhinitis, is lower in patients with certain types of cancer than in individuals who did not have cancer. Further studies are needed to confirm our findings., Competing Interests: The authors have declared that no competing interests exist.

Background: Tree pollens are well-known aeroallergens all over the world. Little is known about the allergenicity of Morus alba (white mulberry) pollen., Objecive: We aimed to explore the potential allergens of this pollen and its clinical relevance in tree pollen allergic patients living in Istanbul, Turkey., Methods: Twenty three seasonal allergic rhinitis patients with a confirmed tree pollen allergy and 5 healthy control subjects underwent skin prick and nasal provocation tests with M.alba pollen extract. The pollen extract was then resolved by gel electrophoresis, and immunoblotted with sera from patients/control individuals to detect the potential allergenic proteins. The prevalent IgE binding proteins from 1D-gel were analyzed by MALDI-TOF/TOF., Results: Eleven out of 23 patients were reactive to the extract with skin prick tests. Seven of those patients also reacted positively to the nasal provocation tests. The most common IgE-binding pollen proteins were detected between 55-100 kDa, and also at molecular weights lower than 30 kDa for some patients. Mass spectrometry analyses revealed that the principal IgE-binding protein was methionine synthase (5-methyltetrahydropteroyltriglutamate homocysteine methyltransferase), which is then proposed as a novel allergen in M.alba pollen., Conclusion: This study provides the first detailed information for the potential allergens of Morus alba pollen of Istanbul. Methionine synthase with an apparent molecular weight of 80 to 85 kDa has been recognized as one of the allergens in Morus alba pollen for the first time.

Olive pollen is a major cause of immunoglobulin E (IgE)-mediated allergy in Mediterranean countries. It is expected to become a worldwide leading allergenic source because olive cultivation is increasing in many countries. Ole e 15 belongs to the cyclophilin pan-allergen family, which includes highly cross-reactive allergens from non-related plant, animal and mold species. Here, the amino acid differences between Ole e 15 and its weak cross-reactive human homolog PPIA were grafted onto Ole e 15 to assess the contribution of specific surface areas to the IgE-binding. Eight Ole e 15-PPIA chimeras were produced in E. coli, purified and tested with 20 sera from Ole e 15-sensitized patients with olive pollen allergy by ELISA experiments. The contribution of linear epitopes was analyzed using twelve overlapping peptides spanning the entire Ole e 15 sequence. All the patients displayed a diverse reduction of the IgE-reactivity to the chimeras, revealing a highly polyclonal and patient-specific response to Ole e 15. IgE-epitopes are distributed across the entire Ole e 15 surface. Two main surface areas containing relevant conformational epitopes have been characterized. This is the first study to identify important IgE-binding regions on the surface of an allergenic cyclophilin.

Background: The temporal sequence in which allergic sensitization to different allergens emerges is not well characterized at the level of general population., Objective: We describe the incidence patterns of atopic sensitization to different allergens from birth up to 12 years of age in an unselected Finnish population., Methods: The study population comprised all children born between 2001 and 2006 identified from the nationwide population register as residents of the province of South Karelia, Finland (n = 5564). The results of allergy tests (22,380 results from skin prick tests, immunoglobulin E [IgE] antibodies, and open food challenges [OFCs], performed in 1827 children) were collected from patient records of all the health care units in the area., Results: The incidence rates of positive results for food and animal allergens as well as positive OFCs for cow's milk showed prominent peaks at 5 months of age. Positive results for pollen allergens started to emerge after 1.5 years of age. The 12-year cumulative incidence of sensitization to food, animal, pollen, and any allergens was 12%, 8%, 10%, and 18%, respectively. The cumulative incidence of sensitization to house dust mites was 1% and to molds or latex less than 1%. Firstborn boys had the highest, and those who were not firstborn girls and children born in rural municipalities had the lowest early incidence of sensitization to inhalation allergens., Conclusion: In the unselected population, the atopic sensitization against food and animal allergens began before 6 months of age and was followed by sensitization to pollen allergens before 2 years of age. Primary prevention of sensitization to food and inhalation allergens should therefore occur in early infancy., (Copyright © 2019 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Plant lipid transfer proteins and homologues of the main birch pollen allergen Bet v 1 are involved in the development of allergic reactions of varying severity to plant foods and pollen. In this study, the sera from patients with tree and weed pollen allergies in the Moscow region were examined. The levels of IL-4, IL-5, IL-9, IL-10, IL-13, IL-17A, IFNγ, TNFα, and TNFβ cytokines were determined in the sera of patients with specific IgE antibodies to Bet v 1 and Pru p 3 allergens. It was confirmed that patients with pollen allergy are often characterized by Th2 response of the immune system, though other mechanisms of allergy development occurred in some cases. The data obtained demonstrate the necessity of detailed analysis of the individual mechanism of allergic reactions and patient-centered approach to the personalized allergy treatment.

Background: The identification of the primary sensitizing pollen is difficult in Southern European patients with Seasonal Allergic Rhinitis (SAR) if sensitized to various pollen sources with overlapping seasonality. A more precise diagnosis is obtained by IgE assays to allergen molecules, currently available as singleplex or microarrays., Objectives: To test the analytical performance of a multi-parameter immunoblot molecular "Pollen Test" specifically designed to test IgE antibodies to pollen extracts and molecules clinically relevant in Southern Europe., Methods: Sera were obtained from 101 children and 98 adults with SAR and tested with a customized multiplex immunoblot assay (EUROLINE Southern European Pollen Profile [ESEP]; EUROIMMUN AG, Luebeck, Germany) containing a comprehensive panel of allergen extracts and molecules. ESEP's outcomes were then compared in selected sera (ESEP positive to negative = 2:1) with those of singleplex IgE assays (ImmunoCAP; ThermoFisher Scientific, Uppsala, Sweden). For each of the examined reagents, qualitative (sensitivity, specificity, accuracy), semi-quantitative (classes) and quantitative (Spearman's rank correlation, Bland-Altmann plots) comparisons were performed., Results: Compared to ImmunoCAP, cumulative ESEP's sensitivity and specificity were 87% (95% CI 84%-90%) and 88% (83%-93%) for extracts and 99% (98%-100%) and 87% (83%-91%) for molecules. Cohen's kappa coefficients (κ C ) ranged for extracts from 0.18 (Pellitory) to 0.50 (Cypress) and for molecules from 0.21 (Ole e 1) to 0.68 (Phl p 7). The quantitative outcomes of the two diagnostic tests were highly correlated, with Spearman's rank correlation coefficients always exceeding 0.80. Bland-Altmann plots showed a tendency of ESEP to overestimate serum specific IgE levels, when compared to ImmunoCAP., Conclusions and Clinical Relevance: Sensitivity and specificity of ESEP in testing serum IgE antibodies against pollen allergen extracts and molecules, in Italian patients with SAR, both exceeded 85%. The advantages and limitations of a multiplex customized immunoblot assay, in the routine clinical use of molecular diagnostics in Southern European pollen allergic patients, deserve to be tested., (© 2018 John Wiley & Sons Ltd.)

Objective: Using multiplex microarray-based component resolved diagnosis (CRD) to investigate the allergen sensitization profile of allergic asthma patients in southern China., Method: Serum samples from 57 polysensitized mite allergic asthmatic patients in a tertiary referral centre of southern China were tested with multiplex CRD (ISAC) for specific immunoglobulin E (sIgE) against 112 single allergen and components. Result was then compared with those from singleplex ImmunoCAP., Results: With ISAC, the highest sensitization was seen for nDer f 1 (71.9%), rDer f 2 (73.7%), nDer p 1 (70.2%) and rDer p 2 (66.7%), whereas rDer p 10 and other storage mites' components only showed 10% positivity. rFel d 1 and rCan f 1 were found positive in 29.8% and 14.0% samples respectively. Other epithelia components had less than 7.0% positive rate. Sensitization to pollen components was dominated by nCyn d 1 (17.5%) and nPhl p 4 (12.3%), Carbohydrate cross-reactive determinants (CCD) was positive in 4 patients who were also positive to nPhl p 4, nCyn d 1 and rPla a 2, and all of them have combined asthma and rhinitis. The sensitivity to mold (rAsp f 3), cockroach (nBla g 7) and Anisakis simplex component (rAni s 3) were all the same at 8.8%. 93.0% patients were sensitive to more than one component, with more than half of them (57.9%) positive to five or more components. Patients with combined asthma and rhinitis (AA + AR) were sensitive to more components than those with asthma only (AA). Positive rate to nPhl p 4 was significantly higher in patients with AA + AR than with AA only (χ 2  = 4.31, P =  0.038). Compared with ImmunoCAP, ISAC showed a similar high detection rate for D. pteronyssinus and D. farinae, but only 10.0% of B. tropicalis sensitive patients were positive to rBlo t 5. Optimal scale analysis on correlation of allergens components showed rDer p 10 was associated to food allergy., Conclusion: Being the first multiplex microarray based CRD study on southern Chinese, ISAC showed house dust mites components were the major allergen components led to sensitization in asthmatic patients. Patients with combined AA + AR were sensitive to more components than those with AA only. Other components with higher positive rate include pollen components nCyn d 1, nPhl P 4 and animal dander components rFel d 1 and rCan f 1. For B. tropicalis, the rBlo t 5 in ISAC may not represent the major Blomia component in southern Chinese patients., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Background: Genetic and environmental factors are proposed to be involved in cedar pollen allergy sensitization and onset. The impact of these factors will provide key information for the prevention of cedar pollen sensitization and allergy onset, which we investigated in this cross-sectional study., Methods: Subjects were 382 young adult volunteers who completed a self-administered questionnaire on self-reported subjective symptoms of pollinosis, physician-diagnosed pollinosis, and background factors. We also measured their serum IgE antibody titers specific for cedar, cypress, and mites. Factors associated with subjective symptoms, physician diagnosis, and the three specific antigens were determined using both univariate and multivariate analyses., Results: Sensitization to cedar, cypress, and mites, defined as specific IgE levels of class 1 or above, was found in 78.8%, 64.4%, and 56.0% of subjects, respectively. The prevalence of cedar pollinosis was 41.2% based on subjective symptoms and 22.2% based on physician diagnosis. Factors associated with increased cedar pollen sensitization were mite sensitization, comorbid allergic rhinitis, and family history of cedar pollinosis. Risk-reducing factors for cedar pollen sensitization were keeping a cat, number of common colds, and hours of sleep. Risk-increasing factors for both subjective pollinosis symptoms and physician-diagnosed pollinosis were comorbid allergic rhinitis and family history of cedar pollinosis., Conclusions: Sensitization to cedar pollen in this population was extremely high. Both common and distinct factors were associated with sensitization to pollen and with the development of pollinosis. The distinct factors were associated with sensitization to cedar and cypress antigens., (Copyright © 2018 Japanese Society of Allergology. Production and hosting by Elsevier B.V. All rights reserved.)

Background: About one-third of the Japanese population suffers from Japanese cedar pollinosis, which is frequently accompanied by Japanese cypress pollinosis. Recently, a novel major Japanese cypress pollen allergen, Cha o 3, was discovered. However, whether a Cha o 3 homolog is present in Japanese cedar pollen remains to be determined., Methods: Western blot analysis was performed using Cha o 3-specific antiserum. In addition, cloning of the gene encoding Cry j 4 was conducted using total cDNA from the male flower of Japanese cedar trees. Allergen potency and cross-reactivity were investigated using a T-cell proliferation assay, basophil activation test, and ImmunoCAP inhibition assay., Results: A low amount of Cha o 3 homolog protein was detected in Japanese cedar pollen extract. The deduced amino acid sequence of Cry j 4 showed 84% identity to that of Cha o 3. Cross-reactivity between Cry j 4 and Cha o 3 was observed at the T cell and IgE levels., Conclusions: Cry j 4 was discovered as a counterpart allergen of Cha o 3 in Japanese cedar pollen, with a relationship similar to that between Cry j 1-Cha o 1 and Cry j 2-Cha o 2. Our findings also suggest that allergen-specific immunotherapy (ASIT) using Japanese cedar pollen extract does not induce adequate immune tolerance to Cha o 3 due to the low amount of Cry j 4 in Japanese cedar pollen. Therefore, ASIT using Cha o 3 or cypress pollen extract coupled with Japanese cedar pollen extract is required in order to optimally control allergy symptoms during Japanese cypress pollen season., (Copyright © 2018 Japanese Society of Allergology. Production and hosting by Elsevier B.V. All rights reserved.)

Background: The overlapping grass and olive pollen seasons in Spain and the phenomenon of cross-reactivity can make it difficult to determine the true causative agent of seasonal allergic rhinitis when only skin prick tests with whole extracts are used. The aim of the GRAMOLE study was to determine sensitization patterns to the major grass and olive pollen allergens detected using specific recombinant IgE and to explore how this knowledge affected physicians' choice of allergen-specific immunotherapy., Methods: Epidemiological, observational, multicenter, cross-sectional study. Results from children under 18 years of age diagnosed with seasonal allergic rhinitis by positive skin prick tests to olive and grass pollen were analyzed. Specific IgE to Phl p 1+5, Ole e 1, and Phl p 7+12 was determined. Investigators specified the optimal composition of allergen immunotherapy before and after knowing the results of the molecular diagnosis., Results: A total of 281 patients with a mean age of 13.4 years were included. Double sensitization to both major allergens was found in vitro in 76% of children for an IgE cutoff point of 0.35 kU/L. When the molecular diagnosis results were known, specialists changed the composition of the prescribed immunotherapy in 52.87% of cases., Conclusions: Double sensitization to grass and olive pollen is common in Spain and also occurs in the pediatric population. Molecular diagnosis using specific IgE may help improve immunotherapy selection in polysensitized patients., (© 2018 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.)

The Salsola kali pollen is considered the main cause of allergic sensitization in desert and semi-desert regions. We have constructed recombinant Lactococcus lactis producing Sal k1 protein with the aim of using it as a mucosal vaccine for specific immunotherapy. The Sal k1 gene was amplified, and transferred into a PNZ 8148 plasmid. The PNZ8148-Sal k1 recombinant plasmid was transformed into competent E.coli strain MC1061 for replication, and then was isolated and cloned into competent L. lactis by electroporation. The cloning was verified by PCR and gene sequencing. The production of recombinant Sal K1 (rSal K1) protein was induced by nisin. The rSal K1 protein was purified by affinity chromatography and dialysis, and confirmed by SDS-PAGE and western blot analyses. The recombinant L. lactis was successfully constructed. Production of a 40-kDa rSal k1 protein with the L. lactis was shown by sodium dodecyl sulfate-polyacrylamid gel electrophoresis (SDS-PAGE) analysis. In addition, western blot analysis using specific mouse anti-Sal k1 polyclonal antibodies and sensitive human sera verified the 40-kD protein as rSal k1 allergen. This study demonstrated that L. lactis may be used as a promising live delivery system for recombinant Sal k1 protein without altering its immunoreactivity; however, its efficacy in the context of the immune system is suggested to be pursued in future studies.

The in vivo model of pollinosis has been established using rodents, but the model cannot completely mimic human pollinosis. We used Callithrix jacchus, the common marmoset (CM), to establish a pollinosis animal model using intranasal weekly administration of cedar pollen extract with cholera toxin adjuvant. Some of the treated CMs exhibited the symptoms of snitching, excess nasal mucus and/or sneezing, but the period was very short, and the symptoms disappeared after several weeks. The CD4+CD25+ cell ratio in the peripheral blood increased in CMs quickly after the nasal administration of cedar pollen extract, but the timing was not parallel with the symptoms. IL-10 mRNA was enhanced in the peripheral blood mononuclear cells (PBMCs), suggesting CM-induced tolerance for cedar pollen administration. Similarly, Foxp3 mRNA was also detected in the PBMC. Additive sensitization of these CMs with Ascaris egg administration did not enhance chronic inflammation of type 1 allergy to induce the symptoms. These results suggest that the environmental immune cells develop transient allergic symptoms and subsequent immune-tolerance in the intranasally sensitized CMs.

Background: Grass pollen-related seasonal allergic rhinoconjunctivitis (SARg) is clinically heterogeneous in severity, comorbidities, and response to treatment. The component-resolved diagnostics disclosed also a high heterogeneity at molecular level. Our study aimed at analyzing the characteristics of the IgE sensitization to Phleum pratense molecules and investigating the diagnostic relevance of such molecules in childhood., Methods: We examined 1120 children (age 4-18 years) with SARg. Standardized questionnaires on atopy were acquired through informatics platform (AllergyCARD™). Skin prick tests were performed with pollen extracts. Serum IgE to airborne allergens and eight P. pratense molecules (rPhl p 1, rPhl p 2, rPhl p 4, rPhl p 5b, rPhl p 6, rPhl p 7, rPhl p 11, rPhl p 12) were tested by ImmunoCAP FEIA., Results: The analysis of IgE responses against eight P. pratense molecules showed 87 profiles. According to the number of molecules recognized by IgE, the more complex profiles were characterized by higher serum total IgE, higher grass-specific serum IgE, and higher number and degree of sensitization to pollens. The most frequent IgE sensitization profile was the monomolecular Phl p 1. Sensitization to Phl p 7 was a reliable biomarker of asthma, whereas Phl p 12 of oral allergy syndrome. Sensitization to Phl p 7 was associated with a higher severity of SARg, and complex profiles were associated with longer disease duration., Conclusions: In a large pediatric population, the complexity of IgE sensitization profiles against P. pratense molecules is related to high atopic features although useless for predicting the clinical severity. The detection of serum IgE to Phl p 1, Phl p 7, and Phl p 12 can be used as clinical biomarkers of SARg and comorbidities. Further studies in different areas are required to test the impact of different IgE molecular profiles on AIT response., (© 2017 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.)

Background/objectives: Studies of the effect of vitamin B12 and folate on the risk of asthma and hay fever have shown inconsistent results that may be biased by reverse causation and confounding. We used a Mendelian randomization approach to examine a potential causal effect of vitamin B12 and folate on hay fever, asthma, and selected biomarkers of allergy by using 11 vitamin B12-associated single-nucleotide polymorphisms (SNPs) and 2 folate-associated SNPs as unconfounded markers., Subjects/methods: We included 162,736 participants from 9 population-based studies including the UK Biobank. Results were combined in instrumental variable and meta-analyses and effects expressed as odds ratios (ORs) or estimates with 95% confidence interval (CI)., Results: Using genetic proxies for B12 and folate, instrumental variable analyses did not show evidence for associations between serum B12 and hay fever: OR = 1.02 (95% CI: 0.98, 1.05), asthma: OR = 0.99 (95% CI: 0.95, 1.04), allergic sensitization: OR = 1.02 (95% CI: 0.74, 1.40), or change in serum IgE: 10.0% (95% CI: -9.6%, 29.6%) per 100 pg/ml B12. Similarly, there was no evidence for association between serum folate and hay fever: OR = 0.74 (95% CI: 0.45, 1.21), asthma: OR = 0.80 (95% CI: 0.43, 1.49), or allergic sensitization: OR = 1.92 (95% CI: 0.11, 33.45), but there was a statistically significant association with change in serum IgE: 2.0% (95% CI: 0.43%, 3.58%) per 0.1 ng/ml serum folate., Conclusions: Our results did not support the hypothesis that levels of vitamin B12 and folate are causally related to hay fever, asthma, or biomarkers of allergy, but we found evidence of a positive association between serum folate and serum total IgE.

Background: English plantain (Plantago lanceolata) is an important weed pollen allergen source triggering allergic symptoms during summer. To elucidate genuine versus cross-reactive sensitization, we investigated IgE reactivity patterns and inhibition capacities of plantain-sensitized patients., Methods: Sera of 35 rhinoconjunctivitis patients from the north-east of France with positive skin prick tests (SPT) to Plantago lanceolata pollen were tested with clinically relevant allergen sources using ELISA, ImmunoCAP, and immunoblot inhibition., Results: The patients were multisensitized with additional reactivity to grass (94.3%), ash (74.3%), birch (71.4%), and mugwort (55.2%) pollen in SPT. Sensitization prevalence to allergen molecules was 34.3% (Pla l 1), 94.3% (Phl p 1/5), 60.0% (Ole e 1), 65.7% (Bet v 1), 37.1% (profilin), and 40.0% (CCD). In immunoblot, IgE reactivity to plantain pollen was inhibited with relevant pollen extracts and purified rPla l 1. Two sera did not reveal any IgE cross-reactivity, while reactivity to plantain was efficiently inhibited by grass pollen in the sera of 10 patients. The sera from 17 different patients could be inhibited by grass, birch, or ash pollen to varying degrees. Thus, only 37.1% of our patients demonstrated true plantain pollen sensitization, while 62.9% were solely positive due to IgE cross-reactive molecules from other clinically relevant pollen., Conclusions: Plantain pollen-sensitized patients are multi-reactors demonstrating varying and complex IgE-reactivity profiles. In vivo and in vitro tests using extracts are typically blurred due to the presence of homologous allergens or CCD in grass, birch, or ash pollen. So far, Pla l 1 represents the only indicative marker allergen for the diagnosis of genuine plantain pollen sensitization., (© 2018 S. Karger AG, Basel.)

Background: It has been suggested that latex-specific IgE analysis may lead to false-positive results, especially in patients with pollen allergy. In the present study, the reasons underlying clinically irrelevant latex-specific IgE positivity were investigated., Methods: Thirty patients with latex allergy (group 1), 89 patients sensitised to aeroallergens (group 2a), and 98 healthy individuals without allergy (group 2b) were enrolled. Participants from all 3 groups were subjected to skin prick tests with aeroallergens including latex, latex-specific IgE analysis (ImmunoCAP), and nasal provocation test with latex. All cases demonstrating positive latex-specific IgE also underwent specific IgE tests (ImmunoCAP) with latex profilin, birch pollen profilin, peach lipid transfer protein, and pineapple bromelain as cross-reactive carbohydrate determinants., Results: Comparison of the atopic and healthy control groups showed that the rate of positive latex-specific IgE was significantly higher in group 2a. Latex profilin-, birch pollen profilin-, and bromelain-specific IgE were remarkably higher in group 2a., Conclusion: False positivity to latex-specific IgE in ImmunoCAP analysis may be observed in approximately 19% of patients with pollen allergy. Profilins and bromelain are the main contributors to clinically irrelevant positive latex-specific IgE., (© 2018 S. Karger AG, Basel.)

Background: An association between childhood asthma and IgE sensitization has been established, but our understanding of the genetic and environmental contribution to it is incomplete. Our aim was to estimate the associations and dose-response relationship between asthma and sensitization to airborne allergens in Swedish 9- to 14-year-old twins. Additionally, we aimed to explore the importance of familial confounding from shared genes and environment using co-twin controls., Methods: In the STOPPA cohort, 752 same-sex twin children were screened with Phadiatop® (Thermo Fisher Scientific; Pharmacia, Uppsala, Sweden); if positive further analysis of IgE antibodies to airborne allergens of pets (cat, horse, dog), pollens (birch, timothy, mugwort), mites, and mold were performed. The associations between asthma and airborne allergens were assessed with generalized estimating equations. The co-twin control analysis was performed by conditional logistic regression., Results: Children with positive Phadiatop® had more than doubled odds of asthma (OR 2.53, 95% CI [1.74, 3.70]). Sensitization to pet allergens was associated with increased odds of asthma; for example, cat OR 4.15 (95% CI [2.67, 6.45]), with similar estimates for pollens; for example, birch OR 3.22 (95% CI [2.12, 4.91]). Associations persisted with sensitization as a categorical variable and for trend, indicating a dose-response relationship. Results remained in the co-twin analyses; for example, cat OR 4.75 (95% CI [1.62, 14.0]) and birch OR 5.00 (95% CI [1.45, 17.3])., Conclusion: The association between childhood asthma and sensitization to airborne allergens remains in co-twin analyses, indicating they are not due to confounding from shared environmental or genetic factors.

Platanus acerifolia (P. acerifolia) is an important cause of pollinosis in cities. The use of allergen extracts on patients with allergic diseases is the most commonly applied method to attempt to treat pollinosis. Pla a 3, a non‑specific lipid transfer protein, is a major allergen present in P. acerifolia pollen extracts. In the present study, the Pla a 3 gene was sub‑cloned into a pSUMO‑Mut vector using Stu I and Xho I sites and transformed into the Arctic Express™ (DE3) RP E. coli host strain. The purified Pla a 3 allergen was analyzed by western blotting and the results revealed that the Pla a 3 allergen has the ability to bind IgE in the P. acerifolia pollen of allergic patients' sera. Moreover, the authors predicted the potential B cell epitopes of the Pla a 3 allergen using the DNAStar Protean system, the Bioinformatics Predicted Antigenic Peptides system and the BepiPred 1.0 server. In addition, the T cell epitopes were predicted by the SYFPEITHI database and the NetMHCII‑2.2 server. As a result, two B cell epitopes (35‑45 and 81‑86) and four potential T cell epitopes including 2‑15, 45‑50, 55‑61 and 67‑73 were predicted in the present study. The current results can be used to contribute to allergen immunotherapies and useful in peptide‑based vaccine designs of pollen allergy.

Background: Eligibility to immunotherapy is based on the determination of IgE reactivity to a specific allergen by means of skin prick or in vitro testing. Biomarkers predicting the likelihood of clinical improvement during immunotherapy would significantly improve patient selection., Methods: Proteins were differentially assessed by using 2-dimensional differential gel electrophoresis and label-free mass spectrometry in pretreatment sera obtained from clinical responders and nonresponders within a cohort of 82 patients with grass pollen allergy receiving sublingual immunotherapy or placebo. Functional studies of Fetuin-A (FetA) were conducted by using gene silencing in a mouse asthma model, human dendritic cell in vitro stimulation assays, and surface plasmon resonance., Results: Analysis by using quantitative proteomics of pretreatment sera from patients with grass pollen allergy reveals that high levels of O-glycosylated sialylated FetA isoforms are found in patients exhibiting a strong decrease in rhinoconjunctivitis symptoms after sublingual immunotherapy. Although FetA is involved in numerous inflammatory conditions, its potential role in allergy is unknown. In vivo silencing of the FETUA gene in BALB/c mice results in a dramatic upregulation of airway hyperresponsiveness, lung resistance, and T H 2 responses after allergic sensitization to ovalbumin. Both sialylated and nonsialytated FetA bind to LPS, but only the former synergizes with LPS and grass pollen or mite allergens to enhance the Toll-like receptor 4-mediated proallergic properties of human dendritic cells., Conclusions: As a reflection of the patient's inflammatory status, pretreatment levels of sialylated FetA in the blood are indicative of the likelihood of clinical responses during grass pollen immunotherapy., (Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Background: Serum vitamin D levels have not been studied in children with seasonal allergic rhinitis (SAR). The aim of this study was to evaluate the vitamin D levels of children with SAR and to compare them to levels in healthy children during pollen season., Methods: This study was conducted in 100 children with SAR and 100 healthy controls. Clinical and laboratory evaluations and vitamin D analyses of all the participants were performed between the months of April and July. Pollen sensitization was detected in the patient group using a skin prick test. 25(OH)D 3 levels were compared between the patient and control groups. Associations among the patient 25(OH)D 3 levels and their demographic, clinical, and laboratory characteristics were analyzed., Results: Overall, 72% of the patients were male, the median age was 12.35 years (range: 6-17.8 years), and the median body mass index value was 19.15 (range: 13.6-27.8). There were no differences between the patients and healthy controls in terms of gender, age, or body mass index. The mean levels of 25(OH)D 3 (20.78±6) in patients were higher than those of the controls (17.92±4). In the patient group, no associations were found among 25(OH)D 3 levels, demographic characteristics, atopy test results, atopy history, severity of rhinitis, and the total four symptoms score (all P>.05)., Conclusions: During pollen season, children with SAR may have higher vitamin D levels than healthy controls. The presence of asthma and/or atopic dermatitis in addition to SAR did not change this result., (© 2017 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.)

Allergic inflammation in the upper airways represents a wide-spread health issue: Little is known about whether it increases sensitivity to airborne chemicals thereby challenging established exposure limits that neglect such differences in susceptibility. To investigate the role of pre-existing allergic inflammation, 19 subjects with seasonal allergic rhinitis (SAR) and 18 control subjects with low risk of sensitization were exposed for 4h to ammonia in two concentrations (cross-over design): 2.5ppm (odor threshold) and 0-40ppm (occupational exposure limit: 20ppm TWA). Prior to the whole-body exposure, it was confirmed that subjects with SAR showed persistent inflammation outside the pollen season as indicated by increased exhaled nitric oxide and total immunoglobulin E in serum compared to controls. Despite concentration-dependent increases in chemosensory perceptions and acute symptoms, SAR status did not modulate subjective effects of exposure. Moreover, SAR status did not affect the investigated physiological endpoints of sensory irritation: While eye-blink recordings confirmed weak ocular irritation properties of ammonia at 0-40ppm, this effect was not enhanced in SAR subjects compared to controls. Irrespective of SAR status, exposure to 0-40ppm ammonia did not result in a cortisol stress response, objective nasal obstruction as measured with anterior active rhinomanometry, or an inflammatory response as indexed by substance P, tumor-necrosis-factor α, and high-mobility-group protein 1 in nasal lavage fluid. At least for the malodorous compound ammonia, these results do not support the hypothesis that SAR enhances chemosensory effects in response to local irritants. Before generalizing this finding, more compounds as well as sensitization to perennial allergens need to be investigated., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

Background: Repeated low-dose grass pollen intradermal allergen injection suppresses allergen-induced cutaneous late-phase responses comparably with conventional subcutaneous and sublingual immunotherapy., Objective: We sought to evaluate the efficacy and safety of grass pollen intradermal immunotherapy in the treatment of allergic rhinitis., Methods: We randomly assigned 93 adults with grass pollen-induced allergic rhinitis to receive 7 preseasonal intradermal allergen injections (containing 7 ng of Phl p 5 major allergen) or a histamine control. The primary end point was daily combined symptom-medication scores during the 2013 pollen season (area under the curve). Analysis was by intention to treat. Skin biopsy specimens were collected after intradermal allergen challenges, and late-phase responses were measured 4 and 7, 10, or 13 months after treatment., Results: There was no significant difference in the primary end point between treatment arms (active, n = 46; control, n = 47; median difference, 14; 95% CI, -172.5 to 215.1; P = .80). Among secondary end points, nasal symptoms were worse in the intradermal treatment group, as measured based on daily (median difference, 35; 95% CI, 4.0-67.5; P = .03) and visual analog scale (median difference, 53; 95% CI, -11.6 to 125.2; P = .05) scores. In a per-protocol analysis intradermal immunotherapy was further associated with worse asthma symptoms and fewer symptom-free days. Intradermal immunotherapy increased serum Phleum pratense-specific IgE levels (P = .001) compared with those in the control arm. T cells cultured from biopsy specimens of subjects undergoing intradermal immunotherapy had higher expression of the T H 2 surface marker CRTH2 (P = .04) and lower expression of the T H 1 marker CXCR3 (P = .01), respectively. Late-phase responses remained inhibited 7 months after treatment (P = .03)., Conclusion: Intradermal allergen immunotherapy suppressed skin late-phase responses but was not clinically effective and resulted in worsening of respiratory allergic symptoms., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Mice models suggest epigenetic inheritance induced by parental allergic disease activity. However, we know little of how parental disease activity before conception influences offspring's asthma and allergy in humans., Objective: We aimed to assess the associations of parental asthma severity, bronchial hyperresponsiveness (BHR), and total and specific IgEs, measured before conception vs. after birth, with offspring asthma and hayfever., Methods: The study included 4293 participants (mean age 34, 47% men) from the European Community Respiratory Health Survey (ECRHS) with information on asthma symptom severity, BHR, total and specific IgEs from 1991 to 1993, and data on 9100 offspring born 1972-2012. Adjusted relative risk ratios (aRRR) for associations of parental clinical outcome with offspring allergic disease were estimated with multinomial logistic regressions., Results: Offspring asthma with hayfever was more strongly associated with parental BHR and specific IgE measured before conception than after birth [BHR: aRRR = 2.96 (95% CI: 1.92, 4.57) and 1.40 (1.03, 1.91), respectively; specific IgEs: 3.08 (2.13, 4.45) and 1.83 (1.45, 2.31), respectively]. This was confirmed in a sensitivity analysis of a subgroup of offspring aged 11-22 years with information on parental disease activity both before and after birth., Conclusion & Clinical Relevance: Parental BHR and specific IgE were associated with offspring asthma and hayfever, with the strongest associations observed with clinical assessment before conception as compared to after birth of the child. If the hypothesis is confirmed in other studies, parental disease activity assessed before conception may prove useful for identifying children at risk for developing asthma with hayfever., (© 2017 The Authors. Clinical & Experimental Allergy Published by John Wiley & Sons Ltd.)

Background: Allergen-specific immunotherapy (AIT) with birch pollen generates Bet v 1-specific immunoglobulin (Ig)G 4 which blocks IgE-mediated hypersensitivity mechanisms. Whether IgG 4 specific for Bet v 1a competes with IgE for identical epitopes or whether novel epitope specificities of IgG 4 antibodies are developed is under debate., Objective: We sought to analyze the epitope specificities of IgE and IgG 4 antibodies from sera of patients who received AIT., Methods: 15 sera of patients (13/15 received AIT) with Bet v 1a-specific IgE and IgG 4 were analyzed. The structural arrangements of recombinant (r)Bet v 1a and rBet v 1a &#95;11x , modified in five potential epitopes, were analyzed by circular dichroism and nuclear magnetic resonance spectroscopy. IgE binding to Bet v 1 was assessed by ELISA and mediator release assays. Competitive binding of monoclonal antibodies specific for Bet v 1a and serum IgE/IgG 4 to rBet v 1a and serum antibody binding to a non-allergenic Bet v 1-type model protein presenting an individual epitope for IgE was analyzed in ELISA and western blot., Results: rBet v 1a &#95;11x had a Bet v 1a - similar secondary and tertiary structure. Monomeric dispersion of rBet v 1a &#95;11x was concentration and buffer-dependent. Up to 1500-fold increase in the EC 50 for IgE-mediated mediator release induced by rBet v 1a &#95;11x was determined. The reduction of IgE and IgG 4 binding to rBet v 1a &#95;11x was comparable in 67% (10/15) of sera. Bet v 1a-specific monoclonal antibodies inhibited binding of serum IgE and IgG 4 to 66.1% and 64.9%, respectively. Serum IgE and IgG 4 bound specifically to an individual epitope presented by our model protein in 33% (5/15) of sera., Conclusion and Clinical Relevance: Patients receiving AIT develop Bet v 1a-specific IgG 4 which competes with IgE for partly identical or largely overlapping epitopes. The similarities of epitopes for IgE and IgG 4 might stimulate the development of epitope-specific diagnostics and therapeutics., (© 2016 John Wiley & Sons Ltd.)

Objective: To observe serum levels of periostin, ECP, IgE in the antibiotic enterprise workers, and study the role of periostin, ECP, IgE in the development of allergic inflammation. Methods: 90 cases with asthma or rhinitis were enrolled as disease group, another 117 workers exposed to 7-ACA、6-APA dust without suffering from allergic illness, are chosen as group of dust exposed, and 192 healthy workers who didn't contact dust were chosen as control group. Questionnaires were used to learn their basic information.Lung function was determined with a portable spirometer.The expression levels of periostin、ECP and IgE in serum were measured by enzyme-linked immuno sorbent assay. Results: The exposure group and disease group had significantly lower forced vital capacity (FVC) , forced expiratory volume in 1 second (FEV(l.0)) , and FEV(l.0)/FVC ratio than the control group ( P <0.05) . The disease group had significantly higher eosinophil than the control group ( P <0.05) . Compared with the control group, the exposure group, the disease group, asthma subgroup, rhinitis subgroup of serum periostin and IgE increased, the differences are statistically significant ( P <0.05) . Serum levels of ECP in the workers of asthma subgroup were significantly higher than that in control group ( P <0.05) . Serum expression levels of periostin were positively correlated with IgE, ECP in workers ( P <0.001) , serum levels of periostin were negatively correlated with FEV(1.0) in workers ( P <0.05) . Multiple logistics regression analysis found that exposure to 7-ACA or 6-APA ( OR =3.09, 95% CI : 1.83-5.21) , age>47years ( OR =2.53, 95% CI : 1.22-5.26) , higher ECP ( OR =1.04, 95% CI : 1.01-1.06) were risk factors for increased serum periostin level. Conclusion: Occupational exposure to 7-ACA or 6-APA can result in higher serum periostin level, exposure to 7-ACA or 6-APA, age>47 years, higher ECP are risk factors for increased serum periostin level.

Background: Overlapping seasons and cross-reactivity, especially to grass pollen profilin, can hamper the diagnosis of birch pollen allergy. To identify the primary sensitizing allergen and the clinical relevance of cross-sensitization, we correlated sensitization profiles with in vitro and in vivo tests, symptom scores, and pollen counts., Methods: A total of 433 patients with positive skin prick test (SPT) to birch pollen were analyzed regarding IgE to major birch and grass pollen allergens Bet v 1 and Phl p 1/p 5 and the profilins Bet v 2 and Phl p 12. Subgroups were analyzed by basophil activation test (BAT) and CAP-FEIA-based cross- and self-inhibition tests., Results: A total of 349 patients were sensitized to Bet v 1, 44 patients to both Bet v 1 and Bet v 2, and 15 patients to Bet v 2 only. From Bet v 2-sensitized patients, 40 were also sensitized to Phl p 12. Ex vivo, Bet v 2 and Phl p 12 induced dose-dependent activation in basophils of these patients. Cross- and self-inhibition tests with both allergens confirmed cross-reactivity. However, semiquantitative analysis of SPTs demonstrated markedly increased reactivity to grass compared to birch pollen extract in Bet v 2 only sensitized patients. Accordingly, in most of those patients, clinical symptoms precisely correlated with grass pollen counts., Conclusion: Identification of the clinically relevant and sensitizing allergen needs correlation of actual pollen counts with clinical symptoms and sensitization status to major allergens. Semiquantitative analysis of SPT or BAT and determining profilin-specific IgE can contribute to making the diagnosis., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Introduction: Field studies for allergic rhinitis (AR) commonly have inconsistent allergen concentrations and subject exposure patterns due to varying environmental conditions and subject behaviors. A technical and clinical validation study was conducted for the Allergen BioCube ® using timothy grass to confirm uniform allergen concentration and clinically relevant subject symptom responses., Methods: Allergen concentrations were verified by laser particle counts. Subjects ( N  = 14) with positive skin test reactions and no symptoms at screening received four 3-h timothy grass exposures in the BioCube over consecutive days. Subjects evaluated nasal itching, sneezing, rhinorrhea, and nasal congestion while in the BioCube; Total Nasal Symptom Score (TNSS) was computed. Peak Nasal Inspiratory Flow (PNIF), Peak Expiratory Flow Rate (PEFR), sIgE blood tests, and Nasal Inflammation Score (NIS) were assessed. A correlation analysis was conducted for mean sIgE, skin test, and TNSS., Results: Uniform timothy grass concentrations were achieved in the BioCube, both spatially and temporally, at all subject positions. Mean TNSS increased substantially from pre-exposure levels (0.36 ± 0.74 to 1.86 ± 2.14) to maximums of 7.07 ± 2.76 at 1.5 h and 6.71 ± 2.70 at 3 h BioCube exposure. Twelve (86%) subjects had TNSS increases ≥6 units. PNIF decreased 12-24% from baseline at 3-h BioCube exposure. NIS increased (baseline = 0) to 3.7 (maximum score = 4). A low/moderate correlation ( r  = 0.485) occurred between mean sIgE blood levels and mean skin tests; neither sIgE or skin tests correlated with mean TNSS. However, subjects with high skin test scores or positive blood IgE tended to also have higher TNSS., Conclusions: The Allergen BioCube achieved technical and clinical validation for uniform timothy grass concentration and clinically meaningful AR sign and symptom responses. The Allergen BioCube can be used to assess the efficacy of therapies for reduction of AR signs and symptoms resulting from grass exposure.

Background: Bet v 6, a minor birch pollen allergen, is commercially available for allergen-specific IgE antibody testing. However, there is hardly any literature on the clinical impact of Bet v 6. The aim of the present study was to analyze if testing specific IgE against Bet v 6 can add valuable benefit to the standard diagnostics of birch pollen-sensitized patients, especially in patients with unknown sensitization-eliciting allergens., Methods: Birch pollen-sensitized patients with missing IgE reactivity against Bet v 1, 2, and 4 were tested for specific IgE antibodies against Bet v 6. For comparison, an equal number of randomly selected patients with birch pollen sensitization, regardless of their individual sensitization patterns, were tested for specific anti-Bet v 6., Results: Of 57 cases with missing reactivity against the standard birch pollen allergens, 2 patients were positive for Bet v 6. In the representative sample, 3 patients showed specific IgE against Bet v 6 - resulting in a total prevalence of 5%. None of the Bet v 6-positive patients showed allergic symptoms after exposure to birch pollen or an oral allergy syndrome. An increased prevalence of asthma and a higher degree of sensitization were the only distinctive clinical features in Bet v 6-positive patients., Conclusions: Among birch pollen-sensitized patients, the prevalence of specific IgE against Bet v 6 is low. Further, sensitization to Bet v 6, which shows characteristics of a panallergen, remains clinically silent. Therefore, determination of anti-Bet v 6 is not considered useful in the clinical routine., (© 2017 S. Karger AG, Basel.)

Cannabis sativa (C.sativa) is well-known for its medicinal, industrial and recreational use. However, allergies in relation to Cannabis sativa (C.sativa) are rarely reported. C. sativa is one of the common weeds found in Pakistan and its pollen grains are common in spring and fall season. Although categorized as an aeroallergen, there are limited number of reports regarding allergenic potential in C. sativa. Therefore, the current study is aimed at exploring the IgE- binding potential among the C. sativa pollen in local pollen allergic patients. Initial screening of C. sativa sensitized individuals was carried out by dot blot from the sera of pollen allergic patients. Proteins from the pollen grains were extracted and resolved on 10% gel. Eight bands were visible on gel however only one protein fragment i.e. of 14KDa size was found to bind to IgE as analyzed through protein gel blot analysis. Strong IgE affinity of a 14 kDa protein fragment from C. sativa pollen extract suggests its allergenic potential. Further study is required to find the exact nature of this protein fragment.

Background: The most emblematic members of Urticaceae at allergic risk level are wall pellitories (Parietaria), whereas nettle (Urtica) pollen is considered as poorly allergenic. No allergen from nettle pollen has yet been characterized, whereas 4 are listed for Parietaria pollen by the International Union of Immunological Societies. Clinical and biological profiles of 2 adult men who developed symptoms against nettle pollen and/or leaves were studied., Objective: To characterize the allergic reaction and identify the potential nettle pollen sensitizing allergens., Methods: IgE-mediated reaction to nettle pollen extract was evaluated by skin prick test, immunoassay, nasal provocation, and basophil activation test. To characterize specific nettle pollen allergens, an allergomic (IgE immunoproteomic) analysis was performed combining 1- and 2-dimensional electrophoresis, IgE immunoblots of nettle pollen extract, identification of allergens by mass spectrometry, and database queries., Results: The results of biological and immunochemical analyses revealed that the allergic rhinitis was due to Urtica dioica pollen in both patients. The allergomic analysis of nettle pollen extract allowed the characterization of 4 basic protein allergens: a thaumatin-like protein (osmotin) with a relative molecular mass of 27 to 29 kDa, a pectinesterase (relative molecular mass, 40 kDa), and 2 other basic proteins with relative molecular masses of 14 to 16 kDa and 43 kDa. There is no or only very weak allergen associations between pellitory and nettle pollen., Conclusion: Exposure to nettle pollen can be responsible of allergic symptoms, and several allergens were characterized. Unravelling the allergens of this underestimated allergy might help to improve diagnosis and care for patients, to predict cross-reactivities and design adapted specific immunotherapy., (Copyright © 2016 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Background: Oral allergy syndrome (OAS) is caused by cross-reacting allergens found in pollen, raw fruits, vegetables, and some tree nuts. The major apple allergen, Mal d 1, is a cause of food allergic reactions in birch pollen sensitized patients., Objective: To explore the allergenicity of the most popular and commonly consumed apple cultivars in Poland in patients with birch pollen allergy with or without OAS., Methods: Data were obtained from 46 adults with clinical symptoms of birch allergy and allergic rhinitis or rhinoconjunctivitis. Patients were divided into 2 groups according to the occurrence of OAS to apple. Skin prick tests (SPTs) were performed with pulp from the 11 most popular apple cultivars in Poland. Specific IgE (sIgE) to Bet v 1 was measured by radioallergosorbent test., Results: Patients with OAS had more positive responses to apple SPT vs patients without OAS (odds ratios, 4.8-11.96). Patients with OAS had distinctive responses to apple cultivars. Szara Reneta and Cortland induced positive responses in 50% and 83% of patients, respectively. Patients with OAS vs patients without OAS who were allergic to apples vs nonallergic had 2-fold (P = .008) and 7-fold (P = .03) higher blood concentrations of sIgE Bet v 1, respectively. There were different profiles of correlations of sIgE Bet v 1 with wheal diameter for low and high allergenic cultivars in patients without and with OAS., Conclusion: We noted a substantial role for Bet v 1 sensitization in the allergic response based on evaluation of the allergenicity of 11 apple cultivars. The sIgE Bet v 1 and SPT results of patients with and without OAS allowed differentiation between low and high allergenic cultivars., (Copyright © 2016 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)