To the Editor,The proportion of the population with allergic diseases has increased rapidly in recent decades1, 2. In addition to affecting the quality of life, a significant economic burden of these diseases was transferred to society and the national health care system1. China is a large country with a rapidly developing economy, wide geography, and diverse climate and lifestyles, which may lead to significantly regional differences in the distribution of allergens. Although a series of studies have explored the prevalence of allergen sensitization in China, the majority of them focus on one part of geography in China3-5. In 2009, a study6 was conducted to estimate the prevalence of common aeroallergens among patients with allergic asthma and/or rhinitis in mainland China. Although the study investigated the differences of the prevalence in different regions of China, it divided China into only four geographical regions, which may neglect detailed information about the characteristics of sensitization prevalence in different places in China. In that study, the skin prick test (SPT) was used to detect the sensitization to allergens. The method has low accuracy for positive results because it is heavily affected by certain factors, such as the skill of the tester, reagent used, interpretation of results and so on. Our research has the following different characteristics compared with previous studies: 1) covering a variety of allergic diseases, 2) exploring both aeroallergens and food allergens simultaneously, 3) including a large set of data from all the seven regions of mainland China, and 4) using an internationally recognized method of sIgE testing, ImmunoCAP, to detect sensitization. These advantages may help us obtain more accurate and reliable results and conclusions.Here, we conducted a large multicenter study on the prevalence patterns of serum allergen-specific IgE (sIgE) sensitization to the four most common food allergens (i.e., egg whites, cow’s milk, crab, and shrimp) and five aeroallergens (i.e., house dust mite, German cockroach, tree pollen mix, mold mix, dog dander) among 44156 patients with allergic symptoms in 52 cities from 26 provinces of all the seven geographical regions in mainland China from July 2015 to June 2018. The sIgE sensitization was tested by a certified third-party laboratory service provider with uniform and standardized procedures. This study was approved by the ethics committee of the First Affiliated Hospital of Guangzhou Medical University (Approval number: GYFYY-2017-18). Details about the methods were in the supplementary materials .Our study showed that the overall prevalence of positive sIgE responses to the 9 allergens across mainland China from the highest to the lowest was 33.74% for house dust mites, 24.5% for cockroaches, 19.97% for shrimp, 17.31% for crab, 11.62% for cow’s milk, 10.92% for egg whites, 9.35% for tree pollen mix, 4.02% for dog dander and 3.92% for mold mix (Table 1 ). Our study confirmed that an observation shown in several previous studies based on certain specific areas in China3-5 that the positive cases in sIgE fell mainly in the two low classes (i.e., classes 1 and 2) was also held in all the seven regions in mainland China (Table 1 ).Our study revealed the distinctive patterns in the prevalence of allergen sensitization among regions, gender, age groups and seasons. Geographically, there is a significant difference in the prevalence among regions for all 9 allergens except for the mold mix (Table S1 ). House dust mites were the allergen with the highest prevalence of sensitization in all seven regions, with the highest in South China (40.79%) and the lowest in Northeast China (11.21%). Allergies to German cockroaches had a higher prevalence in southern regions (Southwest China, South China and East China) than in northern regions (North China and Northeast China). The prevalence of sIgE responses to dog dander was the highest in North China and was very close to each other in the southern regions. The prevalence of the egg whites and milk in Central China, East China and South China was higher than in Southwest China, North China and Northeast China, which means that patients living in eastern, coastal and/or southern areas were more sensitive to egg whites and cow’s milk. The prevalence of crab and shrimp sensitization in Southwest China and South China was higher than that in the northern regions (North China and Northeast China). The heatmap (Figure 1 ) displays the distribution of the prevalence of the sIgE response to allergens in different regions of mainland China.The prevalence of sensitization to all nine allergens was higher overall in males than in females (Table 1 and Figure S1 ) although that may not be true in each age group for each allergen as shown in the forest plot in Figure S1 . Our study showed that, whereas the sensitization to egg whites and milk was the highest in children, the sensitization to other allergens tended to be the highest in teenagers and young adults (Figure S2 ). Figure S3displays he prevalence pattern of allergens by months across years. The prevalence of dog dander and mold mix was very stable across months; however, the prevalence of other allergens fluctuated from January to December. The prevalence of house dust mites, German cockroach, shrimp and crab were higher in the summer months (from June to August) than in other months. The prevalence of tree pollen mix was much higher in April and October than in other months.This should be the first large study to investigate the prevalence of allergen sensitization in the patients with allergic symptoms from all the seven geographic regions of mainland China. Based on this study, we found that the prevalence of sIgE sensitization to allergens displayed obvious and distinctive patterns among regions, gender, age groups and seasons. The reasons for these patterns may include lifestyle factors, socioeconomic factors, genetic predispositions, climate, sexual hormones, cross-reactivity and so on3,4,6-9. Please refer to the supplementary materials for the detailed discussion on the factors that influenced these variations. Our findings may help clinicians find effective individualized treatments for unique patient groups and direct researchers to conduct further studies on the epidemiology of allergic diseases.
To the Editor: Bronchial asthma is characterized by restricted airflow due to chronic airway inflammation, and frequent lower respiratory viral infections in early life are a significant risk factor for development of the disease. Previous studies demonstrated that anti-viral interferon (IFN) production, including of IFN-α, IFN-β and IFN-λ, by leukocytes and bronchial epithelial cells can be impaired in asthma patients.1 An epidemiological study found that allergic sensitization precedes wheeze during asthma development in children, suggesting that Type 2 (T2) conditions play a key role in the impaired anti-viral IFN production. Furthermore, a prospective cohort study showed that, regardless of the type of virus, each successive lower respiratory viral infection with wheeze increases the risk of asthma by about 1.5 fold.2 However, we still don’t have a full understanding of the precise mechanism(s) of how respiratory viral infections under T2 conditions lead to development of asthma.A meta-analysis of large-scale genome-wide association studies revealed that both IL-33 and its receptor, IL-33 receptor(IL-33R ; also known as ST2 ), are closely associated with asthma development.3 Indeed, IL-33 expression was reportedly increased in rhinovirus-infected bronchial epithelial cells and correlated significantly with the disease severity of asthma.4 This suggests that virus-induced IL-33 in the airway may be fundamentally involved in the mechanistic links between viral infection and development and/or exacerbation of asthma. In addition, impairment of anti-viral IFN production was reported to cause necrosis—but not apoptosis—of the virus-infected epithelium,5 which results in release of bioactive IL-33.MicroRNAs (miRNAs) are small, non-coding RNA molecules (containing about 22 nucleotides) that are found in diverse organisms. miRNAs regulate expression of a broad spectrum of target genes through RNA silencing and/or post-transcriptional regulation. Among them, microRNA-29a (miR-29a) was induced by respiratory syncytial virus (RSV) infection in a human lung adenocarcinoma cell line, A549, and suppressed expression of IFN (α, β and ω) receptor 1 (IFNAR1).6 Furthermore, miR-29a regulated the expression of soluble ST2 (sST2), a decoy receptor for IL-33, in human tenocytes.7 Based on those earlier findings, we focused on miR-29 in the present study. We hypothesized that T2 cytokine induces miR-29 expression in bronchial epithelial cells, leading to suppression of both sST2 release and IFNAR1 expression by epithelial cells, and culminating in asthma development and/or exacerbation.Based on that hypothesis, we first examined whether T2 cytokine and inflammatory cytokine induced sST2 production in a human bronchial epithelial cell line, BEAS-2B. The detailed methods are described in Supporting Information. Specific ELISA showed that IL-4 and TNF-α synergistically induced sST2 release from BEAS-2B, in a dose-dependent manner (Figure 1A). Next, to examine the effects of miR-29 overexpression or inhibition on that cytokine-induced sST2 release, BEAS-2B cells were first transfected with miR-29 mimics or inhibitors for 24 hours and then stimulated with a combination of IL-4 and TNF-α for 48 hours.The human miR-29 family consists of three mature members, i.e., miR-29a, miR-29b, and miR-29c. These miR-29s are encoded by the miR-29a/b-1 cluster on chromosome 7q32.3 and the miR-29c/b-2 cluster on chromosome 1q32.2, respectively (Figure 1B).8 The three family members share an identical seed sequence (Figure 1B), and their functional properties are thought to be similar. We examined the effects of miR-29a and miR-29b in this study. ELISA of the culture supernatants showed that inhibition of miR-29a or miR-29b significantly enhanced cytokine-induced sST2 release (Figure 1C). In contrast, overexpression of miR-29a or miR-29b almost completely inhibited that release, indicating that these miR-29s regulate sST2 release from bronchial epithelial cells under T2 conditions. Of note, neither inhibition nor overexpression of miR-29a or miR-29b had any effects on the protein levels of the ST2 receptor in the BEAS-2B cells (Figure 1D, upper panel). These results suggest that T2 cytokine-induced miR-29 plays a critical role in IL-33-dependent allergic inflammation through regulation of sST2 release from bronchial epithelial cells.Furthermore, transfection of either miR-29a or miR-29b inhibitors significantly enhanced IFNAR1 protein expression in the BEAS-2B cells (Figure 1D, middle panel), which is consistent with earlier findings for miR-29a in A549 cells.6 Conversely, transfection of miR-29 mimics resulted in reduced IFNAR1 expression in BEAS-2B cells, suggesting that overproduction of miR-29s in bronchial epithelial cells may lead to suppression of antiviral responses by IFNs. Thus, we found that miR-29s simultaneously regulate the expression of both sST2 and IFNAR1 in bronchial epithelial cells. Our findings suggest the possibility that T2 cytokine-induced miR-29s in airway epithelial cells are key players in the development and/or exacerbation of asthma triggered by respiratory viral infections through both decreasing IFN-regulated antiviral activities and exacerbating IL-33-dependent allergic inflammation.miRNAs are released from cells into the extracellular environment via exosomes, which can then fuse with target cells. This process can deliver various proteins and nucleic acids, including miRNAs, into even distant target/receiving cells.9 We, therefore, examined whether exosomes similarly export miR-29s from bronchial epithelial cells. BEAS-2B cells were stimulated with a combination of IL-4 and TNF-α for 48 hours, and exosomal fractions were collected from the culture supernatants. Although qPCR detected both miR-29a and miR-29b in the exosomes even without that T2 cytokine stimulation (control), both of their copy numbers were significantly increased by that stimulation (Figure 2A). Furthermore, Western blot analysis also found that expression of CD81, an exosome marker, was enhanced by the cytokine stimulation (Figure 2B). These results suggest that T2 cytokine-stimulated epithelial cells release more exosomes containing more miR-29s than unstimulated cells.This study has several limitations. First, no functional experiments were performed in this study to confirm the effects of the changes in sST2 release or IFNAR1 expression. In addition, we did not measure the expression levels of miR-29s in clinical samples.Figure S1 summarizes our findings as a schematic illustration of bronchial epithelial cells. Based on those findings, we hypothesize that elevated nasal, bronchial and/or exosomal levels of miR-29s in infancy may be useful biomarker(s) for predicting later development of asthma, and further studies are needed. Our data suggest a new perspective that miRNAs are crucially involved in the association between viral infection and asthma development. We believe that our research has great significance in pointing to a novel direction for further studies and the existence of a new key player, i.e., miRNAs, in the relationship between viral infections and asthma development.
To the Editor, Immediate hypersensitivity reactions are related to mast cell and/or basophil activation. The mediators released such as tryptase and histamine are involved in clinical symptoms and are key parameters that contribute to diagnosis. Serum tryptase concentrations peak between 30 minutes and 4 hours following the reaction. Tryptase release is considered a robust marker of mast cell degranulation but is not informative in mild reactions.1 Histamine is released at the early beginning of the reaction but has a short half-life. The concentration value can be altered by pre-analytic conditions.2 Anaphylactic reactions can occur during night and week-end when laboratories can’t take charge of samples. To our knowledge, no thorough study of the stability in whole blood of these markers in anaphylaxis has been published. The aim of our study was to evaluate the impact of whole blood sample storage conditions (temperature and delay before centrifugation and plasma collection) on the reliability of tryptase and histamine measurements.Blood samples from 14 patients suspected of anaphylactic reactions (grade 2 to 4 of the Ring and Messmer scale)3 and from 10 volunteers were collected on EDTA after signed informed consent (CCPPRB Caen Basse-Normandie protocol 2004-32). The description of the patient anaphylactic episodes appears in Table 1.When received in the lab, an aliquot of whole blood was processed for diagnostic (reference measurement) and the remaining was divided in aliquots stored at room temperature (RT) or at +4°C for 24, 72 hours or 7 days (patients) or 2, 6, 24 or 72 hours for controls before centrifugation and plasma collection.Total tryptase concentrations were measured by an automated fluoroimmunoassay (ThermoFisher, Phadia SAS, ). Increased tryptase is defined as ≥ 1.2 x basal value + 2 µg.L-1.4 In our hands, tryptase uncertainties of measurement for low and high concentrations (9 µg.L-1 and 38.2 µg.L-1) are 17% and 16% respectively, in accordance with published results.5 Plasma histamine concentrations were measured by a radioimmunoassay (Beckman Coulter, Immunotech, France). Increased values defined by the manufacturer are >10 nmol.L-1, in accordance with published data.6 In our hands, histamine uncertainties of measurement for low and moderate concentrations (4.7 nmol.L-1 and 12.9 nmol.L-1) were 22% and 25%, respectively.The differences between the concentrations measured before and after storage were compared by paired two-tailed t-tests using SAS software. Results were considered significant for p < 0.05.As shown in Figure 1A, storage conditions did not modify tryptase concentrations (linear regression: slope=1.079, R²=0.9675). Tryptase concentrations appeared stable in whole blood left at +4°C for 7 days or 72 hours at RT.Histamine concentrations in patient samples were not modified during 72h at +4°C (Figure 1B) or at RT (Figure 1C). In the control group at RT, histamine concentrations were significantly increased at 6 hours (p=0.005) although moderately increased and staying within the limits of uncertainty measurement and never reaching the positivity threshold (Figure 1E). After 24 hours at RT a false positivity was observed for 8 of 10 samples (p<0.0001) (Figure 1E). At +4°C, histamine concentrations were significantly increased after 24 hours (p<0.0001) but remained in the limits of uncertainty measurement and under the threshold of positivity (Figure 1D). After 72h at +4°C, histamine concentrations exceeded the limits of uncertainty measurement and the positivity threshold for 4 samples of 10 (Figure 1D).Tryptase and histamine measurements are recommended to prove degranulation in anaphylaxis.7 Anaphylactic reactions occur unexpectedly. It is thus important to master sample shipment and processing before mediator measurement.The knowledge of possible artifacts modifying the measured values is necessary for the biochemist to address accreditation criteria of pre-analytic requirements (ISO 15189 standard) and for the physician to rely on trustable diagnostic data.Tryptase stability in whole blood had not been described. Our data has shown no impact on results after 72h at RT or 7 days at +4°C. Tryptase stability in plasma or serum has been evaluated by the manufacturer who ensured stability for 48h at RT or 5 days at +2°C to +8°C (Thermofisher).8 Thus, measured values of tryptase appear highly reliable.Histamine stability in whole blood had only been evaluated in controls and false positive results may be attributed to passive release from basophil during prolonged storage.2 We observed no impact for patient blood samples after 72h at +4°C or at RT. In contrast, false positive results were observed in controls after storage at RT during 24h or at +4°C during 72h. Histamine is known to be stable in the plasma obtained after centrifugation up to 4 days at RT for patients and controls.9According to these results, we suggest that whole blood samples can be stored at +4°C up to 72h for histamine and 7 days for tryptase when the laboratory is not available immediately. In any case, the biochemist must accept all these unrenewable samples. It is his role to take into account the pre-analytical conditions to interpret the results and provide helpful information to the physician.Keywords : pre-analytic; tryptase; histamine; whole blood.
Background The prevalence of allergy to cat is expanding worldwide. Allergen-specific immunotherapy (AIT) has advantages over symptomatic pharmacotherapy and promises long lasting disease control in allergic patients. However, there is still a need to improve cat AIT regarding efficacy, safety and adherence to the treatment. Here we aim to boost immune tolerance to the major cat allergen Fel d 1 by increasing the anti-inflammatory activity of AIT with the established immunomodulatory adjuvant CpG, but at a higher dose than previously used in AIT. Methods Together with CpG, we used endotoxin-free Fel d 1 as therapeutic allergen throughout the study in a BALB/c model of allergy to Fel d 1, thus mimicking the conditions of human AIT trials. Multidimensional immune phenotyping including mass cytometry was applied to analyze AIT-specific immune signatures. Results We show that AIT with high-dose CpG in combination with endotoxin-free Fel d 1 reverts all major hallmarks of allergy. High dimensional CyTOF analysis of the immune cell signatures initiating and sustaining the AIT effect indicates the simultaneous engagement of both, the pDC-Treg and -B cell axis, with the emergence of a systemic GATA3+ FoxP3hi biTreg population. The regulatory immune signature also suggests the involvement of the anti-inflammatory TNF/TNFR2 signaling cascade in NK and B cells at an early stage and in Tregs later during AIT. Conclusion Our results highlight the potential of CpG adjuvant in a novel formulation to be further exploited for inducing allergen-specific tolerance in patients with cat allergy or other allergic diseases in the future.
To the Editor.The beneficial effects of Allergen Specific Immunotherapy (AIT) relies on the induction of allergen-specific Regulatory T-cells (Tregs) (1). Tregs, a subpopulation of CD4+CD25+T-cells expressing the specific transcription factor Foxp3, are not functionally homogeneous and their detection is complex and uncertain due to FoxP3 intracellular localization. Furthermore, FoxP3+ Tregs might become unstable and halt the production of their functional suppressive cytokines in inflammatory conditions (2) (1). In its place, the surface antigen CD127, whose expression inversely correlates with FoxP3, conveniently identifies Tregs as CD4+CD25+CD127negcells (3) (2), so surmounting the problems of FoxP3 stability and intracellular detection. Tregs also constitutively express the inhibitory antigen CD39, enhanced in highly suppressive memory Tregs (4) (3). Furthermore, HLA-DR expression is a monitor of Treg differentiation status and identifies a functionally and greatly suppressive population (1,5) (1,5). Lack of CD45RA characterizes memory T cells enabled to survive for long periods, even in absence of specific antigen, showing increased activity upon re-exposure and able to induce apoptosis in target cells (6) (3). CD4+CD25highCD39+CD127negcells are subtyped as Resting (CD45RA+/HLA-DRneg: rTreg), Activated (CD45RAnegHLA-DRneg: aTreg) and Effector (CD45RAnegHLA-DRlow/high: eTreg) Tregs (6) (6). This latter subtype includes terminally differentiated Tregs, the most highly suppressive (5) (7) (Supplementary Figure 1). They are different from secreting or type III Tregs expressing CD127 that represent a short-lived terminally differentiated population (5,6,8) . In order verify possible correlations between specific subsets of Treg and the effectiveness of AIT, we applied this analytical approach to study Treg profile in adolescents suffering from mite allergic rhinitis, pre and 12 months post Sublingual Immunotherapy (SLIT) with mite monomeric allergoid, an acid-resistant allergen known to elicit early T reg-activation (7,8). The study was approved by the Ethical Committee of University “G. d’Annunzio”, Chieti-Pescara. All patients and parents signed a written informed consent after having been informed about the procedures of the study.Twenty patients diagnosed with mite-allergic persistent rhinitis with or without asthma were enrolled. Allergic rhinitis (AR) was graded according to ARIA guidelines in 1) intermittent mild, 2) intermittent moderate/severe, 3) persistent mild and 4) persistent moderate/severe. At the enrollment, each patient marked in a 100 mm visual analogic scale (VAS) the level of its health status related to allergy with 0 the best status and 100 the worst.All patients were treated by SLIT with mite monomeric allergoid (LAIS - Lofarma, Milan, Italy) at 1000 UA four times/week every other day, for 12-months. No adverse local and systemic reactions were detected. The effectiveness of SLIT was established comparing VAS, ARIA grading and ACT questionnaire performed after 12-months of treatment with their basal values. Two blood samples were drawn pre/post SLIT to be analyzed for Regulatory T-cells. Clinical and demographic details of the studied population, analytical methods, statistical approach and the outline of the study are detailed in the online supplementary material .Rhinitis scores VAS and ARIA significantly decreased after SLIT (Table 1), with the same statistical significance (Wilcoxon z -3.7236; p = 0.0002). Improvement was evidenced also in the subgroup of asthmatic patients (n=7) since ACT scores significantly increased from the baseline value of 18 (16-19) up to 24 (20-25) after 12 months of treatment (the low number of patients does not allow application of efficient statistics).Tregs were analyzed as frequency of total Treg cells and their three subsets, namely Resting (rTregs), Activated (aTregs) and Effector (eTregs), within the parental population of CD4+cells. Total Tregs did not change significantly; rTreg significantly decreased (Wilcoxon z-3.6214, p<0.0003), while, the abundance of aTregs and eTregs significantly incremented (Wilcoxon z-2.9011, p<0.05 and z-3.077, p=0.002, respectively) (Table 1). A significant negative correlation has been observed between the decrease in rTreg and the increase in aTreg (Spearman’s ρ-0.69391, p<0.02) and increase in eTreg cells (Spearman’s ρ-0.56845, p<0.02) (Figure 3 in supplementary material).HLA-DR resulted significantly up-regulated in all Tregs from 4.93±3.1 to 6.92±5.1 MFI (Wilcoxon z-4.2026, p <0.00001). HLA-DR increased on aTregs from 3.4±3.03 to 4.91±3.2 MFI (Wilcoxon z-3.2479, p=0.001) and on eTregs from 1.54±0.66 to 2.0±1.45 MFI (Wilcoxon z-2.9664, p=0.005). CD39 was found differently expressed in the three subsets of Tregs at baseline, with Resting<Activated<Effector. After 12 months of SLIT, CD39 surface expression was found significantly increased in all Tregs from 6.9±4 to 8.02±5 MFI (Wilcoxon z-3.1049, p=0.001) (HLA-DR and CD39 changes are reported in Table 1). We found some interesting correlations between laboratory data and clinical parameters. Changes in eTregs significantly correlated with both ARIA (Spearman’s ρ=0.58728, p=0.013) (Figure 1A) and VAS (Spearman’s ρ=0.49172, p=0.044) (Figure 1B) variations after SLIT. While a significant negative correlation was found between rTregs and clinical parameter changes after treatment (Spearman’s ρ-0.48482, p=0.0491). Changes in HLA-DR expression on all Treg cells significantly correlated with variation in VAS pre-/post-SLIT (Spearman’s ρ=0.54104, p= 0.01376) (Figure 1C). No other correlations were found except for the lowest increase (< 8%) of memory Tregs (CD45RAneg) detected in patients with the lowest levels of mite-specific serum IgE (not shown).To our knowledge this is the first report on successful SLIT being associated with re-patterning of the differentiation status of Tregs, with high rates of the most suppressive Treg subtypes: activated and effector, characterized by higher expression of HLA-DR and CD39 both playing inhibitory function in Tregs. Moreover, effective SLIT seems to be associated with the generation of cells lacking CD45RA that characterizes memory T cells with increased activity upon re-exposure to the antigen. Our results suggest that SLIT also induced empowerment of Treg inhibitory function, likely compensating the under-representation of Tregs observed in allergic patients (9) (9). In AR children, there are evidences that Tregs have defect in suppressing IgE production and that they can be incremented by mite SLIT.Next step of our study will be to evidence if such relationship between effective SLIT and Treg re-patterning is present in the first months of SLIT, with a view to profiling Tregs for the early identification of SLIT responders/non-responders by mean of a straightforward and non-invasive blood test.
Immediate and non-immediate hypersensitivity reactions to iodinated contrast media (ICM) have been reported to occur in a frequency of about 0.5-3% of patients receiving non-ionic ICM. The diagnosis and management of these patients is controversial among guidelines published by various national and international scientific societies, with recommendations ranging from avoidance or premedication to drug provocation test. This position paper aims to give recommendations for the management of patients with ICM hypersensitivity reactions and analyze controversies in this area. Skin tests are recommended as the initial step for diagnosing patients with immediate and non-immediate hypersensitivity reactions; besides, they may also help guide on tolerability of alternatives. Drug provocation test is the gold-standard; although, as it is a risky procedure, the decision for performing it needs to be taken based on a risk-benefit analysis. Another source of controversy is the role of in vitro tests for diagnosis and pretreatment for preventing reactions.
Modern healthcare requires a proactive and individualized response to diseases, combining precision diagnosis and personalized treatment. Accordingly, the approach to patients with allergic diseases encompasses novel developments in the area of personalized medicine, disease phenotyping and endotyping and the development and application of reliable biomarkers. A detailed clinical history and physical examination followed by the detection of IgE immunoreactivity against specific allergens still represents the state of the art. However, nowadays, further emphasis focuses on the optimization of diagnostic and therapeutic standards and a large number of studies have been investigating the biomarkers of allergic diseases, including asthma, atopic dermatitis, allergic rhinitis, food allergy, urticaria and anaphylaxis. Various biomarkers have been developed by omics technologies, some of which lead to a better classification of the distinct phenotypes or endotypes. The introduction of biologicals to clinical practice increases the need for biomarkers for patient selection, prediction of outcomes and monitoring, to allow for an adequate choice of the duration of these costly and long-lasting therapies. Escalating healthcare costs together with questions on the efficacy of the current management of allergic diseases requires further development of a biomarker-driven approach. Here, we review biomarkers in diagnosis and treatment of asthma, atopic dermatitis, allergic rhinitis, viral infections, chronic rhinosinusitis, food allergy, drug hypersensitivity and allergen-immunotherapy with a special emphasis on specific IgE, microbiome and epithelial barrier. In addition, EAACI guidelines on biologicals are discussed within the perspective of biomarkers.
Background: There is limited information on risk factors for eczema in adults. Recent evidence suggests that air pollution may be associated with increased incidence of eczema in adults. We aimed to assess this possible association. Methods: Ambient air pollution exposures (distance from a major road, nitrogen dioxide [NO2], fine particulate matter with an aerodynamic diameter ≤2.5 µm [PM2.5]) were assessed for the residential address of Tasmanian Longitudinal Health Study participants at ages 43 and 53 years. Eczema incidence (onset after age 43 years), prevalence (at 53 years) and persistence were assessed from surveys, while sensitisation was assessed using skin prick tests. The presence or absence of eczema and sensitisation was classified into four groups: no atopy or eczema, atopy alone, non-atopic eczema, and atopic eczema. Adjusted logistic and multinomial regression models were fitted to estimate associations between ambient air pollution and eczema, and interaction by sex was assessed. Results: Of 3153 participants in both follow ups, 2369 had valid skin prick tests. For males, a 2.3 ppb increase in baseline NO2 was associated with increased risk of prevalent eczema (OR=1.15 [95%CI 0.98-1.36]), both non-atopic (OR=1.39 [1.02-1.90]) and atopic eczema (OR=1.26 [1.00-1.59]). These associations were not seen in females (P for interaction=0.08, <0.01). For both sexes, a 1.6 µg/m3 increase in PM2.5 exposure at follow-up was associated with increased odds of aeroallergen sensitisation (OR=1.15 [1.03-1.30]). Conclusion: Increased exposure to residential ambient air pollutants was associated with an increased risk of eczema, only in males, and aeroallergen sensitisation in both genders.
EDITORIAL The average global temperatures on our planet are increasing due to rising anthropogenic greenhouse gases in the atmosphere, in particular carbon dioxide (CO2).1,2 There is an urgent need to call for action on global warming, which is resulting in extreme weather and related catastrophes.1 ,2 The Earth’s rising temperature is evidenced by warming of the oceans, melting glaciers, rising sea levels, and the diminished snow cover in the Northern Hemisphere. Climate-related factors can affect interactive atmospheric components (chemical and biological) and their interrelationship with human health.Climate change, a physics and meteorological event that affects health in the whole biosphere started to receive attention around the mid-twentieth century. Air pollution is the driving force of the Earth’s warming powered by the greenhouse effect (Figure 1). Environmental changes are occurring in frequency, intensity, type of precipitation, and extreme weather events, such as heatwaves, droughts, floods, blizzards, thunderstorms, sandstorms, and hurricanes. These are real and daunting challenges for the human and biosphere health, impacting the food and water supplies.1 ,2 Urbanization, with its high level of vehicle emissions and westernized lifestyle, is linked to the rising levels of particulate matter in the air, food supplies, soil, freshwater, and oceans. These environmental changes are correlated with the increased frequency of respiratory allergic diseases and bronchial asthma observed over recent decades in most industrialized countries and is continuously rising in developing countries.1-5This issue of Allergy focuses on the interrelationship between climate change, air pollution and human health.3-7Climate change is an important medical aspect in allergology as we are observing an increasing incidence of allergic diseases indirectly related to rising temperatures and are becoming a high socio-economic burden.1-3,8 Allergies and asthma appear to be at the front line of the sequelae of climate change along with infectious and cardiovascular diseases.1,5Cecchi et al. focus on the development and exacerbation of allergic diseases can be explained in terms of the exposome, a concept that includes all the environmental exposures from conception onwards. Multiple factors can trigger a pollen-induced respiratory allergy, such as airborne endotoxin levels and microbial composition of pollen, and these comprise a “pollen exposome”.4,9Susan Prescott has written an editorial in this issue bringing the attention to climate change and bidiversity aspects. At the time of Neil Armstrong’s lunar landing 50 years ago, Prof. Rene Dubos, a renowned microbiologist, delivered the seminal lecture “The Spaceship Earth”. He was ahead of his time and warned of an “altered immunity” driven by environmental problems and loss of biodiversity. Most of his predictions proved correct and we are now understanding at a molecular level the pathophysiological mechanisms involved in allergic diseases.8Climate change indirectly affects allergies by altering the pollen concentrations, allergenic potential, composition, migration of species and growth of new ones. Air pollution and climate change have resulted in the faster growth of allergenic plants, increasing the aeroallergen load for patients with inhalant allergy. Phenological studies indicate longer pollen seasons and emerge earlier in the year.1,4,5,8 Pollen and mold allergies are generally used to evaluate the interrelationship between air pollution and allergic respiratory diseases, such as rhinitis and asthma. Studies show that plants exhibit enhanced photosynthesis and reproductive effects and produce more pollen as a response to high atmospheric levels of CO2. 1,4,8 Pollen allergens have been demonstrated to trigger the release of pro-inflammatory and immunomodulatory mediators that accelerate the onset of allergy and the IgE-mediated sensitization. Lightning storms or wet conditions rupture the pollen grains releasing the allergenic proteins that cause asthma exacerbations in patients with pollinosis (thunderstorm-asthma).1,3,4,7,10 As a result of climate change, patients with seasonal allergic rhinoconjunctivitis and asthma have more intense symptoms and need stronger medication.1,4,8 In addition to respiratory illnesses, Fairweather et al. demonstrate the effect of environmental changes on cardiovascular, brain and mind, gastrointestinal, skin, immunologic and metabolic effects.1,3,4,7 The migration of stinging and biting insects to cooler climates has caused an increase in insect allergies in those areas.Prunicki et al. focus on the contribution of wildfires and deforestation and their contribution to global warming and immunological effects. It should be noted that in the last fifty years, half of the pluvial forests on Earth have been lost. Deforestation and forestation degradation is estimated to occur at a rate of 13 million hectares per year, mostly for agricultural purposes. Wildfires are becoming increasingly frequent, posing a serious risk to human health. The fine particulate matter (PM2.5) in wildfire smoke exacerbates asthma attacks, among other health problems. A study of 67 subjects demonstrated that those exposed to wildfire smoke had significantly higher levels of C-reactive protein and IL-1β compared with controls.6 The elevated levels of these two biomarkers are indicative of airway inflammation.Global warming and climate change need actions throughout the whole world with joined forces of all capabilities. These efforts are sometimes hampered by the unresponsiveness of governmental institutions and the general population, the lack of infrastructure and poverty. An action plan is needed to disseminate information on health-related problems associated with climate change. Patients with pollen allergies or asthma should be educated on the higher health risk during a thunderstorm or pollen season and the need for appropriate medication if staying outdoors. In collaboration with environmental organizations, physicians should take the lead to promote actions to mitigate air pollution and advocate the need to reduce global warming to protect our health.
EDITORIAL Coronavirus disease‐19 (COVID‐19) is a new disease caused by SARS‐CoV2. Since the beginning of 2020, it has become one of the main challenges of our times, causing a high incidence of severe pneumonia, acute respiratory distress syndrome (ARDS), multiorgan failure and death1. At the root of COVID-19 lies the sudden development of ‘cytokine storms’, hyper-inflammatory responses involving the release of pro-inflammatory cytokines (e.g., TNF-α, IL-6, IL-1, IL-8, and MCP-1) that impair the gas exchange function of the lung and lead in select patients, mostly with underlying comorbidities, to multiorgan failure and death1,2. Additional complications triggered by ‘cytokine storms’ include endothelial dysfunction and hypercoagulation, increasing the risk of thromboembolytic events, and life-threatening cardiovascular complications. Anti-inflammatory therapies are thus being considered for alleviating the damaging side effects of hyper-inflammation with many trials including anti-cytokine biologicals, disease-modifying antirheumatic drugs (DMARDs) and corticosteroids being ongoing3. Surprisingly, among them dexamethasone has taken center stage as initial results from the RECOVERY trial, a large multicenter randomized open-label trial for hospitalized patients run in the United Kingdom, revealed notable efficacy in the treatment of critically ill COVID-19 patients4.Dexamethasone is one of the oldest synthetic glucocorticoid agonists synthesized in 1957 and introduced into the clinic in 1961. When administered at 6 mg daily, either orally or intravenously for 10 days, dexamethasone was shown in the RECOVERY trial to improve survival rates of hospitalized patients with severe COVID-19 receiving oxygen or being on mechanical ventilation by a remarkable 30%4. Benefit was restricted to patients requiring respiratory support whereas in milder cases this was not clear. This notable efficacy of dexamethasone treatment goes against the current view of corticosteroid use in respiratory viral infections which remains contradictory. Although corticosteroids improve ventilator weaning and can lower the intensity of the host response to the virus, tempering the ‘cytokine storm’ and limiting immunopathology, they can also reduce viral clearance and lead to more severe disease. Understanding therefore how dexamethasome mediates its effects is of paramount importance.Dexamethasone, as other corticosteroids, is held to mediate its anti-inflammatory and immunosuppressive effects via the glucocorticoid receptor. Upon ligand binding, the receptor-corticosteroid molecule complex moves into the cell nucleus, where it dimerizes and binds to glucocorticoid response elements (GRE), acting as transcriptional repressor or transactivator of diverse sets of genes. This results in the inhibition of inflammatory cell activity, including neutrophils, macrophages and lymphocytes, and the suppression of pro-inflammatory cytokines such as TNF and interleukins and other genes such as cyclooxygenase-2 and inducible nitric oxide synthase5. However, we have recently uncovered that dexamethasone can also induce the D-series proresolving lipid mediator pathway leading to the production of 17-HDHA and the protectins D1 and DX6. These are potent major players of the molecular machinery driving the resolution of inflammation, i.e. the proper regulated termination of pro-inflammatory responses involving the catabolism of pro-inflammatory mediators, the removal of inflammatory cells and the restoration of the tissue in a timely and highly coordinated manner7. Although resolution of inflammation has long been considered to occur spontaneously as a result of the waning of pro-inflammatory responses, this is now known to be an ordered and highly regulated process involving the timely production of enzymatically oxygenated lipid-derived mediators such as protectins, D-series resolvins and maresins derived from the omega-3 fatty acid docosahexaenoic acid (DHA), E-series resolvins derived from eicosapentaenoic acid (EPA), and lipoxins biosynthesized from omega-6 fatty acids following eicosanoid class switching7. Interestingly, certain lipid mediators have been shown to exert additional non-conventional functions; resolvin D4 can attenuate pathologic thrombosis, reduce NETosis and promote clot removal8 which is now recognized as a key pathology of COVID-19 infection, while resolvin E4 (RvE4) stimulates efferocytosis of senescent erythrocytes in hemorrhagic exudates especially under hypoxic conditions that characterize COVID-199. Moreover, corticosteroids have been reported to reduce fibrinogen and procoagulant factors under pro-inflammatory conditions and increase anticoagulant factors10.The ability of viral infections to induce proresolving lipids has been reported earlier. Toll-like receptor 7 (TLR7), a major pattern recognition receptor of viral RNA, activates PD1 and PDX production11. Moreover, influenza virus infection has been demonstrated to drive proresolving lipid mediator networks including the production of PD1 which limits influenza pathogenicity by directly interacting with the RNA replication machinery to inhibit viral RNA nuclear export12,13. Notably, in particularly virulent strains of influenza virus such as the H5N1 avian strain, PD1 formation is not sufficiently upregulated, leading to more efficient viral replication and host demise12. It is therefore plausible that the efficacy of dexamethasone in COVID-19 is due at least in part to its ability to induce proresolving lipid mediators that possess multiple anti-inflammatory and proresolving actions tempering down inflammation and promoting its resolution, preventing coagulation and enhancing viral and bacterial clearance (Figure 1) yet are not immunosuppressive . Whether other corticosteroids beyond dexamethasone can also mediate such effects, and to what extent, is not known. Whether inhalable corticosteroids, such as those given to asthmatic patients, can also induce proresolving lipid mediator networks locally and thus prevent the development of severe SARS‐CoV‐2 infection remains to be determined. There is evidence that asthmatic patients exhibit reduced incidence of severe and/or critical COVID-1914.Recently, COVID-19 patients showed increased association of serum arachidonate-derived proinflammatory lipid mediators, e.g. prostaglandins, in severe COVID -19 infections while some pro-resolving mediators such as resolvin E3 were up-regulated in the moderate COVID-19 group suggesting that an imbalance in lipid mediators with a swift toward pro-inflammatory mediators in severe disease may contribute to COVID-19 disease severity15. Although the involvement of proresolving lipid mediator pathways in COVID-19 is an attractive hypothesis, further evidence from human trials is needed as there are no studies at present reporting the induction or modulation of such networks in the context of the various disease stages and treatments. It is thus of uttermost priority to investigate proresolving lipid mediators in COVID-19, in a temporal and longitudinal manner, as modulating these networks either through drug treatment or direct administration of resolvin and protectins agonists has the potential to affect this highly lethal and devastating disease in a way other approaches cannot. Such studies are therefore eagerly awaited.
To the Editor, Sulforaphane [1-isothiocyanato-4-(methylsulfinyl)butane] is a clinically relevant nutraceutical compound present in cruciferous vegetables (Brassicaceae). It is used for the prevention and treatment of chronic diseases and may be involved in ageing.1Along with other natural nutrients, sulforaphane has been suggested to have a therapeutic value for the treatment of the coronavirus disease 2019 (COVID-19).2 Sulforaphane is an isothiocyanate stored in its inactive form glucoraphanin.3 The enzyme myrosinase, found in plant tissue and in the gut microbiome, is involved in the conversion of glucoraphanin to its active form sulforaphane.4
Dear Editor: I read with interest the report by Antonella et al.1 This report described a case of the acute scrotum caused by Anisakis . As the authors write, this condition is rare in its own right. However, I would like to discuss two other rare aspects of this case: that it occurred during childhood and that acute scrotal disease and anaphylaxis occurred simultaneously.There has been a long debate as to whether anaphylaxis caused by Anisakis occurs with the ingestion of live insect bodies only or with dead insect bodies as well.2 Since several allergen components of Anisakis have been identified and their tolerance to heat has been reported, it is theoretically possible that anaphylaxis could occur with the ingestion of dead larvae body parts. However, some reports suggest that even patients sensitized to Anisakis may not develop allergic symptoms with the ingestion of frozen Anisakis larvae.3Nevertheless, there have been very few cases of gastrointestinal anisakiasis and anaphylaxis occurring simultaneously. In fact, previous literature has shown that in 40 cases of anaphylaxis which occurred due to the ingestion of live fish, upper gastrointestinal endoscopy revealed no difference in phenotype between the 20 cases in which live larvae were found and the 20 cases in which they were not found, and even in the case of live Anisakis bodies, the abdominal symptoms were minor.4 Of the 128 cases included in our previous study, only one could be said to have developed anaphylaxis and gastric anisakiasis simultaneously.5The patient we experienced was a 36-year-old woman with a previous history of gastric anisakiasis. Urticaria, watery diarrhea and vomiting, and respiratory distress developed three hours after eating sashimi (sliced raw fish) of young yellowtail. The patient was rapidly administered adrenaline intramuscular injection, followed by H1/H2 blockers and methylprednisolone, and admitted to the hospital for observation. However, after a day of admission, she continued to complain of intermittent epigastric pain and underwent upper gastrointestinal endoscopy. A live Anisakis larva was found in the gastric cavity, and the epigastric pain disappeared after its removal. This case was negative for fish-specific IgE and positive forAnisakis -specific IgE (ImmunoCAP🄬 fluorescent enzyme immunoassay). Similar cases have been reported recently by Shikino et al.6The reason for such phenotypic variations after the ingestion of liveAnisakis is a direction for future research. From this perspective, it would be very interesting to explore what pathological changes, e.g., eosinophilic granulomatous changes, had occurred in the scrotum or lungs of the boy described in Antonella et al. I believe that these characteristics are important to determine the cause of the respiratory impairment in this case.Further, it is interesting to note that this phenomenon occurred in an 8-year-old boy. Only one in our 128 cases of fish-associated anaphylaxis was under 10 years of age, and this case was positive for the IgE specific to horse mackerel and mackerel.5 Therefore, the group I analyzed did not include cases of Anisakisanaphylaxis under the age of 10 years. The case described in Antonella’s manuscript does not appear to have undergone a specific IgE test or other skin tests. However, given the rarity of Anisakisanaphylaxis in this age group, anaphylaxis due to other culprits such as parvalbumin caused by fish ingestion should also be considered.Ryo Morishima MDDepartment of Neurology, Tokyo Metropolitan Neurological Hospital,Tokyo, JapanReferenceAntonella C, Stellario C, Aurelio M, Domenico S, Domenico S, Ilaria PP, et al. Acute scrotum in a 8-year-old Italian child caused by extraintestinal anisakiasis in a seaside area. Allergy 2020 [in press]Nieuwenhuizen NE. Anisakis – immunology of a foodborne parasitosis. Parasite Immunology 2016 Sep;38(9):548-57. doi: 10.1111/pim.12349. PMID: 27428817Alonso-Gómez A, Moreno-Accillo A, López-Serrano MC, Suarez-de-Parga JM, Daschner A, Cabañas R, et al. Anisakis simplex only provokes allergic symptoms when the worm parasitizes the gastrointestinal tract. Parasitol Res. 2004 Aug;93(5):378-84. doi: 10.1007/s00436-004-1085-9. PMID: 15221464Daschner A, Alonso-Gómez A, Cabañas R, Suarez-de-Parga JM, López-Serrano MC. J Allergy Clin Immunol. 2000 Jan;105(1 Pt 1):176-81. doi: 10.1016/s0091-6749(00)90194-5. PMID: 10629469Morishima R, Motojima S, Tsuneishi D, Kimura T, Nakashita T, Nishino H, et al. Anisakis is a major cause of anaphylaxis in seaside areas: an epidemiological study in Japan. Allergy. 2020 Feb;75(2):441-444. doi: 10.1111/all.13987. PMID: 31315145Shikino K, Ikusaka M. Anaphylaxis induced by Anisakis . Intern Med 2019 Jul 15;58(14):2121. doi: 10.2169/internalmedicine.2428-18. PMID: 30918192
The basophil activation test (BAT) is a functional assay that measures the degree of degranulation following stimulation with allergen or controls by flow cytometry and is directly correlated with histamine release. From the bell-shaped curve resulting from BAT in allergic patients, basophil reactivity (given by %CD63+ basophils) and basophil sensitivity (given by EC50 or similar) are the main outcomes of the test. BAT takes into account all characteristics of IgE and allergen and thus can be more specific than sensitization tests in the diagnosis of allergic disease. BAT reduces the need for in vivo procedures, such as intradermal tests and allergen challenges, which can cause allergic reactions of unpredictable severity. As it closely reflects the patients’ phenotype, it can potentially be used to monitor the natural resolution of food allergies and to predict and monitor clinical response to immunomodulatory treatments, such as allergen-specific immunotherapy and biologicals. Clinical application of BAT requires analytical validation, clinical validation, standardization of procedures and quality assurance to ensure reproducibility and reliability of results. Currently, efforts are ongoing to establish a platform that could be used by laboratories in Europe and in the USA for certification.
Background: The prevalence of tree nut allergy has increased worldwide, and cashew has become one of the most common food allergens. More critically, cashew allergy is frequently associated with severe anaphylaxis. Despite the high medical need, no approved treatment is available and strict avoidance and preparedness for prompt treatment of allergic reactions are considered dual standard of care. In the meantime, Phase III study results suggest investigational epicutaneous immunotherapy (EPIT) may be a relevant and safe treatment for peanut allergy and may improve the quality of life for many peanut allergic children. Objective: We aimed to evaluate the capacity of EPIT to provide protection against cashew-induced anaphylaxis in a relevant mouse model. Methods: A mouse model of IgE-mediated cashew anaphylaxis was first developed. Based upon this model, the efficacy of EPIT was evaluated by applying patches containing cashew allergens to cashew-sensitized mice. Cashew-specific antibody titers were measured throughout treatment. Treated mice were challenged orally to cashew and anaphylactic symptoms were monitored. Additionally, plasma levels of mast-cell proteases (mMCP)-1/7 were quantified from blood samples collected after challenge to evaluate IgE-induced mast-cell activation. Results: EPIT significantly decreased anaphylactic symptoms following challenge and increased cashew-specific IgG2a (equivalent of human IgG1). Interestingly, this protection was associated with a sharp decrease in mast-cell reactivity. Conclusion: We demonstrate that EPIT markedly reduced IgE-mediated allergic reactions in a mouse model of cashew allergy, which suggests that EPIT may be a relevant approach to treating cashew allergy.
This systematic review evaluates the efficacyand safety of omalizumab for chronic spontaneous urticaria (CSU). Pubmed, EMBASE and Cochrane Library were searched for RCTs. Critical and important CSU-related outcomes were considered. The risk of bias and the certainty of the evidence were assessed using GRADE. Ten RCTs including 1620 subjects aged 12 to 75 years old treated with omalizumab for 16 to 40 weeks were evaluated. Omalizumab 150 mg: does not result in clinically meaningful improvement(high certainty) of the urticaria activity score (UAS)7 (mean difference (MD) -5; 95%CI -7.75 to -2.25) and the itch severity score(ISS)7 (MD -2.15; 95% CI -3.2 to -1.1); does not increase (moderate certainty) quality of life (QoL) (Dermatology Life Quality Index (DLQI); MD -2.01; 95%CI -3.22 to -0.81); decreases (moderate certainty) rescue medication use (MD -1.68; 95%CI -2.95 to -0.4). Omalizumab 300 mg:results in clinically meaningful improvements(moderate certainty)of the UAS7 (MD -11.05; 95%CI -12.87 to -9.24), theISS7 (MD -4.45; 95%CI -5.39 to -3.51), and QoL (high certainty)(DLQI; MD -4.03; 95% CI -5.56 to -2.5); decreases (moderate certainty) rescue medication use (MD -2.04; 95%CI -3.19 to -0.88) and drug-related serious AEs (RR 0.77; 95%CI 0.20 to 2.91).
This systematic review evaluates the efficacy, safety and economic impact of dupilumabcompared to standard of care for uncontrolled moderate-to-severe atopic dermatitis (AD). Pubmed, EMBASE and Cochrane Library were searched for RCTs and health economic evaluations. Critical and important AD-related outcomes were considered. The risk of bias and the certainty of the evidence were assessed using GRADE. Seven RCTs including 1845 subjects > 12 years treated with dupilumab16 to 52 weeks were evaluated. For adultsthere is high certainty that dupilumabdecreasesSCORAD (MD -30,72; 95%CI -34,65% to -26,79%) and EASI-75 (RR 3.09; 95%CI 2.45 to 3.89), pruritus (RR 2.96; 95%CI 2.37 to 3.70), rescue medication (RR 3.46; 95%CI 2.79 to 4.30), sleep disturbance (MD -7.29; 95%CI -8.23 to -6.35), anxiety/depression (MD -3.08; 95% CI -4.41 to -1.75) and improves quality of life (MD -4.80; 95% CI -5.55 to -4.06). The efficacy for adolescents is similar. Dupilumab-related adverse events (AEs) slightly increase (low certainty). The evidence for dupilumab-related serious AE is uncertain. The incremental cost-effectiveness ratio ranged from 28,500 £ (low certainty) to 124,541 US$ (moderate certainty).More data on long term safety are needed both for children and adults, together with more efficacy data in the paediatric population.