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Sensitization to lipid transfer protein (LTP), the most frequent cause of food allergy in southern Europe, still shows several controversial, but also intriguing, aspects. Some of these include the degree of cross-reactivity between LTPs from botanically distant sources, the definition of risk factors, the role of some cofactors, clinical outcomes, geographical differences and the identification of the primary sensitizer in different areas. This review article tries to analyse and comment on these aspects point by point suggesting some explanatory hypotheses with the final scope to stimulate critical thoughts and elicit the scientific discussion about this issue in the readership.

BACKGROUND Clinically complex phenotypes require more and more sophisticated and comprehensive diagnostic approaches, able to discriminate genuine sensitizations from cross-reactivity. Interpretative complexity of multiplex diagnostic arrays has somewhat limited their diffusion. This study compares two currently available methods, namely ISAC® test and ALEX2® test. METHODS In total, 140 allergic individuals, with a history of atopic dermatitis, adverse food reactions, allergic rhinitis and/or bronchial asthma were studied by Allergy Explorer-ALEX2® macroarray and ImmunoCAP ISAC112® . Lin's concordance correlation coefficient, intraclass correlation coefficient and Bland-Altman plots were used to verify the agreement between continuous values. Cohen's kappa coefficient (k) was assessed for the molecules available in both tests. The degree of relationship was analysed using Spearman's correlation (quantitative variables) and Pearson's χ2 or Fisher's exact test (categorical variables). RESULTS A substantial agreement (κ = 0.795) was observed between the two methods with 94,3% concordant results when results were dichotomized as negative or positive, but if double-negative results were discarded, the agreement dropped to 71%. Conversely, little or no concordance was observed comparing raw data. Considering the 102 molecules shared by both systems, 28/102 (27%) showed an almost perfect agreement (k > 0.81), and concordance was good (k > 0.61) in a further 32 (31%) cases. A perfect to substantial agreement was observed by comparing species-specific aeroallergens. Heterogeneous results emerged comparing panallergens (co-recognition ranging from 30% for tropomyosin/serum albumins to 70% for PR-10/profilin). The correlation among LTP, profilin and PR-10 assayed with ISAC® was better than ALEX2® , but the latter identified more positive cases due to the wider number of molecules available. The CCD blocker provided by ALEX® test abolishes the carbohydrate determinants signal in 60% of the 33 cases reactive to MUXF3 on the ISAC® test. CONCLUSION Despite the excellent concordance of the species-specific markers, the analysis of the panallergens provided in both methods suggests a better performance of the ISAC® test on those components, while the ALEX2® test, which includes a larger number of allergens, allowing a broader molecular detection.

Background: The so-called “thunderstorm asthma” (TA) is an uncommon but dramatic outbreak of asthma attacks occurring during a thunderstorm in the pollen and moulds season. Mechanisms which make the pollen able to enter the deeper airways and provoke severe asthma symptoms are still unclear. Objective: To test the hypothesis that sub-pollen particles (SPPs) originated from the rupture by an osmotic shock of pollen associated with TA contain allergens. Methods: After hydration, SPPs released from pollen grains of grass, pellitory, olive, cypress, ragweed and birch were isolated and determined by microscopy. Allergens were determined by in vitro ELISA inhibition tests indirectly using the sera from 10 polyreactive patients. An inhibition 75% complete. Results: The inhibition experiments showed that the SPPs from birch and cypress were unable to inhibit serum IgE reactivity to Bet v 1 and Cup a 1, respectively. Ragweed SPPs inhibited ragweed pollen extract and Amb a 1 by 75.8±0.11% and 81.2±0.15%, respectively. Olive and pellitory SPPs retained almost the whole IgE-binding capability in all cases tested. Grass SPPs inhibited 32±0.06% of Lolium perenne Lol p 1 and 65% of Phleum pratense extracts, but results were highly variable for individual allergens (97.5%-0.03% for Phl p 2, 45.3±0.12% for Phl p 5, 24.7±0.22% for Phl p 6, and 38.3±0.2% for Phl p 1). Conclusions: Inhibition experiments confirm the hypothesis that SSPs obtained after the osmotic shock of pollen involved in TA, namely grass, pellitory and olive tree pollen, contain allergens and therefore they can induce severe asthma attacks during thunderstorms.

Hymenoptera stings mostly cause acute urticaria but we describe a case of CU after wasp stings which remitted during venom immunotherapy. IgE‐mechanisms have not been fully clarified in CU, except for isolated circumstances. In our case immunotherapy has played a positive role reducing immune cells reactivity and improving urticaria symptoms.

BACKGROUND: Severe Covid-19 is associated with a cytokine storm leading to the hyper-expression of cytokines such as IFN-gamma, TNF-alpha, and IL-6, which may be responsible for most severe symptoms and signs of the disease. The immune response of the atopic patient is mostly Th2-oriented, associated with the expression of cytokines such as IL-4, IL-5, and IL-13, and atopic patients express fewer ACE-2 receptors than non-atopic individuals do. We assessed whether atopic status may protect from the most severe consequences of Covid-19. METHODS: Atopic status along with co-factors such as diabetes, hypertension, coronary heart disease, and thrombosis was investigated in severe Covid-19 patients admitted to different Italian hospitals. Patients were classified as having a mild, severe, or very severe disease based on the need of respiratory assistance; severity was plotted against the different co-factors and underwent multivariate analysis. RESULTS: 531 adults aged 25 - 100 years were studied; 57 (11.7%) were atopic. Atopic status showed a significant association with a milder disease irrespective of all other co-factors considered (p

This evidence- and consensus-based guideline was developed following the methods recommended by Cochrane and the Grading of Recommendations Assessment, Development and Evaluation (GRADE) working group. The conference was held on 1 December 2016. It is a joint initiative of the Dermatology Section of the European Academy of Allergology and Clinical Immunology (EAACI), the EU-founded network of excellence, the Global Allergy and Asthma European Network (GA²LEN), the European Dermatology Forum (EDF) and the World Allergy Organization (WAO) with the participation of 48 delegates of 42 national and international societies. This guideline was acknowledged and accepted by the European Union of Medical Specialists (UEMS). Urticaria is a frequent, mast cell-driven disease, presenting with wheals, angioedema, or both. The lifetime prevalence for acute urticaria is approximately 20%. Chronic spontaneous urticaria and other chronic forms of urticaria are disabling, impair quality of life and affect performance at work and school. This guideline covers the definition and classification of urticaria, taking into account the recent progress in identifying its causes, eliciting factors and pathomechanisms. In addition, it outlines evidence-based diagnostic and therapeutic approaches for the different subtypes of urticaria.

Nine hundred and thirty-nine rPla a 1, nPla a 2, and rPla a 3 ImmunoCAP ISAC reactors were studied. nPla a 2pos MUXF3pos but Pla a 1/2neg subjects were excluded from the study because they were cross-reactive carbohydrate determinant reactors. Among the 764 remaining participants, 71.9% were Pla a 3pos, 54.1% Pla a 2pos, and 10.9% Pla a 1pos. Among Pla a 3 reactors, 89.6% were Pru p 3pos and 86.8% Jug 3pos, but the strongest IgE recognition relationship was observed between Pla a 3 and Jug r 3. Distinctive clinical subsets could be documented among plane tree-allergic patients. Pla a 3 reactors had both local and systemic food-induced reactions, but lower past respiratory symptoms occurrence. Pla a 2 reactivity was associated with respiratory symptoms but inversely related to systemic reactions to food. Cosensitization to Pla a 2 and Pla a 3 was associated with a lower past incidence of severe food-induced reactions.

This guideline is the result of a systematic literature review using the 'Grading of Recommendations Assessment, Development and Evaluation' (GRADE) methodology and a structured consensus conference held on 28 and 29 November 2012, in Berlin. It is a joint initiative of the Dermatology Section of the European Academy of Allergy and Clinical Immunology (EAACI), the EU-funded network of excellence, the Global Allergy and Asthma European Network (GA(2)LEN), the European Dermatology Forum (EDF), and the World Allergy Organization (WAO) with the participation of delegates of 21 national and international societies. Urticaria is a frequent, mast cell-driven disease, presenting with wheals, angioedema, or both. the life-time prevalence for acute urticaria is approximately 20%. Chronic spontaneous urticaria and other chronic forms of urticaria do not only cause a decrease in quality of life, but also affect performance at work and school and, as such, are members of the group of severe allergic diseases. This guideline covers the definition and classification of urticaria, taking into account the recent progress in identifying its causes, eliciting factors and pathomechanisms. in addition, it outlines evidence-based diagnostic and therapeutic approaches for the different subtypes of urticaria. This guideline was acknowledged and accepted by the European Union of Medical Specialists (UEMS). Charite, Allergy Ctr Charite, Dept Dermatol & Allergy, D-10117 Berlin, Germany Med Univ Graz, Dept Dermatol, Graz, Austria Clin San Carlo, Allergy Clin, Paderno Dugnano, MI, Italy Univ Southern Denmark, Odense Univ Hosp, Dept Dermatol & Allergy Ctr, Odense, Denmark Med Univ Silesia, Allergol & Clin Immunol Katowice, Dept Internal Dis, Zabrze, Poland Univ Genoa, IRCCS AOU SanMartino, Genoa, Italy Universidade Federal de São Paulo, Dept Clin Immunol & Allergy, São Paulo, Brazil Univ Autonoma Barcelona, Hosp Mar Parc Salut Mar, E-08193 Barcelona, Spain Dr DY Patil Med Coll & Hosp, Dept Dermatol, Navi Mumbai, India Fac Med, Dermatol Clin, Coimbra, Portugal Univ Hosp, Coimbra, Portugal Guys & St Thomas Hosp NHS Fdn Trust, St Johns Inst Dermatol, London, England Ctr Appl Res Allergy Quebec, Quebec City, PQ, Canada Hiroshima Univ, Inst Biomed & Hlth Sci, Dept Dermatol, Hiroshima, Japan Med Univ S Carolina, Dept Med, Div Pulm & Crit Care Med, Charleston, SC USA Hannover Med Sch, Dept Dermatol & Allergy, Hannover, Germany Pantai Hosp Kuala Lumpur, Dept Paediat, Kuala Lumpur, Malaysia Univ Hosp Tenon, Dept Dermatol & Allergy, Paris, France Johns Hopkins Asthma & Allergy Ctr, Baltimore, MD USA Ctr Medicodocente Trinidad, Allergy & Clin Immunol Dept, Caracas, Venezuela Univ Hosp, Dept Dermatol, Allergy Unit, Zurich, Switzerland Univ Manitoba, Dept Pediat & Child Hlth, Winnipeg, MB R3T 2N2, Canada Univ Manitoba, Dept Immunol, Winnipeg, MB, Canada Univ Med Ctr Mainz, Dept Dermatol, Mainz, Germany Univ Toronto, Div Allergy & Clin Immunol, Toronto, ON, Canada Technion, Fac Med, Bnai Zion Med Ctr, Haifa, Israel Univ Bari, Dept Biomed Sci & Human Oncol, Unit Dermatol & Venereol, Bari, Italy Peking Univ, Hosp 1, Dept Dermatol, Beijing 100871, Peoples R China Universidade Federal de São Paulo, Dept Clin Immunol & Allergy, São Paulo, Brazil Web of Science

Chronic urticaria (CU) is a widespread skin disease, characterized by the recurrence of transient wheals and itch for more than 6 weeks. Besides autoimmune mechanisms, coagulation factors, in particular tissue factor and thrombin, might also participate in the disease pathophysiology. Tissue factor expressed by eosinophils can induce activation of blood coagulation generating thrombin which in turn can increase vascular permeability both directly, acting on endothelial cells, and indirectly, inducing degranulation of mast cells with release of histamine, as demonstrated in experimental models. D-dimer, a fibrin degradation product, generated following activation of the coagulation cascade and fibrinolysis, has been found to be increased during urticaria exacerbations; moreover, it has been proposed as a biomarker of severity and resistance to H1-antihistamines in CU patients. The possible role of coagulation in CU is also supported by case reports, case series and a small controlled study showing the efficacy of anticoagulant therapy in this disease. The purpose of this review was to summarize the available data on the possible contribution of coagulation to the pathophysiology of CU focusing on clinical aspects and possible future therapeutic developments.

An autoimmune subset of chronic spontaneous urticaria is increasingly being recognized internationally, based on laboratory and clinical evidence that has accrued over the last 20 years. This evidence has been reviewed by a taskforce of the Dermatology section of the European Academy of Allergy and Clinical Immunology. Functional autoantibodies in chronic urticaria (CU) patient sera have been demonstrated against IgE and FcεRIα by basophil and mast cell histamine release assays and by basophil activation assays. Antibody specificity has been confirmed by immunoassay, but there is a poor correlation between functionality and immunoreactivity. Approximately 25% of CU patients have a positive basophil histamine release assay and show autoreactivity (a positive autologous serum skin test), whereas 50% are negative regarding both. Functionality of CU sera appears to be complement dependent on mast cells but not exclusively on basophils. Basophil activation by CU sera is predominantly restricted to IgG1 and IgG3 subclasses. Circumstantial evidence for CU being an autoimmune disease comes from an observed association with other autoimmune diseases, a strong association between serum functionality and HLA-DR4 haplotype and the good response of CU patients to immunotherapies. It was proposed that a study should be undertaken to prospectively validate potentially relevant clinical criteria (from the history, examination and routinely available clinical investigations) against a new 'gold standard' for the diagnosis of ACU (positive autoreactivity, functional bioassay and immunoassay) to define preliminary criteria sets for the diagnosis of ACU based on clinical and laboratory features with highest individual sensitivity and specificity.

Background Most cases of beer allergy reported so far have been associated with hypersensitivity to the non-specific lipid transfer protein (LTP). In view of the marked differences in brewing processes we assessed IgE reactivity as well as tolerance to many different beers in an allergic patient. Methods A 45 year-old man hypersensitive to grass pollen, cat dander and Alternaria tenuis with a history of urticaria and dyspnoea after drinking beer and a weak skin reactivity to commercial corn extract was studied. The patient underwent SPT with 36 different brands of beer and an open challenge with those scoring negative was performed. An immunoblot analysis was carried out using 2 SPT-positive beers, 2 SPT-negative beers, and barley, wheat, and maize extracts using both patient's serum and a maize LTP-specific in-house developed polyclonal antibody from rabbit. Further, the immune reactive LTP of one beer was separated by HPLC and the chromatogram was compared to that of purified maize LTP. Results Beer SPT scored positive in 30/36 cases. The immunoblot analysis showed IgE reactivity at about 10 kDa against the two SPT-positive beers and against maize with both patient's serum and the polyclonal anti-LTP rabbit serum, whereas the two SPT-negative beers, and barley extract scored negative. The immunodetected protein co-migrated with maize LTP. Conclusion In beer-allergic patients the diagnostic workup may point to the detection of some tolerated products that can be consumed risk-free.

Background: Pollutants may affect pollen allergenicity and thus the prevalence of allergies. Although a few studies are available in literature, the connection between pollution and the allergenic potential of pollen has yet to be clearly defined. The objective of this study was to evaluate the effect of traffic-related pollution on the allergenicity of ragweed (Ambrosia artemisiifolia L.) pollen through a field-based experiment. Methods: Mature pollen grains were collected from ragweed plants grown along main roadsides and in vegetated areas of Po river plain. The percentage of subpollen particle-releasing grains (SPPGs) was evaluated immediately after sampling by microscope and image analysis. Immunochemistry and LC-MS/MS were applied to assess the whole allergenicity and the allergen pattern characterizing the different pollen samples. Results: No statistical difference was detected in the percentage of SPPGs among pollen samples. Specifically, after hydration, the mean percentage was very low (>4%) in all the samples, regardless of the site of origin. On the contrary, pollen collected along high-traffic roads showed a higher whole allergenicity than pollen from low-traffic roads and vegetated areas which showed a reactivity similar to that of the commercial pollen 'Allergon', used as a standard. The detected higher allergenicity levels were attributed to both quantitative and qualitative differences in allergen pattern. Conclusion: Our findings show that pollen collected at different sites contains different amount and number of allergens and suggest that traffic-related pollution enhances ragweed pollen allergenicity, which may contribute to the increasing prevalence of ragweed allergy in Lombardy plain. © 2012 John Wiley & Sons A/S.

To cite this article: Cochrane SA, Salt LJ, Wantling E, Rogers A, Coutts J, Ballmer-Weber BK, Fritsche P, Fernandez-Rivas M, Reig I, Knulst A, Le T-M, Asero R, Beyer K, Golding M, Crevel R, Clare Mills EN, Mackie AR. Development of a standardized low-dose double-blind placebo-controlled challenge vehicle for the EuroPrevall Project. Allergy 2012; 67: 107–113. Abstract Background: Double-blind placebo-controlled food challenge (DBPCFC) is the gold standard for diagnosing food allergy. Standardized materials and protocols are essential for comparing DBPCFC results for multicentre studies such as EuroPrevall. This required the development and piloting of a standardized vehicle and low-dose protocol for confirming food allergy and determination of minimum eliciting doses (MEDs). Methods: A low-dose DBPCFC protocol was developed, with eight titrated protein doses from 3 μg to 1 g. This was delivered using a simple, microbiologically stable food base incorporating allergenic food ingredients manufactured at three sites and centrally distributed to clinical centres. Allergen blinding was assessed by a professional sensory testing panel using a triangle test. Homogeneity and allergen content were confirmed by ELISA and clinical efficacy was assessed in a pilot study, using celeriac and hazelnut as exemplars. Results: Celeriac and hazelnut ingredients were sufficiently blinded in the dessert. The dessert meals were successfully piloted with hazelnut in allergy clinics in Spain, the Netherlands and Italy and with celeriac and hazelnut in Zurich. The challenges elicited a range of subjective and objective reactions ranging in severity from mild itching of the oral mucosa to bronchospasm. Conclusions: A standardized challenge vehicle proven to sufficiently blind processed, powdered hazelnut and celeriac ingredients and that can be reproducibly manufactured has been developed. This pilot study shows that the vehicle is promising for the confirmation of food allergy and determination of MEDs in adults and children with body weight >28.8 kg (approximately 7–11 years old).

To cite this article: Bossi F, Frossi B, Radillo O, Cugno M, Tedeschi A, Riboldi P, Asero R, Tedesco F, Pucillo C. Mast cells are critically involved in serum-mediated vascular leakage in chronic urticaria beyond high-affinity IgE receptor stimulation. Allergy 2011; 66: 1538–1545. Abstract Background: Chronic urticaria (CU) is one of the most common skin disorders whose pathogenic mechanisms are not fully clarified. Autoimmune aetiology can be ascribed to 45% of patients with CU, and basophil histamine release is positive in 40% of cases. Our aim was to use a novel approach to evaluate the serum permeabilizing effect to identify the mediators of endothelial cell (EC) leakage and to define the role of mast cells (MCs) in the process. Methods: Permeabilizing activity of sera from 19 patients with CU and 11 healthy blood donors was evaluated by measuring serum-induced degranulation of two MC lines, expressing (LAD2) or lacking (HMC-1) the IgE receptor. Mast cell supernatant (SN) was then incubated with an EC monolayer, and endothelial permeability was evaluated by Fluorescein isothiocyanate–bovine serum albumin leakage in a transwell system. Results: All 19 patient sera failed to induce direct EC leakage, but 15/19 and 17/19 promoted degranulation of HMC-1 and LAD2, respectively. Interestingly, 85% of autologous serum skin test-negative sera were able to cause MC degranulation. Also, 17/19 SNs from HMC-1 and all SNs from LAD2 incubated with CU sera increased endothelial permeability. Endothelial cell leakage remained unchanged after Ig depletion and was prevented by antihistamine, platelet-activating factor or leukotriene antagonist. Conclusions: Our study shows that CU sera are able to degranulate MCs through an IgE- and IgG-independent mechanism. The nature of histamine-releasing factors involved is still unclear, but our finding opens new ways to the understanding of the pathogenesis of CU, particularly in patients not showing circulating autoantibodies to FceRI or IgE.

To cite this article: Asero R., Tedeschi A., Cugno M. Markers of autoreactivity, coagulation and angiogenesis in patients with nonallergic asthma. Allergy 2011; 66: 1339–1344. Abstract Background: Patients with nonallergic asthma frequently show autoreactivity as do subjects with chronic urticaria (CU). Activation of the coagulation cascade and hyper-expression of vascular endothelial growth factor (VEGF) were recently found in CU, and there is sparse evidence that the same may occur in asthma. Objective: To investigate autoreactivity, activation of the coagulation cascade, and expression of VEGF in patients with nonallergic asthma. Methods: Twenty-one adults with nonallergic asthma underwent autologous plasma skin test (APST) and the measurement of plasma levels of the prothrombin fragment F1+2, D-dimer, VEGF, and the inflammatory marker C-reactive protein (CRP). Twenty-one healthy sex- and age-matched subjects served as normal controls. Results: The APST scored positive in 19 of 21 (90%) patients vs 0 controls. Mean fragment F1+2 plasma levels were significantly higher in patients with asthma (267 ± 243 pM) than in controls (150 ± 51 pM; P = 0.0001). Similarly, plasma levels of both D-dimer and VEGF were significantly higher in patients than in controls (D-dimer: 2364 ± 1467 vs 1301 ± 525 pM; P = 0.0001; VEGF: 1721 ± 2566 vs 76 ± 375 fM; P = 0.0001). A trend toward increased levels of F1+2, D-dimer, VEGF, and CRP was found in patients with a more severe disease according to GINA classification. Conclusion: Nonallergic asthma is characterized by autoreactivity as well as increased coagulation and angiogenesis markers, which are known to enhance vascular permeability. The presence of circulating vasoactive factors may be relevant to understand the disease pathophysiology and to detect novel therapeutic strategies in nonallergic asthma.

To cite this article: Heffler E, Nebiolo F, Asero R, Guida G, Badiu I, Pizzimenti S, Marchese C, Amato S, Mistrello G, Canaletti F, Rolla G. Clinical manifestations, co-sensitizations, and immunoblotting profiles of buckwheat-allergic patients. Allergy 2011; 66: 264–270. Abstract Background: Buckwheat allergy is a rare food allergy in Europe and North America, whereas it is often described and studied in Asia. The aim of this study was to describe a series of patients with proven buckwheat allergy evaluated in an Italian allergy clinic. Co-sensitization to other food and inhalant allergens and immunoblotting profiles of buckwheat-allergic patients were studied. Methods: Patients with suspected buckwheat allergy who attended the allergy clinic between January 1, 2006, and September 30, 2008, were evaluated. All patients underwent skin prick tests for a standard panel of inhalant and food allergens, prick-by-prick with buckwheat flour, buckwheat-specific IgE determinations, and double-blind placebo-controlled food challenge (DBPCFC) with buckwheat flour. Immunoblotting with buckwheat flour extract was performed on sera from buckwheat-allergic patients. Results: Among 72 patients with suspected buckwheat allergy, 30 (41.7%) were sensitized to buckwheat and 24 had a positive DBPCFC. The mean buckwheat IgE level was 6.23 kUA/l (range, 0.16 to >100 kUA/l). Several IgE-binding proteins were identified and grouped into three patterns: a 16-kDa band in patients with predominantly gastrointestinal symptoms with grass and wheat flour co-sensitization, a 25-kDa band in patients with predominantly cutaneous symptoms and a low frequency of co-sensitization, and a 40-kDa band in patients with anaphylaxis and a low frequency of co-sensitization. Conclusions: Buckwheat allergy is an emerging food allergy in Italy. We identified three distinct patterns of clinical and laboratory characteristics, suggesting that specific allergens could be more frequently associated with clinical manifestations of different severity.

Background Skin reactivity to the intradermal injection of autologous serum (autologous serum skin test – ASST) and/or plasma (autologous plasma skin test – APST) is thought to identify chronic urticaria (CU) patients with an autoimmune/autoreactive disease. Immune-mediated inflammation and coagulation are strictly linked, and coagulation activation has been described in CU patients as shown by the elevation of plasma prothrombin fragment F1 + 2 and, in severe cases, of d-dimer as well. Objective The aim of this study was to evaluate whether the coagulation cascade is activated in APST-negative CU patients as it has been described in CU patients with an autoreactive disease. Methods A total of 43 adults with CU (M/F 15/28; mean age 43.5 years; 16 APST-negative patients and 27 APST-positive) and 30 healthy subjects were studied. Prothrombin fragment F1 + 2, d-dimer and C-reactive protein (CRP) plasma levels were measured by ELISA. Results Prothrombin fragment F1 + 2 and d-dimer were elevated in seven of 16 APST-negative CU patients. The activation of the coagulation cascade was associated with disease severity. Men were more prevalent in idiopathic than in autoreactive CU patients (M/F: 10/6 vs. 5/22; P

Summary Background Matrix metalloproteinase-9 (MMP-9) is an endopeptidase produced by many inflammatory cells that has been found in increased amounts in plasma from patients with chronic urticaria (CU). Objective To evaluate plasma levels of MMP-9 and its tissue inhibitor of metalloproteinase-1 (TIMP-1) in CU patients in relation with disease severity, C-reactive protein (CRP) and circulating histamine-releasing factors. Methods Fifty-two consecutive CU patients were included in the study and disease activity was graded from 0 to 3. Plasma MMP-9, TIMP-1 and CRP levels were measured by enzyme immunoassays. Circulating histamine-releasing factors were assessed using in vivo (autologous serum skin test) and in vitro (basophil histamine release) tests. Seven CU patients were studied both during active disease and during remission. Thirty healthy subjects were used as normal controls. Results Plasma levels of MMP-9, TIMP-1 and CRP were significantly higher in CU patients than in healthy controls (P=0.0001, 0.003 and 0.005, respectively) and a trend towards a higher MMP-9/TIMP-1 molar ratio was found (P=0.051). A significant correlation was found between plasma MMP-9 levels and urticaria severity score (r=0.48, P

To cite this article: Aina R, Asero R, Ghiani A, Marconi G, Albertini E, Citterio S. Exposure to cadmium-contaminated soils increases allergenicity of Poa annua L. pollen. Allergy 2010; 65: 1313–1321. Abstract Background: Pollution is considered as one main cause for the increase of allergic diseases. Air pollutants may cause and worsen airway diseases and are probably able to make pollen allergens more aggressive. Previous studies looked at traffic-related air pollution, but no data about the effects of polluted soils on pollen allergens are available. We aimed to assess the effects of plant exposure to cadmium-contaminated soil on allergenicity of the annual blue grass, Poa annua L, pollen. Methods: Poa plants were grown in soil contaminated or not contaminated (control) with cadmium. At flowering, mature pollen was analyzed by microscopy, to calculate the percentage of pollen grains releasing cytoplasmic granules, and by proteomic techniques to analyze allergen proteins. Allergens were identified by sera from grass pollen–allergic patients and by mass spectrometry. Results: Pollen from Cd-exposed plants released a higher amount of allergenic proteins than control plants. Moreover, Cd-exposed pollen released allergens-containing cytoplasmic grains much more promptly than control pollen. Group 1 and 5 allergens, the major grass pollen allergens, were detected both in control and Cd-exposed extracts. These were the only allergens reacting with patient’s sera in control pollen, whereas additional proteins strengthening the signal in the gel region reacting with patient’s sera were present in Cd-exposed pollen. These included a pectinesterase, a lipase, a nuclease, and a secretory peroxydase. Moreover, a PR3 class I chitinase-like protein was also immunodetected in exposed plants. Conclusion: Pollen content of plants grown in Cd-contaminated soils is more easily released in the environment and also shows an increased propensity to bind specific IgE.

This guideline, together with its sister guideline on the classification of urticaria (Zuberbier T, Asero R, Bindslev-Jensen C, Canonica GW, Church MK, Gimenez-Arnau AM et al. EAACI/GA(2)LEN/EDF/WAO Guideline: definition, classification and diagnosis of urticaria. Allergy 2009;64: 1417-1426), is the result of a consensus reached during a panel discussion at the Third International Consensus Meeting on Urticaria, Urticaria 2008, a joint initiative of the Dermatology Section of the European Academy of Allergology and Clinical Immunology (EAACI), the EU-funded network of excellence, the Global Allergy and Asthma European Network (GA(2)LEN), the European Dermatology Forum (EDF) and the World Allergy Organization (WAO). As members of the panel, the authors had prepared their suggestions regarding management of urticaria before the meeting. The draft of the guideline took into account all available evidence in the literature (including Medline and Embase searches and hand searches of abstracts at international allergy congresses in 2004-2008) and was based on the existing consensus reports of the first and the second symposia in 2000 and 2004. These suggestions were then discussed in detail among the panel members and with the over 200 international specialists of the meeting to achieve a consensus using a simple voting system where appropriate. Urticaria has a profound impact on the quality of life and effective treatment is, therefore, required. The recommended first line treatment is new generation, nonsedating H(1)-antihistamines. If standard dosing is not effective, increasing the dosage up to four-fold is recommended. For patients who do not respond to a four-fold increase in dosage of nonsedating H(1)-antihistamines, it is recommended that second-line therapies should be added to the antihistamine treatment. In the choice of second-line treatment, both their costs and risk/benefit profiles are most important to consider. Corticosteroids are not recommended for long-term treatment due to their unavoidable severe adverse effects. This guideline was acknowledged and accepted by the European Union of Medical Specialists (UEMS).

Serological tests for immunoglobulin G4 (IgG4) against foods are persistently promoted for the diagnosis of food-induced hypersensitivity. Since many patients believe that their symptoms are related to food ingestion without diagnostic confirmation of a causal relationship, tests for food-specific IgG4 represent a growing market. Testing for blood IgG4 against different foods is performed with large-scale screening for hundreds of food items by enzyme-linked immunosorbent assay-type and radioallergosorbent-type assays in young children, adolescents and adults. However, many serum samples show positive IgG4 results without corresponding clinical symptoms. These findings, combined with the lack of convincing evidence for histamine-releasing properties of IgG4 in humans, and lack of any controlled studies on the diagnostic value of IgG4 testing in food allergy, do not provide any basis for the hypothesis that food-specific IgG4 should be attributed with an effector role in food hypersensitivity. In contrast to the disputed beliefs, IgG4 against foods indicates that the organism has been repeatedly exposed to food components, recognized as foreign proteins by the immune system. Its presence should not be considered as a factor which induces hypersensitivity, but rather as an indicator for immunological tolerance, linked to the activity of regulatory T cells. In conclusion, food-specific IgG4 does not indicate (imminent) food allergy or intolerance, but rather a physiological response of the immune system after exposition to food components. Therefore, testing of IgG4 to foods is considered as irrelevant for the laboratory work-up of food allergy or intolerance and should not be performed in case of food-related complaints.

Background Recently, distinct studies have shown that: (a) chronic idiopathic urticaria (CIU) is autoimmune in 30–50% of cases; (b) in patients with CIU the autologous serum skin test is inhibited by heparin; and (c) basophil histamine release induced in vitro by CIU sera maybe complement-dependent. Objective To carry out a comprehensive clinical and serological study on CIU based upon these observations. Methods Three hundred and six adults with CIU underwent intradermal (ID) test with autologous serum; 57 of them with autologous heparinized plasma as well. Sera from 121 patients (plasmas from 17) were employed to induce in vitro histamine release from basophils of normal donors. The effects of heating (56 °C, 60 min), filtration through membrane, and preincubation with heparin were evaluated as well. Results Autologous serum and plasma induced a weal and flare reaction in 205 out of 306 (205/306; 67%) and in 8/57 (14%) patients, respectively. Positive plasma skin tests were observed only in patients showing strongly positive serum skin tests. Plasma did not elicit any skin reaction in 3/3 patients with dermatographism who showed a positive intradermal test with saline. Sera from 20/121 (16.5%) patients induced significant histamine release from basophils of normal donors. 19/20 sera were from patients with a positive intradermal test; thus, basophil histamine release assay was positive in 19/87 (21.8%) patients with a positive serum skin test. Heating at 56 °C × 1 h markedly reduced the histamine-releasing activity of both serum and plasma from in vitro reactors. Ultrafiltered fractions > 100 kDa of both sera tested retained the histamine-releasing activity, whereas fractions

Background: Allergens in Plantago lanceolata have not been characterized yet. The objective was to characterize some plantain-pollen allergens and to investigate the cross-reactivity between plantain and grass pollens. Methods: Sera from four patients monosensitive to plantain pollen and from eight grass-pollen-allergic patients showing strong skin reactivity to plantain pollen in the skin prick test (SPT) underwent immunoblot analysis with both Plantago and grass mix extract. Moreover, immunoblot inhibition experiments were done with grass mix extract as inhibitor. Results: All four sera from plantain-allergic patients reacted to two distinct bands at 17 and 19 kDa, and 2/4 sera showed further reactivity to a 40-kDa protein, which in one case represented the most prominent IgE-binding allergen. Plantain-monosensitive subjects did not show any reactivity to grass-pollen extract, and preabsorption of their sera with grass-pollen extract did not cause any loss of reactivity to plantain pollen. Sera from all eight grass-pollen-allergic controls reacted to a 30-kDa protein in plantain pollen, and some sera showed cross-reactivity to higher and lower molecular-weight structures as well. In all cases, plantain reactivity was totally abolished by preabsorption of sera with grass-pollen extract. A preliminary investigation by immunoblot showed that polyclonal IgG anti-Phl p 5 (but not polyclonal Phl p 1) from rabbit reacted to a 30-kDa protein in plantain pollen. Conclusions: Three specific allergens (of 17, 19, and 40 kDa, respectively) have been detected in plantain pollen. Further studies on a larger number of patients will determine whether these proteins may be considered major allergens. Cross-reactivity between grass and plantain pollen is mainly caused by a 30-kDa protein in plantain pollen. Group 5 grass-pollen allergen is probably responsible for most grass/plantain cross-reactivity.

Background Most patients with birch pollen allergy report oral allergy symptoms after eating fresh apples and other vegetable foods. Major birch pollen and apple allergens, Bet v 1 and Mal d 1, are highly homologous; as a consequence, pollen-specific immunotherapy (SIT) might be expected to improve apple hypersensitivity. Objective To evaluate the clinical and immunological effects of birch pollen SIT on oral allergy syndrome (OAS) induced by apples. Methods A prospective study carried out in 49 birch pollen-sensitive patients with apple-induced OAS who received injection immunotherapy for 12, 24, or 36 months. Twenty-six patients not submitted to SIT and followed up for 12–48 months were used as controls. Both SPT and open oral challenges with fresh golden delicious apple were performed, as well as specific IgE measurements, before and after SIT. Results Forty-one patients (84%) vs no control (0%) reported a significant reduction (50–95%) or a total disappearance (100%) of OAS symptoms after SIT (P

SOME patients with chronic idiopathic urticaria (CIU) do not respond to antihistamines and require long-term steroid administration or even cyclosporin to control their disease. Recently, several studies reported good results with antileukotriene drugs in CIU (1–4); however, a prospective investigation aiming to detect which patients are more likely to benefit from antileukotriene treatment has not been carried out yet. Twelve consecutive patients with unremitting, steroid-dependent urticaria were studied. Clinical features and results of the main laboratory investigations are shown in Table 1. Montelukast 10 mg once a day (n=5) or zafirlukast 20 mg twice a day (n=7) was started. The usual therapies were not changed at the start of the scheduled 3-week treatment period, but patients were left free to reduce or withdraw antihistamine treatment in the case of a remission of the disease. Patients recorded both the severity of urticaria and the drugs used, and were followed up weekly. The primary end point of the study was an adequate control of the disease, defined as absence or marked reduction of pruritus and urticaria without the need for systemic steroid therapy. One patient was excluded because of intolerance of both montelukast and zafirlukast (severe headache). Six of the remaining 11 patients (55%) responded to zafirlukast (n=3) or montelukast (n=3) (Table 1). In these subjects, leukotriene receptor antagonists (LRA) induced a nearly total remission of the disease: four could easily control their urticaria with cetirizine 10 mg every other day, and two could discontinue antihistamine treatment altogether. The effect of LRA was already appreciable after some days of treatment. Responders have now been followed up for 6–24 months; they are still taking LRA and are all still in remission. No clinical feature or specific ‘‘marker’’ (sex, age, duration of the disease, NSAID intolerance, autologous serum skin test [ASST] result, thyroid autoimmunity, or histamine-releasing activity of serum) was associated with the response to LRA (Table 1). Leukotrienes might well play a central pathogenic role in some patients with CIU. Their injection into human skin produces a wheal and flare reaction (5), and, on a molar basis, they are 100 times more potent than histamine; moreover, serum from some patients with CIU, particularly those positive on ASST, induces both histamine release and de novo production of sulfidoleukotrienes in human basophils (6). Interestingly, in one study (3), one nonresponder to LRA showed a complete response to the lipoxygenase inhibitor, zileuton, suggesting a possible pathogenic role for LTB4, a substance that enhances cutaneous microvascular permeability by a not yet identified neutrophil-derived mediator (7). Because zileuton is not available in Italy, we could not confirm these observations. Good result in 50% of cases of therapy resistance.

Background: Patients with chronic urticaria (CU) frequently show signs of thrombin generation as a result of the activation of the extrinsic pathway of coagulation and signs of fibrinolysis as shown by slightly increased mean D-dimer plasma levels. Here, we studied patients with severe CU to see whether the activation of coagulation and fibrinolysis parallels the severity of the disease. Methods: Eight consecutive patients with severe exacerbations of CU and 13 with slight CU were studied. Plasma prothrombin fragment F 1 + 2 as well as D-dimer were measured by ELISA. Serum histamine-releasing activity was assessed by basophil histamine release assay. Seventy-four normal subjects were used as controls. Results: In patients with severe CU, median levels of both D-dimer (11.20 nmol/ 1) and F 1 + 2 (592 pmol/1) largely exceeded those found in patients with slight CU [D-dimer: 2.66 nmol/1 (P = 0.001) and F 1 +2 : 228 pmol/1 (P = 0.003)] and in normal subjects [D-dimer: 1.41 nmol/l (P = 0.0001) and F 1 + 2 : 159 pmol/1 (P = 0.0001)]. Sera from 25% of patients with severe CU and 31% of those with slight CU, but from none of normal subjects, showed in vitro histamine-releasing activity. D-dimer and F 1 +2 levels were significantly correlated each other (r = 0.64, P = 0.002) and with CU severity score (r = 0.80-0.90, P = 0.0001), but no correlation was observed between serum histamine-releasing activity and coagulation parameters or severity score. Conclusions: Severe exacerbations of CU are associated with a strong activation of coagulation cascade and fibrinolysis. Whether this activation is the cause of CU or acts as an amplification system is still a matter of debate.