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Therapeutic antibodies have predominantly been IgG-based. However, the ongoing clinical trial of MOv18 IgE has highlighted the potential of using IgE antibodies in cancer therapy. While extensive studies targeting IgG glycosylation resulted in a rational basis for the development of enhanced biotherapeutics, IgE glycosylation remains an area with limited analyses. Previous studies on the role of IgE glycosylation present conflicting data with one study emphasizing the importance of N275 and T277 residues for FcεRI binding whereas another asserts the nonsignificance of IgE glycosylation in receptor interaction. While existing literature underscores the significance of glycans at the N275 position for binding to FcεR1 receptor and initiation of anaphylaxis, the role of other IgE glycosylation sites in folding or receptor binding remains elusive. This study systematically investigates the functional significance of N-linked glycosylation sites in the heavy chain of IgE which validates the pivotal role of N275 residue in IgE secretion and stability. Replacement of this asparagine to non-amine group moieties does not affect IgE function in vitro, yet substitution with aspartic acid compromises antibody yield. The deglycosylated IgE variant exhibits superior efficacy, challenging the conventional importance of glycosylation for effector function. In summary, our study unveils an intricate relationship between N-glycosylation sites and the structural-functional dynamics of IgE antibodies. Furthermore, it offers novel insights into the IgE scaffold, paving the way for the development of more effective and stable IgE-based therapeutics., (© 2024 Wiley Periodicals LLC.)

By whole exome sequencing, we identified a homozygous c.2086 C→T (p.R696C) TERT mutation in patients who present with a spectrum of variable bone marrow failure (BMF), raccoon eyes, dystrophic nails, rib anomalies, fragility fractures (FFs), high IgE level, extremely short telomere lengths (TLs), and skewed numbers of cytotoxic T cells with B and NK cytopenia. Haploinsufficiency in the other family members resulted in short TL and osteopenia. These patients also had the lowest bone mineral density Z-score compared to other BMF-patients. Danazol/zoledronic acid improved the outcomes of BMF and FFs. This causative TERT variant has been observed in one family afflicted with dyskeratosis congenita (DC), and thus, we also define a second report and new phenotype related to the variant which should be suspected in severe cases of DC with co-existent BMF, FFs, high IgE level and rib anomalies., (© 2023 British Society for Haematology and John Wiley & Sons Ltd.)

Background: Piwi-interacting RNAs (piRNAs), comprising the largest noncoding RNA group, regulate transcriptional processes. Whether piRNAs are associated with type 2 (T2)-high asthma is unknown., Objective: We sought to investigate the association between piRNAs and T2-high asthma in childhood asthma., Methods: We sequenced plasma samples from 462 subjects in the Childhood Asthma Management Program (CAMP) as the discovery cohort and 1165 subjects in the Genetics of Asthma in Costa Rica Study (GACRS) as a replication cohort. Sequencing reads were filtered first, and piRNA reads were annotated and normalized. Linear regression was used for the association analysis of piRNAs and peripheral blood eosinophil count, total serum IgE level, and long-term asthma exacerbation in children with asthma. Mediation analysis was performed to investigate the effect direction. We then ascertained if the circulating piRNAs were present in asthmatic airway epithelial cells in a Gene Expression Omnibus (GEO; www.ncbi.nlm.nih.gov/geo) public data set., Results: Fifteen piRNAs were significantly associated with eosinophil count in CAMP (P ≤ .05), and 3 were successfully replicated in GACRS. Eleven piRNAs were associated with total IgE in CAMP, and one of these was replicated in GACRS. All 22 significant piRNAs were identified in epithelial cells in vitro, and 6 of these were differentially expressed between subjects with asthma and healthy controls. Fourteen piRNAs were associated with long-term asthma exacerbation, and effect of piRNAs on long-term asthma exacerbation are mediated through eosinophil count and serum IgE level., Conclusion: piRNAs are associated with peripheral blood eosinophils and total serum IgE in childhood asthma and may play important roles in T2-high asthma., (Copyright © 2023 American Academy of Allergy, Asthma & Immunology. All rights reserved.)

Background: IgE has been known for mediating endothelial cell dysfunction and mast cell (MC) activation to fuel asthma-aggravated high-fat diet-induced atherosclerosis. However, it remains unclear for the mechanism of asthma-mediated atherosclerosis, especially the potential involvement of IgE in the exacerbation of asthma-mediated atherosclerosis with a standard laboratory diet, and the cross talk between endothelial cells and MCs., Methods: Asthma-mediated atherosclerosis mice models under a standard laboratory diet and FcεR1 knock-out mice were used to determine the role of IgE-FcεR1 signaling in asthma-mediated atherosclerosis, which was assessed by Oil Red O staining and immunohistochemistry. Various in vitro assays including nanoparticle tracking analysis and transmission electron microscopy were used to evaluate exosome characteristics. Immunofluorescence and fluorescent in situ hybridization approaches were used to evaluate the effect and mechanism of MC-secreted exosomes encapsulated circular RNA CDR1as (cerebellar degeneration-related 1 antisense) on endothelial cells in vivo and in vitro. Finally, cohort studies examined the plasma CDR1as levels in patients with atherosclerosis with or without allergies., Results: Asthma mice with a standard laboratory diet showed increased atherosclerotic lesions and inflammatory infiltration depending on IgE-FcεR1 signal. FcεR1 knockout mice and blockage of IgE-FcεR1 signaling with IgE monoclonal antibody, omalizumab, all significantly alleviated asthma-mediated atherosclerosis and vascular inflammatory remodeling. Anti-inflammation with dexamethasone and stabilization of MC with cromolyn partially alleviated atherosclerotic lesions and mitigated the inflammatory infiltration in arteries. Mechanistically, IgE stimulation upregulates MC CDR1as expression in exosomes and upregulates the endothelial cell adhesive factors VCAM-1 (vascular cell adhesion molecule-1) and ICAM-1 (intercellular adhesion molecule-1) via the CDR1as-FUS (fused in sarcoma)-phos-p65 axis. Knockdown of CDR1as in vivo significantly decreased the endothelial adhesion function and mitigated asthma-mediated atherosclerosis. Furthermore, a cohort study indicated higher plasma CDR1as levels in patients with atherosclerosis with allergies than in patients with atherosclerosis and healthy controls., Conclusions: Exosomes from IgE-stimulated MCs aggravated atherosclerosis through circular RNA CDR1as-mediated endothelial dysfunction, providing a novel insight into asthma-mediated atherosclerosis and potential diagnostic and therapeutic targets., Competing Interests: Disclosures None.

Purpose: Idiopathic generalized epilepsies (IGEs) are a common group of genetic generalized epilepsies with high genetic heterogeneity and complex inheritance. However, the genetic basis is still largely unknown. This study aimed to explore the genetic etiologies in IGEs., Methods: Trio-based whole-exome sequencing was performed in 60 cases with IGEs. The pathogenicity of candidate genetic variants was evaluated by the criteria of the American College of Medical Genetics and Genomics (ACMG), and the clinical causality was assessed by concordance between the observed phenotype and the reported phenotype., Results: Seven candidate variants were detected in seven unrelated cases with IGE (11.7%, 7/60). According to ACMG, a de novo SLC2A1 (c.376C>T/p.Arg126Cys) variant identified in childhood absence epilepsy was evaluated as pathogenic with clinical concordance. Six variants were assessed to be uncertain significance by ACMG, but then considered causative after evaluation of clinical concordance. These variants included CLCN4 hemizygous variant (c.2044G>A/p.Glu682Lys) and IQSEC2 heterozygous variant (c.4315C>T/p.Pro1439Ser) in juvenile absence epilepsy, EFHC1 variant (c.1504C>T/p.Arg502Trp) and CACNA1H (c.589G>T/p.Ala197Ser) both with incomplete penetrance in juvenile myoclonic epilepsy, and GRIN2A variant (c.2011C>G/p.Gln671Glu) and GABRB1 variant (c.1075G>A/p.Val359Ile) both co-segregated with juvenile myoclonic epilepsy. Among them, GABRB1 was for the first time identified as potential novel causative gene for IGE., Significance: Considering the genetic heterogeneity and complex inheritance of IGEs, a comprehensive evaluation combined the ACMG scoring and assessment of clinical concordance is suggested for the pathogenicity analysis of variants identified in clinical screening. GABRB1 is probably a novel causative gene for IGE, which warrants further studies., Competing Interests: Declarations of Competing Interest None of the authors has any conflict of interest to disclose., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Objective: We investigated the frequency of coexistence of temporal lobe epilepsy (TLE) and idiopathic generalized epilepsy (IGE) in a retrospective database study. We also explored the underlying pathomechanisms of the coexistence of TLE and IGE based on the available information, using bioinformatics tools., Methods: The first phase of the investigation was a retrospective study. All patients with an electro-clinical diagnosis of epilepsy were studied at the outpatient epilepsy clinic at Shiraz University of Medical Sciences, Shiraz, Iran, from 2008 until 2023. In the second phase, we searched the following databases for genetic variations (epilepsy-associated genetic polymorphisms) that are associated with TLE or syndromes of IGE: DisGeNET, genome-wide association study (GWAS) Catalog, epilepsy genetic association database (epiGAD), and UniProt. We also did a separate literature search using PubMed., Results: In total, 3760 patients with epilepsy were registered at our clinic; four patients with definitely mixed TLE and IGE were identified; 0.1% of all epilepsies. We could identify that rs1883415 of ALDH5A1, rs137852779 of EFHC1, rs211037 of GABRG2, rs1130183 of KCNJ10, and rs1045642 of ABCB1 genes are shared between TLE and syndromes of IGE., Conclusion: While coexistence of TLE and IGE is a rare phenomenon, this could be explained by shared genetic variations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Allergic asthma (AA) is a prevalent chronic airway inflammation disease. In this study, this study aims to investigate the biological functions and potential regulatory mechanisms of the insulin receptor ( INSR ) in the progression of AA., Methods: BALB/c mice (n = 48) were randomly divided into the following groups: control group, AA group, AA+Lentivirus (Lv)-vector short hairpin RNA (shRNA) group, AA+Lv-vector group, AA+Lv- INSR shRNA group, and AA+Lv- INSR group. The pulmonary index was calculated. mRNA and protein expression levels of INSR , signal transducer and activator of transcription 3 (STAT3), Janus kinase 2 (JAK2), phosphorylated-STAT3 (p-STAT3), phosphorylated-JAK2 (p-JAK2), alpha-smooth muscle actin ( α-SMA ), febrile neutropenia ( FN ), mucin 5AC ( MUC5AC ), and mucin 5B ( MUC5B ) were examined using reverse-transcription quantitative PCR (RT-qPCR) and western blot assays. Positive expressions of INSR , retinoic acid-related orphan receptor gamma-t (RORγt), and forkhead box protein P3 (Foxp3) were quantified by immunohistochemistry. Fluorescence intensities of α-SMA and FN were detected by immunofluorescence. Pathological morphology was observed through hematoxylin-eosin (H&E) staining, Masson staining, and Periodic Acid-Schiff (PAS) staining. Contents of immunoglobulin E (IgE), interleukin-6 (IL-6), eotaxin, interleukin-4 (IL-4), interleukin-13 (IL-13), interferon-γ (IFN-γ), interleukin-17 (IL-17), and interleukin-10 (IL-10) were quantified using enzyme-linked immunosorbent assay (ELISA). The percentage of T helper 17 (Th17) and regulatory T (Treg) cells was determined through flow cytometry., Results: Compared to the control group, expression levels of INSR , p-STAT3, p-JAK2, α-SMA , FN , MUC5AC , MUC5B , RORγt, and Foxp3, as well as IgE, IL-6, eotaxin, IL-4, IL-13, and IL-17 contents, pulmonary index, glycogen-positive area (%), and Th17 cell percentage significantly increased ( p < 0.05). Additionally, pulmonary histopathological deterioration and collagen deposition were aggravated, while Treg cell percentage and IFN-γ and IL-10 contents remarkably decreased ( p < 0.05). The overexpression of INSR further exacerbated the progression of allergic asthma, but the down-regulation of INSR reversed the trends of the above indicators., Conclusions: The down-regulation of INSR alleviates airway hyperviscosity, inflammatory infiltration, and airway remodeling, restoring Th17/Treg immune balance in AA mice by inactivating the STAT3 pathway.

The hyper-immunoglobulin E syndrome (HIES) is a primary immunodeficiency disease originally described as Job syndrome. The fundamental causative variant of the HIES is an autosomal dominant mutation in the signal transducer and activator of transcription 3 (STAT3) gene. It is characterized by recurrent staphylococcal cold skin abscess, sinopulmonary infection, eczema, head and face anomalies, frequent bone fractures, eosinophilia and extremely high serum IgE levels (IgE ≥ 2000 IU/mL). However, multiple other genetic defects are also known as HIES-like disorders. Apart from infectious manifestations, STAT3, DOCK8 and TYK2 gene mutations are associated with various malignancies. The most common malignancies reported in these patients are lymphomas, including Hodgkin's and non-Hodgkin's lymphomas (NHL) of B and T cells. This systematic review aimed to investigate the prevalence of malignancies in HIES and the factors associated with malignancy in these patients. In this survey, all articles published until April 1st, 2023, in Scopus, PubMed and Web of Science databases based on three groups of keywords related to HIES syndrome and malignancy were reviewed by three different researchers. Finally, 26 articles were evaluated from which 24 papers were meta-analyzed. In the current study, the demographic information of 1133 patients with HIES, which was mentioned in 24 articles enrolled in the project, was collected, and the information related to patients who had malignancy was analyzed and meta-analyzed. A total of 96 patients out of 1133 studied patients had at least one type of malignancy, the overall prevalence of malignancies reported in the articles was 6.5% (95% confidence interval 4.1-9%), and the total prevalence of malignancy in patients with NHL type and patients with squamous cell carcinoma (SCC) was 2.9% (95% confidence interval 1.7-4.4%) and 2.2% (95% confidence interval 0.3-4.1%), respectively. The results of this study indicated that in 6.5% of cases, HIES was complicated with malignancy, and considering the higher rate of these malignancies in women as well as in DOCK8 mutation sufferers, it is necessary for physicians to be aware of this association and includes malignancy screening in follow-up and periodic examinations of these patients. Indeed, more studies in this field will help to clarify the precise figures and predisposing factors of the relationship between HIES and malignancy., (© 2023. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Background: Evidence has shown that glucocorticoid-induced transcript 1 (GLCCI1) single nucleotide polymorphism (SNP) rs37937 is associated with asthma., Objectives: The objective of this study was to investigate whether the GLCCI1 SNP rs37937 is a risk factor for allergic rhinitis (AR) in a Chinese Han population., Methods: A total of 220 individuals including 109 AR patients and 111 healthy subjects were included. The genotyping of GLCCI1 rs37973 was performed by the SNaPshot method. The correlations of rs37973 polymorphism, AR risk, and clinical characteristics were further analyzed, as well as the treatment response to intranasal corticosteroids (INCS) in AR patients of different genotypes., Results: Three GLCCI1 rs37973 SNP genotypes were identified in both AR patients and healthy subjects. Significant association between rs37973 polymorphism and AR under allele model, dominant model, heterozygote model, and homozygote model were shown. The A allele frequency of SNP rs37973 in AR was significantly higher than that in controls. The serum total immunoglobulin E (IgE) in AR patients of AA genotype was significantly higher than in patients of GA and GG genotype, and the serum total IgE in GA genotype was significantly higher than in GG genotype. Interestingly, after 4 weeks of INCS treatment for AR patients, the improvement of the nasal itching score, sneezing score, runny nose score, total nasal symptom score, and visual analog scale score of the GG genotype were worse than the AA or GA genotype., Conclusion: The GLCCI1 rs37937 polymorphism is associated with the risk of developing AR and the response to INCS treatment in the Chinese Han population., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated nuclease 9 (Cas9) has emerged as a powerful tool to generate targeted loss-of-function mutations for functional genomic studies. As a next step, tools to generate genome modifications in a spatially and temporally precise manner will enable researchers to further dissect gene function. Here, we present two heat shock-inducible genome-editing (IGE) systems that efficiently edit target genes when the system is induced, thus allowing us to target specific developmental stages. For this conditional editing system, we chose the natural heat-inducible promoter from heat-shock protein 18.2 (HSP18.2) from Arabidopsis thaliana and the synthetic heat-inducible promoter heat shock-response element HSE-COR15A to drive the expression of Cas9. We tested these two IGE systems in Arabidopsis using cyclic or continuous heat-shock treatments at the seedling and bolting stages. A real-time quantitative polymerase chain reaction analysis revealed that the HSP18.2 IGE system exhibited higher Cas9 expression levels than the HSE-COR15A IGE system upon both cyclic and continuous treatments. By targeting brassinosteroid-insensitive 1 (BRI1) and phytoene desaturase (PDS), we demonstrate that both cyclic and continuous heat inductions successfully activated the HSP18.2 IGE system at the two developmental stages, resulting in highly efficient targeted mutagenesis and clear phenotypic outcomes. By contrast, the HSE-COR15A IGE system was only induced at the seedling stage and was less effective than the HSP18.2 IGE system in terms of mutagenesis frequencies. The presented heat shock-IGE systems can be conditionally induced to efficiently inactivate genes at any developmental stage and are uniquely suited for the dissection and systematic characterization of essential genes., (© 2023 The Authors. The Plant Genome published by Wiley Periodicals LLC on behalf of Crop Science Society of America.)

Background: Genomic prediction of breeding values (GP) has been adopted in evolutionary genomic studies to uncover microevolutionary processes of wild populations or improve captive breeding strategies. While recent evolutionary studies applied GP with individual single nucleotide polymorphism (SNP), haplotype-based GP could outperform individual SNP predictions through better capturing the linkage disequilibrium (LD) between the SNP and quantitative trait loci (QTL). This study aimed to evaluate the accuracy and bias of haplotype-based GP of immunoglobulin (Ig) A (IgA), IgE, and IgG against Teladorsagia circumcincta in lambs of an unmanaged sheep population (Soay breed) based on Genomic Best Linear Unbiased Prediction (GBLUP) and five Bayesian [BayesA, BayesB, BayesCπ, Bayesian Lasso (BayesL), and BayesR] methods., Results: The accuracy and bias of GPs using SNP, haplotypic pseudo-SNP from blocks with different LD thresholds (0.15, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, and 1.00), or the combinations of pseudo-SNPs and non-LD clustered SNPs were obtained. Across methods and marker sets, higher ranges of genomic estimated breeding values (GEBV) accuracies were observed for IgA (0.20 to 0.49), followed by IgE (0.08 to 0.20) and IgG (0.05 to 0.14). Considering the methods evaluated, up to 8% gains in GP accuracy of IgG were achieved using pseudo-SNPs compared to SNPs. Up to 3% gain in GP accuracy for IgA was also obtained using the combinations of the pseudo-SNPs with non-clustered SNPs in comparison to fitting individual SNP. No improvement in GP accuracy of IgE was observed using haplotypic pseudo-SNPs or their combination with non-clustered SNPs compared to individual SNP. Bayesian methods outperformed GBLUP for all traits. Most scenarios yielded lower accuracies for all traits with an increased LD threshold. GP models using haplotypic pseudo-SNPs predicted less-biased GEBVs mainly for IgG. For this trait, lower bias was observed with higher LD thresholds, whereas no distinct trend was observed for other traits with changes in LD., Conclusions: Haplotype information improves GP performance of anti-helminthic antibody traits of IgA and IgG compared to fitting individual SNP. The observed gains in the predictive performances indicate that haplotype-based methods could benefit GP of some traits in wild animal populations., (© 2023. The Author(s).)

Background: Bronchiolitis is the leading cause of infant hospitalization in U.S. and is associated with increased risk for childhood asthma. Immunoglobulin E (IgE) not only plays major roles in antiviral immune responses and atopic predisposition, but also offers a potential therapeutic target., Objective: We aimed to identify phenotypes of infant bronchiolitis by using total IgE (tIgE) and virus data, to determine their association with asthma development, and examine their biological characteristics., Methods: In a multicenter prospective cohort study of 1,016 infants (age <1 year) hospitalized for bronchiolitis, we applied clustering approaches to identify phenotypes by integrating tIgE and virus (respiratory syncytial virus [RSV], rhinovirus [RV]) data at hospitalization. We examined their longitudinal association with the risk of developing asthma by age 6 years and investigated their biological characteristics by integrating the upper airway mRNA and microRNA data in a subset (n=182)., Results: In infants hospitalized for bronchiolitis, we identified 4 phenotypes: 1) tIgE low virus RSV-high , 2) tIgE low virus RSV-low/RV , 3) tIgE high virus RSV-high , and 4) tIgE high virus RSV-low/RV phenotypes. Compared to phenotype 1 infants (resembling "classic" bronchiolitis), phenotype 4 infants (tIgE high virus RSV-low/RV ) had a significantly higher risk for developing asthma (19% vs. 43%; adjOR, 2.93; 95% CI, 1.02-8.43; P =.046). Phenotypes 3 and 4 (tIgE high ) had depleted type I interferon and enriched antigen presentation pathways; phenotype 4 also had depleted airway epithelium structure pathways., Conclusions: In this multicenter cohort, tIgE-virus clustering identified distinct phenotypes of infant bronchiolitis with differential risks of asthma development and unique biological characteristics., Competing Interests: Dr. ST receives royalties from UpToDate as a Section Editor and honoraria for lecture from Medscape. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Shibata, Zhu, Ooka, Freishtat, Mansbach, Pérez-Losada, Ramos-Tapia, Teach, Camargo and Hasegawa.)

STAT6 (signal transducer and activator of transcription 6) is a transcription factor that plays a central role in the pathophysiology of allergic inflammation. We have identified 16 patients from 10 families spanning three continents with a profound phenotype of early-life onset allergic immune dysregulation, widespread treatment-resistant atopic dermatitis, hypereosinophilia with esosinophilic gastrointestinal disease, asthma, elevated serum IgE, IgE-mediated food allergies, and anaphylaxis. The cases were either sporadic (seven kindreds) or followed an autosomal dominant inheritance pattern (three kindreds). All patients carried monoallelic rare variants in STAT6 and functional studies established their gain-of-function (GOF) phenotype with sustained STAT6 phosphorylation, increased STAT6 target gene expression, and TH2 skewing. Precision treatment with the anti-IL-4Rα antibody, dupilumab, was highly effective improving both clinical manifestations and immunological biomarkers. This study identifies heterozygous GOF variants in STAT6 as a novel autosomal dominant allergic disorder. We anticipate that our discovery of multiple kindreds with germline STAT6 GOF variants will facilitate the recognition of more affected individuals and the full definition of this new primary atopic disorder., (© 2023 Sharma et al.)

Background: House dust mites (HDM) are among the most important sources for airborne allergens with high relevance for atopic diseases. Routine tests contain only 4 of 32 registered allergens of Dermatophagoides pteronyssinus. Clinical relevance and pathomechanistic properties of many allergens are not well understood., Objective: The association of several HDM allergens with allergic rhinitis, allergic asthma, and atopic dermatitis was investigated to identify allergens with biomarker potential and to transfer them into diagnostics., Methods: Eight out of nine D. pteronyssinus allergens (nDer p 1, rDer p 2, rDer p 5, rDer p 7, rDer p 10, rDer p 13, rDer p 20, rDer p 21, rDer p 23) were recombinantly expressed and purified. Sensitization patterns of 384 HDM-allergic individuals exhibiting different clinical phenotypes were analyzed with a serum-saving multiplex array., Results: Sensitization to more than three mite allergens (sensitization count) was associated with allergic asthma and/or atopic dermatitis. Reactions to Der p 5 and Der p 21 were more frequent in allergic asthma compared to allergic rhinitis. Atopic dermatitis patients were more often sensitized to Der p 5, Der p 20, and Der p 21 among others. Der p 20-IgE > 80 kU/L was associated with severe atopic dermatitis in 75% of patients., Conclusion: This study demonstrates the clinical importance of the sensitization count and of certain allergens (Der p 5, Der p 20, and Der p 21) not available for routine diagnostics yet. Implementing them as well as the sensitization count in diagnostic measures will improve diagnosis and risk assessment of HDM-allergic patients., (© 2022 The Authors. Allergy published by European Academy of Allergy and Clinical Immunology and John Wiley & Sons Ltd.)

Aims: Hypertension is the major cause of cardiovascular diseases and global mortality. Immunoglobulin E (IgE), which plays crucial roles in allergic diseases, has been implicated in the pathogenesis of vascular and cardiac remodelling via its receptor (FcεR1). In this study, we aimed to reveal the role of IgE and FcεR1 in hypertension., Methods and Results: Herein, we reported that IgE levels were significantly increased in hypertensive patients as well as in hypertensive mice induced by angiotensin II (Ang II). Ang II-induced vascular remodelling and hypertension were significantly alleviated in FcεR1 genetic knockout mice or in mice treated with anti-IgE monoclonal antibody. Similarly, treatment with omalizumab (a clinical IgE antagonist) also markedly inhibited Ang II-induced hypertension. Furthermore, the cellular contribution of IgE-FcεR1 in hypertension was evaluated in mice with FcεR1 conditional knockout in mast cell (MC), smooth muscle cell (SMC), or endothelial cell (EC). Our data revealed that IgE-mediated hypertension is largely dependent on FcεR1 in MCs but not SMCs and ECs. Finally, RNA-seq and signalling pathway analyses of mouse bone marrow-derived MCs suggested that interleukin 6 (IL-6) is one of critical mediators in IgE-mediated hypertension. IL-6 derived from IgE-stimulated MCs promoted reactive oxygen species production and decreased the levels of phosphorylated endothelial nitric oxide synthase in ECs, leading to endothelial dysfunction., Conclusion: Our findings reveal that IgE contributes to the pathogenesis of hypertension, at least partially through activating the IgE-FcεR1 signalling in MCs. Thus, IgE may represent a new therapeutic target for IgE-mediated hypertension., Competing Interests: Conflict of interest: none declared., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2022. For permissions, please email: journals.permissions@oup.com.)

Overproduction of mucins in the airways donates largely to airway blockage in asthma patients. Glycoprotein MUC7 plays a role in the clearance of bacteria and has anti-candidacidal criteria. Our goal was to investigate the association between the MUC7 variable number of tandem repeats (VNTR) polymorphism and bronchial asthma among Egyptian children. The MUC7 VNTR polymorphism was investigated among 100 children with bronchial asthma and 100 healthy controls using polymerase chain reaction (PCR) method. Serum levels of immunoglobulin E (IgE), tumor necrosis factor-alpha (TNF-α), and transforming growth factor-beta1 (TGF-β1) were assessed by enzyme-linked immunosorbent assay (ELISA) technique. The frequencies of 6*5 genotype, 5*5 genotype, (6*5 + 5*5) genotypes, and MUC7*5 allele of the MUC7 VNTR variant were significantly lower among asthmatic patients than controls (p < 0.015, OR = 0.39, 95% CI = 0.19-0.81; p = 0.03, OR = 0.18, 95% CI = 0.04-0.86; p < 0.001, OR = 0.29, 95% CI = 0.15-0.58; p < 0.001, OR = 0.3, 95% CI = 0.17-0.55, respectively). The (6*5 + 5*5) genotypes of the MUC7 VNTR variant were not associated with the clinical manifestations and serum levels of IgE, TNF-α, and TGF-β1 among asthmatic patients (p ˃ 0.05). In conclusion, the (6*5 + 5*5) genotypes of the MUC7 VNTR variant may have a protective role for bronchial asthma in Egyptian children., (© 2022. The Author(s).)

Dominant negative mutations in the STAT3 gene account for autosomal dominant hyper-IgE syndrome (AD-HIES). Patients typically present high IgE serum levels, recurrent infections, and soft tissue abnormalities. While current therapies focus on alleviating the symptoms, hematopoietic stem cell transplantation (HSCT) has recently been proposed as a strategy to treat the immunological defect and stabilize the disease, especially in cases with severe lung infections. However, because of the potentially severe side effects associated with allogeneic HSCT, this has been considered only for a few patients. Autologous HSCT represents a safer alternative but it requires the removal of the dominant negative mutation in the patients' cells prior to transplantation. Here, we developed allele-specific CRISPR-Cas9 nucleases to selectively disrupt five of the most common STAT3 dominant negative alleles. When tested ex vivo in patient-derived hematopoietic cells, allele-specific disruption frequencies varied in an allele-dependent fashion and reached up to 62% of alleles harboring the V637M mutation without detectable alterations in the healthy STAT3 allele. However, assessment of the gene expression profiles of the STAT3 downstream target genes revealed that, upon activation of those edited patient cells, mono-allelic STAT3 expression (functional haploinsufficiency) is not able to sufficiently restore STAT3-dependent signaling in edited T cells cultured in vitro. Moreover, the stochastic mutagenesis induced by the repair of the nuclease-induced DNA break could further contribute to dominant negative effects. In summary, our results advocate for precise genome editing strategies rather than allele-specific gene disruption to correct the underlying mutations in AD-HIES.

CBL-B is an E3 ubiquitin ligase that ubiquitinates proteins downstream of immune receptors to downregulate positive signaling cascades. Distinct homozygous mutations in CBLB were identified in 3 unrelated children with early-onset autoimmunity, one of whom also had chronic urticaria. Patient T cells exhibited hyperproliferation in response to anti-CD3 cross-linking. One of the mutations, p.R496X, abolished CBL-B expression, and a second mutation, p.C464W, resulted in preserved CBL-B expression. The third mutation, p.H285L in the SH2 domain of CBL-B, was expressed at half the normal level in the patient's cells. Mice homozygous for the CBL-B p.H257L mutation, which corresponds to the patient's p.H285L mutation, had T and B cell hyperproliferation in response to antigen receptor cross-linking. CblbH257L mice had increased percentages of T regulatory cells (Tregs) that had normal in vitro suppressive function. However, T effector cells from the patient with the p.H285L mutation and CblbH257L mice were resistant to suppression by WT Tregs. Bone marrow-derived mast cells from CblbH257L mice were hyperactivated after FcεRI cross-linking, and CblbH257L mice demonstrated exaggerated IgE-mediated passive anaphylaxis. This study establishes CBL-B deficiency as a cause of immune dysregulation.

A commonly consumed legume in India, the kidney bean (Phaseolus vulgaris) is associated with allergy. We report molecular and immunological characterization of cysteine protease allergen and its cross-reactivity. In silico allergenicity assessment and phylogenetic analysis of kidney bean cysteine protease showed significant sequence homology (upto 67%) with allergens from kiwi, papaya, soybean, ragweed pollen and mites. Physicochemical properties and motif-analysis depicted cysteine protease as probable allergen. Multiple sequence alignment and phylogenetic analysis indicated structural conservation between kidney bean and homologous cysteine protease sequences. The gene was cloned, expressed and affinity purified. Cysteine protease was resolved at 42 kDa and exhibited high IgE binding (up to 89%) with hypersensitive sera. Cysteine protease showed functional property on cross-linking IgE receptors and upregulated expression of CD203c on activated basophils. In inhibition studies, 8.4 ng of cysteine protease was required for 50% self-inhibition, whereas significant inhibition was also observed with kidney bean (52 ng), black gram (155 ng), chick pea (437 ng), mesquite pollen (36 ng), house dust mite (64.85 ng), Alternaria alternata (78.8 ng) and Curvularia lunata (73.6 ng) extracts. ConSurf analysis indicated conserved active site and catalytic residues in mature domain among proteases from legumes, fruits, pollens, mites and fungus. In summary, P. vulgaris cysteine protease was molecularly characterized having functional activity. This study demonstrated, cross-reactivity between food and aeroallergens based on evolutionary conservancy that showed its clinical importance as cross-reactive allergen. PRACTICAL APPLICATIONS: Adaptation of sustainable lifestyle has led to a surge in consumption of plant-based foods especially legumes. Their high nutritional content lowers the risk of developing cardiovascular diseases, diabetes, obesity, and stroke. Kidney beans, a commonly consumed legume in Indian subcontinent, have a potential to be used as nutraceutical and functional food. Despite its alimentary nature, it elicits allergic reactions. Being a major sensitizer, trivial information regarding its allergic components has led to an urgent need for exploring its allergen repertoire. Our study reported biochemical and immunological characterization of its major cysteine protease allergen. Cysteine proteases are major cross-reactive allergens from insects, fruits and fungal sources. Identification and molecular characterization of such immunodominant allergens by RDT offers the prospect of using recombinant proteins for accurate diagnosis and therapeutic purposes. This study suggests that a potential major cross-reactive allergen may aid in developing allergy management interventions for a wide range of allergenic sources., (© 2022 Wiley Periodicals LLC.)

Background: Asthma is chronic immune-mediated airway inflammation, and it is affected by a complex network of interacting cytokines. To date, the exact role of each cytokine and its genetic polymorphisms in childhood asthma development and its severity has remained poorly understood. The purpose of this study was to explore potential roles of four cytokine genes polymorphism and serum levels l [(T helper-2 (Th2) cytokine); Interleukin-4 (IL-4) 590, (Th3 cytokine); and transforming growth factor β1 (TGF-β1) 509T; (Th17) including tumor necrosis factor-alpha (TNF-α), and IL17A rs8193036] in childhood asthma risk and control in Egyptian children, for the 1st time., Materials and Methods: This case-control study included two children subgroups; Group1 included 216 non-asthmatic controls and (Group 2) 216 cases diagnosed with asthma (clinically and spirometry-based) were classified as controlled, partly controlled, and uncontrolled. Polymorphisms of TGF-β1-509, IL-4 590, and TNF-α-308 genes were detected using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). IL-17 was genotyped using tetra-primer amplification refractory mutation system polymerase chain reaction (ARMS-PCR). Serum cytokines levels were measured by enzyme-linked immunosorbent assay (ELISA)., Results: Serum total IgE, TGF-β1, IL4, TNF-α, and IL17A levels were significantly higher in asthmatic compared to controls. Also, significant increases in serum total IgE, IL-4, TGF-β1, and TNF-α levels are combined with poor asthma control, while no significant IL17A changes. There were significant changes of IL-4-590, TNF-α-308, and IL17A genotypes and allele distributions between asthmatic and controls groups as well as different asthma control levels; while no impact of TGF-β1 SNP on asthma risk and control level. Four cytokines SNPs affected their serum levels among asthmatic patients., Conclusion: There are impacts of cytokine gene polymorphisms (IL-4-590, TNF-α-308, and IL17A); but not TGF-β1 on asthma susceptibility and poor asthma control in Egyptian children., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Understanding how individual differences arise and how their effects propagate through groups are fundamental issues in biology. Individual differences can arise from indirect genetic effects (IGE): genetically based variation in the conspecifics with which an individual interacts. Using a clonal species, the Amazon molly ( Poecilia formosa ), we test the hypothesis that IGE can propagate to influence phenotypes of the individuals that do not experience them firsthand. We tested this by exposing genetically identical Amazon mollies to conspecific social partners of different clonal lineages, and then moving these focal individuals to new social groups in which they were the only member to have experienced the IGE. We found that genetically different social environments resulted in the focal animals experiencing different levels of aggression, and that these IGE carried over into new social groups to influence the behaviour of naive individuals. These data reveal that IGE can cascade beyond the individuals that experience them. Opportunity for cascading IGE is ubiquitous, especially in species with long-distance dispersal or fission-fusion group dynamics. Cascades could amplify (or mitigate) the effects of IGE on trait variation and on evolutionary trajectories. Expansion of the IGE framework to include cascading and other types of carry-over effects will therefore improve understanding of individual variation and social evolution and allow more accurate prediction of population response to changing environments.

Cat allergy is a major trigger factor for respiratory reactions (asthma and rhinitis) in patients with immunoglobulin E (IgE) sensitization. In this study, we used a comprehensive panel of purified cat allergen molecules (rFel d 1, nFel d 2, rFel d 3, rFel d 4, rFel d 7, and rFel d 8) that were obtained by recombinant expression in Escherichia coli or by purification as natural proteins to study possible associations with different phenotypes of cat allergy (i.e., rhinitis, conjunctivitis, asthma, and dermatitis) by analyzing molecular IgE recognition profiles in a representative cohort of clinically well-characterized adult cat allergic subjects ( n = 84). IgE levels specific to each of the allergen molecules and to natural cat allergen extract were quantified by ImmunoCAP measurements. Cumulative IgE levels specific to the cat allergen molecules correlated significantly with IgE levels specific to the cat allergen extract, indicating that the panel of allergen molecules resembled IgE epitopes of the natural allergen source. rFel d 1 represented the major cat allergen, which was recognized by 97.2% of cat allergic patients; however, rFel d 3, rFel d 4, and rFel d 7 each showed IgE reactivity in more than 50% of cat allergic patients, indicating the importance of additional allergens in cat allergy. Patients with cat-related skin symptoms showed a trend toward higher IgE levels and/or frequencies of sensitization to each of the tested allergen molecules compared with patients suffering only from rhinitis or asthma, while there were no such differences between patients with rhinitis and asthma. The IgE levels specific to allergen molecules, the IgE levels specific to cat allergen extract, and the IgE levels specific to rFel d 1 were significantly higher in patients with four different symptoms compared with patients with 1-2 symptoms. This difference was more pronounced for the sum of IgE levels specific to the allergen molecules and to cat extract than for IgE levels specific for rFel d 1 alone. Our study indicates that, in addition to rFel d 1, rFel d 3, rFel d 4, and rFel d 7 must be considered as important cat allergens. Furthermore, the cumulative sum of IgE levels specific to cat allergen molecules seems to be a biomarker for identifying patients with complex phenotypes of cat allergy. These findings are important for the diagnosis of IgE sensitization to cats and for the design of allergen-specific immunotherapies for the treatment and prevention of cat allergy.

Early diagnosis of primary immunodeficiencies is crucial for timely treatment and preventing unwanted complications. Next-generation sequencing (NGS) and detailed clinical and immunological evaluation can help early detect such disorders. This study aimed to confirm the diagnosis of two cases of autosomal recessive hyper-immunoglobulin E (IgE) syndrome (AR-HIES), presenting with irreversible eye involvement. Two unrelated patients with suspected AR-HIES were referred to the Immunology, Asthma and Allergy Research Institute (IAARI), Tehran, Iran. Immunological screening tests were performed for AR-HIES, which showed elevated serum IgE levels, eosinophilia, and low T-lymphocyte responses. NGS was performed, and the results were confirmed by Sanger sequencing. Sequence analysis showed a mutation in intron 17 of the dedicator of cytokinesis 8 (DOCK8) gene in the first patient, and a homozygous three base-pair deletion in exon 45 of DOCK8 in the second patient. This is the first time such mutations are reported and these variants are predicted to be damaging. Both patients suffered from persistent viral infections along with cytomegalovirus (CMV) retinitis. Suspicion of these two novel DOCK8 mutations can benefit patients presenting with recalcitrant ophthalmic viral involvements and relevant immunological test results. This would lead to earlier referrals for immunologic and genetic confirmation and thus, a more timely intervention with hematopoietic stem cell transplantation (HSCT).

Background: Peptide-based immunotherapy (PIT) was introduced as an attractive approach in allergen-specific immunotherapy (AIT). However, PIT clinical trials have shown variable results, and immune response to peptides is not precisely predictable. On the other hand, induction of antigen-specific tolerance may be augmented when allergens are combined with the regulatory T cell epitope (Tregitope). This study aimed to evaluate the therapeutic administration of a plasmid DNA encoding Tregitope and ovalbumin (OVA) immunodominant epitope in the murine model of allergy., Methods: Following the induction of allergic rhinitis by ovalbumin, vaccinated group received three doses of recombinant plasmid containing Signal peptide-Tregitope-OVA T cell epitope. After the final OVA challenge, clinical symptoms, histopathological changes, OVA-specific IgE level, and cytokine secretion pattern of spleen cells were examined., Results: Our data are showing that AIT with the recombinant DNA vaccine significantly suppressed airway inflammation; reduced eosinophilic infiltration in the nasal mucosa; decreased expression level of IL-4 and IL-17 in spleen cells, while IFN-γ, IL-10, and TGF-β expression were increased. Moreover, OVA-specific IgE levels were also decreased., Conclusion: These results suggest that Tregitope-immunodominant T cell epitope fusion can act as a safe and effective approach in DNA-based allergen-specific immunotherapy., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Storage mites (SM) may induce allergic respiratory symptoms in sensitized individuals, in both rural and urban settings. The relationship among specific IgE reactions to determined groups of SM allergens in the coincident asthma pheno-endotypes has not yet been investigated. We aimed to study a Precision Allergy Molecular Diagnosis (PAMD@) model to depict the SM molecular profile in individuals presenting with Type-2 inflammation, in two different (moderate and severe) asthma phenotypes. A customized PAMD@ panel, including SM allergens and their concurrent protein allergenic characterization was investigated. Mite group 2 allergens were most frequently recognized, including Lep d 2 (83.45%), followed by Gly d 2 (69.17%) and Tyr p 2 (47,37%), in 133/164 asthmatic subjects. Blo t 5 and Blo t 21 exhibited significant higher titres in both asthma groups. Although relevant mite group 2 allergens cross-reactivity is suggested, individualized sensitization patterns were relevantly identified. The present PAMD@ panel confirmed the dominance of mite group 2 allergens in moderate-to-severe T2 asthmatics. A broadly heterogeneous molecular repertoire of SM allergens was found in all subjects, regardless of their asthma severity. Blomia tropicalis deserves special attention in certain territories, as diagnostic and/or therapeutic approaches merely based on Pyroglyphidae mites may be insufficient.

Background: Allergic diseases figure among the most common immune-mediated diseases worldwide, affecting more than 25% of the world's population. Allergic reactions can be triggered by house dust mite (HDM) allergens, of which the so-called group 21 of allergens is considered as clinically relevant., Methods: Herein, we used a structural bioinformatics and immunoinformatics approach to design hypoallergenic mutant variants of the Der p 21 allergen of Dermatophagoides pteronyssinus, which were then recombinantly expressed in bacteria and tested for their IgE-reactivities. For this, we scanned the wild-type Der p 21 protein for all possible single amino acid substitutions in key IgE-binding regions that could render destabilization of the major epitope regions., Results: Four main substitutions (D82P, K110G, E77G, and E87S) were selected to build mutant variants of the Der p 21 allergen, which were produced in their recombinant forms; two of these variants showed reduced reactivity with IgE. Molecular dynamic simulations and immune simulations demonstrated the overall effects of these mutations on the structural stability of the Der p 21 allergen and on the profile of immune response induced through immunotherapy., Conclusions: When produced in their recombinant forms, two of the Der p 21 mutant variants, namely proteins K110G and E87S, showed significantly reduced IgE reactivities against sera from HDM-allergic individuals (n = 20; p < 0.001)., General Significance: This study successfully translated a rational in silico mutagenesis design into low IgE-binding mutant variants of the allergen rDer p 21. These novel hypoallergens are promising to compose next-generation allergen-immunotherapy formulations in near future., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Crosslinking of FcεRI-bound IgE triggers the release of a large number of biologically active, potentially anaphylactic compounds by mast cells. FcεRI activation ought to be well-controlled to restrict adverse activation. As mast cells are embedded in tissues, adhesion molecules may contribute to limiting premature activation. Here, we report that E-Cadherin serves that purpose. Having confirmed that cultured mast cells express E-Cadherin, a mast-cell-specific E-Cadherin deficiency, Mcpt5-Cre E-Cdh fl/fl mice, was used to analyze mast cell degranulation in vitro and in vivo. Cultured peritoneal mast cells from Mcpt5-Cre E-Cdh fl/fl mice were normal with respect to many parameters but showed much-enhanced degranulation in three independent assays. Soluble E-Cadherin reduced the degranulation of control cells. The release of some newly synthesized inflammatory cytokines was decreased by E-Cadherin deficiency. Compared to controls, Mcpt5-Cre E-Cdh fl/fl mice reacted much stronger to IgE-dependent stimuli, developing anaphylactic shock. We suggest E-Cadherin-mediated tissue interactions restrict mast cell degranulation to prevent their precocious activation., (© 2021 The Authors. European Journal of Immunology published by Wiley-VCH GmbH.)

MicroRNAs (miRNAs, miRs) regulate cell fate decisions by post-transcriptionally tuning networks of mRNA targets. We used miRNA-directed pathway discovery to reveal a regulatory circuit that influences Ig class switch recombination (CSR). We developed a system to deplete mature, activated B cells of miRNAs, and performed a rescue screen that identified the miR-221/222 family as a positive regulator of CSR. Endogenous miR-221/222 regulated B cell CSR to IgE and IgG1 in vitro, and miR-221/222-deficient mice exhibited defective IgE production in allergic airway challenge and polyclonal B cell activation models in vivo. We combined comparative Ago2-HITS-CLIP and gene expression analyses to identify mRNAs bound and regulated by miR-221/222 in primary B cells. Interrogation of these putative direct targets uncovered functionally relevant downstream genes. Genetic depletion or pharmacological inhibition of Foxp1 and Arid1a confirmed their roles as key modulators of CSR to IgE and IgG1., Competing Interests: Disclosures: The authors declare no competing interests exist., (© 2021 Wigton et al.)

Background: Anaphylaxis represents one of the most severe and fatal forms of allergic reactions. Like most other allergies, it is caused by activation of basophils and mast cells by allergen-mediated cross-linking of IgE bound to its high-affinity receptor, FcεRI, on the cell surface. The systemic release of soluble mediators induces an inflammatory cascade, rapidly causing symptoms with peak severity in minutes to hours after allergen exposure. Primary treatment for anaphylaxis consists of immediate intramuscular administration of adrenaline., Objective: While adrenaline alleviates life-threatening symptoms of an anaphylactic reaction, there are currently no disease-modifying interventions available. We sought to develop potent and fast-acting IgE inhibitors with the potential to rapidly terminate acute allergic reactions., Methods: Using affinity maturation by yeast display and structure-guided molecular engineering, we generated 3 optimized disruptive IgE inhibitors based on designed ankyrin repeat proteins and assessed their ability to actively remove IgE from allergic effector cells in vitro as well as in vivo in mice., Results: The engineered IgE inhibitors rapidly dissociate preformed IgE:FcεRI complexes, terminate IgE-mediated signaling in preactivated human blood basophils in vitro, and shut down preinitiated allergic reactions and anaphylaxis in mice in vivo., Conclusions: Fast-acting disruptive IgE inhibitors demonstrate the feasibility of developing kinetically optimized inhibitors for the treatment of anaphylaxis and the rapid desensitization of allergic individuals., (Copyright © 2021 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Mouse IgE and mast cell (MC) functions have been studied primarily using inbred strains. Here, we (a) identified effects of genetic background on mouse IgE and MC phenotypes, (b) defined the suitability of various strains for studying IgE and MC functions, and (c) began to study potentially novel genes involved in such functions. We screened 47 Collaborative Cross (CC) strains, as well as C57BL/6J and BALB/cJ mice, for strength of passive cutaneous anaphylaxis (PCA) and responses to the intestinal parasite Strongyloides venezuelensis (S.v.). CC mice exhibited a diversity in PCA strength and S.v. responses. Among strains tested, C57BL/6J and CC027 mice showed, respectively, moderate and uniquely potent MC activity. Quantitative trait locus analysis and RNA sequencing of BM-derived cultured MCs (BMCMCs) from CC027 mice suggested Sp140 as a candidate gene for MC activation. siRNA-mediated knock-down of Sp140 in BMCMCs decreased IgE-dependent histamine release and cytokine production. Our results demonstrated marked variations in IgE and MC activity in vivo, and in responses to S.v., across CC strains. C57BL/6J and CC027 represent useful models for studying MC functions. Additionally, we identified Sp140 as a gene that contributes to IgE-dependent MC activation.

We collected peripheral blood from thirty-nine elite male endurance runners at rest (24 hours after the last exercise session) and used the Allergy Questionnaire for Athletes score and plasma specific IgE level to separate them into atopic and non-atopic athletes. Neutrophils obtained from atopic and non-atopic athletes were subsequently stimulated in vitro with fMLP (N-formyl-methionyl-leucyl-phenylalanine), LPS (lipopolysaccharide), or PMA (phorbol 12-myristate 13-acetate). Neutrophils from non-atopic runners responded appropriately to LPS, as evidenced by the production of pro (IL-8, TNF-α, and IL-6) and anti-inflammatory (IL-10) cytokines. Neutrophils from atopic elite runners exhibited lower responses to LPS stimulus as indicated by no increase in IL-1β, TNF-α, and IL-6 production. Neutrophils from non-atopic and atopic runners responded similarly to fMLP stimulation, indicating that migration function remained unaltered. Both groups were unresponsive to PMA induced reactive oxygen species (ROS) production. Training hours and training volume were not associated with neutrophil IgE receptor gene expression or any evaluated neutrophil function. Since non-atopic runners normally responded to LPS stimulation, the reduced neutrophil response to the stimuli was most likely due to the atopic state and not exercise training. The findings reported are of clinical relevance because atopic runners exhibit a constant decline in competition performance and are more susceptible to invading microorganisms., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zambonatto, Teixeira, Poma, da Silva, de Almeida, Leite, dos Santos, Alves, Gorjão, Pithon-Curi, Carvalho, Curi and Levada-Pires.)

Rationale: Cardiac fibrosis is an important feature of cardiac remodeling and is a hallmark of heart failure. Recent studies indicate that elevated IgE plays a causal role in pathological cardiac remodeling. However, the underlying mechanism of how IgE promotes cardiac fibrosis has not been fully elucidated. Methods and Results: To explore the function of IgE in cardiac fibrosis, we stimulated mouse primary cardiac fibroblasts (CFs) with IgE and found that both IgE receptor (FcεR1) and fibrosis related proteins were increased after IgE stimulation. Specific deletion of FcεR1 in CFs alleviated angiotensin II (Ang II)-induced cardiac fibrosis in mice. To investigate the mechanisms underlying the IgE-mediated cardiac fibrosis, deep miRNA-seq was performed. Bioinformatics and signaling pathway analysis revealed that IgE upregulated Col1a1 and Col3a1 expression in CFs by repressing miR-486a-5p, with Smad1 participating downstream of miR-486a-5p in this process. Lentivirus-mediated overexpression of miR-486a-5p was found to alleviate Ang II-induced myocardial interstitial fibrosis in mice. Moreover, miR-486-5p serum levels were lower in patients with heart failure than in healthy controls, and were negatively correlated with NT-proBNP levels. Conclusions: Our study demonstrates that elevated IgE promotes pathological cardiac fibrosis by modulating miR-486a-5p and downstream factors, such as Smad1 . These findings suggest new targets for pathological cardiac fibrosis intervention., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Introduction: Asthma with airway mucus hypersecretion is an inadequately characterized variant of asthma. While several studies have reported that hypersecreting patients may carry genetic variants in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, many of those studies have been questioned for their numerous limitations and contradictory results., Objectives: (1) To determine the presence of genetic variants of the CFTR gene in patients with asthma with and without airway mucus hypersecretion. (2) To identify the clinical, inflammatory and functional characteristics of the asthma phenotype with airway mucus hypersecretion., Method: Comparative multicentre cross-sectional descriptive study that included 100 patients with asthma (39 hypersecretors and 61 non-hypersecretors). Asthmatic hypersecretion was defined as the presence of cough productive of sputum on most days for at least 3 months in 2 successive years. The patients were tested for fractional exhaled nitric oxide, spirometry, induced sputum cell count, total immunoglobulin E (IgE), peripheral blood eosinophil count, C-reactive protein, blood fibrinogen and blood albumin and underwent a skin prick test. Asthma control and quality of life were assessed by the Asthma Control Test and Mini Asthma Quality of Life questionnaires, respectively. Blood DNA samples were collected from the patients and next-generation sequencing using a MiSeq sequencer and the Illumina platform was used for the CFTR gene analysis., Results: Genetic differences were observed in the c.1680-870T>A polymorphism of the CFTR gene, significantly more evident in hypersecretors than in non-hypersecretors: 78.94% vs. 59.32% in the majority allele and 21.05% vs. 40.67% in the minority allele (p = 0.036). Clinically, asthma hypersecretors compared to non-hypersecretors were older (57.4 years vs. 49.4 years; p = 0.004); had greater asthma severity (58.9% vs. 23.7%; p = 0.005); experienced greater airway obstruction (FEV1/FVC% 64.3 vs. 69.5; p = 0.041); had poorer asthma control (60% vs. 29%; p = 0.021); had lower IgE levels (126.4 IU/mL vs. 407.6 IU/mL; p = 0.003); and were less likely to have a positive prick test (37.5% vs. 68.85%; p = 0.011)., Conclusion: The results suggest that patients with asthma and with mucus hypersecretion (1) may have a different phenotype and disease mechanism produced by an intronic polymorphism in the CFTR gene (NM&#95;000492.3:c.1680-870T>A), and (2) may have a poorer clinical outcome characterized by severe disease and poorer asthma control with a non-allergic inflammatory phenotype., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: AC has received fees in the last three years for talks at meetings sponsored by Chiesi, Esteve Laboratories, GlaxoSmithKline, Novartis, Ferrer, Zambón and Boehringer Ingelheim, has received travel and attendance expenses for conferences from Novartis, Bial, Teva and FAES Farma and has received funds/grants for research projects from several state agencies and non-profit foundations and from AstraZeneca. EC reports non-financial support from Astrazeneca, personal fees from Boehringer-Ingleheim, personal fees and non-financial support from Chiesi, non-financial support from Novartis, non-financial support from Menarini, non-financial support from ALK, outside the submitted work. SB, EdR, ER, CM, NM, AP, SP, JG, AP, CC, JS and MB have no conflicts of interest to declare. VP has received fees in the last three years for talks at meetings sponsored by AstraZeneca, Boehringer-Ingelheim, MSD and Chiesi, has received travel and attendance expenses for conferences from AstraZeneca, Chiesi and Novartis, has acted as a consultant for ALK, AstraZeneca, Boehringer, MSD, MundiPharma and Sanofi, and has received funds/grants for research projects from several state agencies and non-profit foundations and from AstraZeneca, Chiesi and Menarini. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

STAT3-hyper IgE syndrome (STAT3-HIES) is a primary immunodeficiency presenting with destructive lung disease along with other symptoms. CRISPR-Cas9-mediated adenine base editors (ABEs) have the potential to correct one of the most common STAT3-HIES causing heterozygous STAT3 mutations (c.1144C>T/p.R382W). As a proof-of-concept, we successfully applied ABEs to correct STAT3 p.R382W in patient fibroblasts and induced pluripotent stem cells (iPSCs). Treated primary STAT3-HIES patient fibroblasts showed a correction efficiency of 29% ± 7% without detectable off-target effects evaluated through whole-genome and high-throughput sequencing. Compared with untreated patient fibroblasts, corrected single-cell clones showed functional rescue of STAT3 signaling with significantly increased STAT3 DNA-binding activity and target gene expression of CCL2 and SOCS3 . Patient-derived iPSCs were corrected with an efficiency of 30% ± 6% and differentiated to alveolar organoids showing preserved plasticity in treated cells. In conclusion, our results are supportive for ABE-based gene correction as a potential causative treatment of STAT3-HIES.

IgE induced by type 2 immune responses in atopic dermatitis is implicated in the progression of atopic dermatitis to other allergic diseases, including food allergies, allergic rhinitis, and asthma. However, the keratinocyte-derived signals that promote IgE and ensuing allergic diseases remain unclear. Herein, in a mouse model of atopic dermatitis-like skin inflammation induced by epicutaneous Staphylococcus aureus exposure, keratinocyte release of IL‑36α along with IL-4 triggered B cell IgE class-switching, plasma cell differentiation, and increased serum IgE levels-all of which were abrogated in IL-36R-deficient mice or anti-IL‑36R-blocking antibody-treated mice. Moreover, skin allergen sensitization during S. aureus epicutaneous exposure-induced IL-36 responses was required for the development of allergen-specific lung inflammation. In translating these findings, elevated IL‑36 cytokines in human atopic dermatitis skin and in IL‑36 receptor antagonist-deficiency patients coincided with increased serum IgE levels. Collectively, keratinocyte-initiated IL‑36 responses represent a key mechanism and potential therapeutic target against allergic diseases.

Introduction: Hyperimmunoglobulin E syndromes (HIESs) are characterized by a high serum immunoglobulin E (IgE) level, eczematoid rashes, recurrent staphylococcal skin abscesses, and recurrent pneumonia and pneumatocele formation. Autosomal dominant HIES is the most common form of HIES and mainly occurs due to loss-of-function mutations in the Signal Transducer and Activator of Transcription 3 ( STAT3 ) gene ( STAT3 LOF). Case Presentation: We report the case of an 11-year-old Peruvian girl diagnosed with STAT3 LOF caused by p.R382W mutation. She presented with recurrent staphylococcal pneumonia and empyema caused by the rarely reported Achromobacter xylosoxidans , which led to severe destruction of the lung parenchyma, multiple lung surgeries, and the development of bronchopleural fistulas. A laparotomy was also performed, which showed evidence of sigmoid colon perforation. The patient received immunoglobulin replacement therapy (IRT) and antibiotic prophylaxis, and the frequency of her infections has decreased over the past 3 years. Conclusion: This is the first case of STAT3 LOF diagnosed by genomic sequencing in Peru. Patients with this mutation have recurrent pulmonary infections, and require multiple surgical procedures with frequent complications. A. xylosoxidans infection could be related to the prolonged stay in intensive care leading to high mortality; therefore, additional care must be taken when treating patients with this infection. In addition, colonic perforation is a rare complication in STAT3 LOF patients. IRT and antibiotic prophylaxis appear to decrease the frequency of infections and hospitalizations.

Allergic disease is on the rise and yet the underlying cause and risk factors are not fully understood. While lifesaving in many circumstances, the use of antibiotics and the subsequent disruption of the microbiome are positively correlated with the development of allergies. Here, we describe the use of the antibiotic vancomycin in combination with the papain-induced mouse model of allergic disease that allows for the assessment of microbiome perturbations and the impact on allergy development.

Mouse models of allergic conjunctivitis mimic various aspects of human allergic conjunctivitis. They are useful as acute models of allergic conjunctivitis to study immunological aspects of this condition. In this chapter, we will describe ragweed-pollen-induced experimental allergic conjunctivitis (mostly driven by adaptive immunity), and papain-soaked contact lens-induced experimental allergic conjunctivitis (mostly driven by innate immunity). Giemsa staining of histological sections is used for quantification of the number of infiltrating eosinophils, which is useful to evaluate the severity of the allergic inflammation. Immunohistochemical staining and quantitative PCR are used to clarify spatiotemporal expression of proinflammatory molecules in the conjunctival tissue. Flow cytometric analysis of conjunctival tissue is used for the detection of innate lymphoid cell type 2 (ILC2) in the ocular surface tissues.

Mouse models of allergic asthma have been utilized to establish the role of T helper type 2 (Th2) cells in driving lung inflammation, airway hyperresponsiveness, and obstruction. Here, we present the allergic asthma models, in which mice are hypersensitized to ovalbumin (OVA) and house dust mite (HDM). These models mimic the major characteristics of human asthma including the eosinophilic inflammation and hyperactivity of the airway, overproduction of Th2 cytokines in the lung, and elevated total and allergen-specific immunoglobulin E (IgE) in serum.

Allergic asthma is characterized by airway hyperresponsiveness, remodeling, and reversible airway obstruction. This is associated with an eosinophilic inflammation of the airways, caused by inhaled allergens such as house dust mite or grass pollen. The inhaled allergens trigger a type-2 inflammatory response with the involvement of innate lymphoid cells (ILC2) and Th2 cells, resulting in high immunoglobulin E (IgE) antibody production by B cells and mucus production by airway epithelial cells. As a consequence of the IgE production, subsequent allergen reexposure results in a classic allergic response with distinct early and late phases, both resulting in bronchoconstriction and shortness of breath. Allergen-specific immunotherapy (AIT) is the only treatment that is capable of modifying the immunological process underlying allergic responses including allergic asthma. Both subcutaneous AIT (SCIT) as well as sublingual AIT (SLIT) have shown clinical efficacy in long-term suppression of the allergic response. Although AIT treatments are very successful for rhinitis, application in asthma is hampered by variable efficacy, long duration of treatment, and risk of severe side effects. A more profound understanding of the mechanisms by which AIT induces tolerance to allergens in sensitized individuals is needed to be able to improve its efficacy. Mouse models have been very valuable in preclinical research for characterizing the mechanisms of desensitization in AIT and evaluating novel approaches to improve its efficacy. Here, we present a rapid and reproducible mouse model for allergen-specific immunotherapy. In this model, mice are sensitized with two injections of allergen adsorbed to aluminum hydroxide, followed by subcutaneous injections (SCIT) or sublingual administrations (SLIT) of allergen extracts as an immunotherapy treatment. Finally, mice are challenged by intranasal allergen administrations. We will also describe the protocols as well as the most important readout parameters for the measurements of invasive lung function, serum immunoglobulin levels, isolation of bronchoalveolar lavage fluid (BALF), and preparation of cytospin slides. Moreover, we describe how to perform ex vivo restimulation of lung single-cell suspensions with allergens, flow cytometry for identification of relevant immune cell populations, and ELISAs and Luminex assays for assessment of the cytokine concentrations in BALF and lung tissue.

Food allergy has been rising in prevalence over the last two decades, affecting more than 10% of the world population. Current management of IgE-mediated food allergy relies on avoidance and rescue medications; research into treatments that are safer and providing guaranteed and durable curative effects is, therefore, essential. T-cell epitope-based immunotherapy holds the potential for modulating food allergic responses without IgE cross-linking. In this chapter, we describe the methods in evaluating the therapeutic capacities of immunodominant T-cell epitopes in animal models of food allergy. Moreover, we explain in detail the methods to measure the allergen-specific antibody levels, prepare single-cell suspension from spleen, and prepare small intestine for immunohistochemical analysis of eosinophils and Foxp3+ cells.

Loss-of-function (LOF) mutations in signal transducer and activator of transcription 3 (STAT3) gene causes autosomal dominant hyper immunoglobulin E syndrome (AD-HIES or Job's Syndrome), a rare and complex primary immunodeficiency (PID) syndrome characterized by increased levels of IgE (>2000 IU/mL), eosinophilia, recurrent staphylococcal skin abscesses, eczema, recurrent pneumonia, skeletal and connective tissue abnormalities. Although bacterial and fungal infections are common in AD-HIES, susceptibility to parasitic infections has not been reported. Alveolar echinococcosis (AE), a zoonosis caused by the growth of the Echinococcus multilocularis (EM) metacestode, mimics slow-growing liver cancer. The mortality rate of AE is very high when it is diagnosed late or under-treated. Here, we report a 14-year-old boy with AE infections of the liver and the lung resulting in liver failure and diagnosed as STAT3-LOF. To our knowledge, the association between these two conditions has not been reported in the literature before.

Particulate matter 2.5 (PM 2.5 ) exposure can increase the prevalence of allergic rhinitis (AR), the mechanism underlying which may include oxidative stress and inflammatory response. As a ROS quenching agent, N-acetylcysteine (NAC) can attenuate the accumulation of inflammatory cells and hyper-responsiveness in animal asthma models. To explore the effect of NAC on the oxidative stress and inflammatory reactions in AR rats exposed to PM 2.5 , we analyzed the components of PM 2.5 and examined the nasal symptoms, redox level in nasal mucosa, Th1/Th2-related serum cytokines, nasal mucosal histopathology and ultrastructure in AR rat models with NAC intervention after PM 2.5 exposure. The results showed that the high concentrations of metal cations and PAHs in PM 2.5 could aggravate Th2-dominant allergic inflammation in AR model and cause redox imbalance, accompanied by nasal epithelial cell stripping and eosinophil infiltration, while NAC intervention could alleviate the clinical symptoms of AR model after PM 2.5 exposure, correct the redox imbalance, reduce the Th2 cytokines, reduce eosinophil infiltration, and promote the moderate regeneration of epithelial cells. The mechanism of NAC reversing PM 2.5 -mediated action may be related to its anti-oxidant and anti-inflammatory effects, which may provide some new insights for the prevention of AR exacerbated by exposure to PM 2.5 ., Competing Interests: Declaration of competing interest The authors have no competing interests to declare., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Boosting the production of recombinant therapeutic antibodies is crucial in both academic and industry settings. In this work, we investigated the usage of varying signal peptides by antibody V-genes and their roles in recombinant transient production, systematically comparing myeloma and the native signal peptides of both heavy and light chains in 168 antibody permutation variants. We found that amino acids count and types (essential or non-essential) were important factors in a logistic regression equation model for predicting transient co-transfection protein production rates. Deeper analysis revealed that the culture media were often incomplete and that the supplementation of essential amino acids can improve the recombinant protein yield. While these findings are derived from transient HEK293 expression, they also provide insights to the usage of the large repertoire of antibody signal peptides, where by varying the number of specific amino acids in the signal peptides attached to the variable regions, bottlenecks in amino acid availability can be mitigated., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Ling, Su, Lua, Poh, Ng, Wipat and Gan.)

Adipose tissue is a major source of circulating exosomal microRNAs (miRNAs) that are modulators of the immune response in various types of tissues and organs, including airways. Still, no evidence exists if allergic airway inflammation may affect fat tissue inflammation via alterations in the miRNA expression profile. Therefore, we investigated the miRNA expression profile in the adipose tissue upon induced allergic inflammation in the airways in the rat. Brown Norway rats were chronically sensitized to house dust mite extract for seven weeks. Body composition was performed using MiniSpec Plus. The eosinophil count and the total IgE level were determined to confirm the induction of allergic inflammation. MiRNA expression profiling was done using the next-generation sequencing with validation by qPCR. We found that allergic airway inflammation significantly increased fat in adipose tissue, glucose concentration, and the gene expression of adipose tissue-derived proinflammatory peptides (leptin, TNFα). In miRNA-seq analysis, we showed significant differences in the expression of 36 mature miRNAs, three precursors, and two miRNA families in adipose tissue of allergic rats. Two miRNAs-miRNA-151-5p and miRNA-423-3p-showed significantly increased expression in qPCR in adipose tissue and lungs of sensitized animals. Allergic airway inflammation affects fat tissue and alters miRNA expression profile in adipose tissue in the rat.

Mast cells (MCs) are immune cells that act as environment resident sentinels playing a crucial role in Th2-mediated immune responses, including allergic reactions. Distinguishing features of MCs are the presence of numerous cytoplasmic granules that encapsulate a wide array of preformed bio-active molecules and the constitutive expression of the high affinity receptor of IgE (FcεRI). Upon FcεRI engagement by means of IgE and multivalent antigens, aggregated receptors trigger biochemical pathways that ultimately lead to the release of granule-stored and newly synthesized pro-inflammatory mediators. Additionally, MCs are also able to release exosomes either constitutively or upon stimulation. Exosomes are nanosized vesicles of endocytic origin endowed with important immunoregulatory properties, and represent an additional way of intercellular communication. Interestingly, exosomes generated upon FcεRI engagement contain co-stimulatory and adhesion molecules, lipid mediators, and MC-specific proteases, as well as receptor subunits together with IgE and antigens. These findings support the notion that FcεRI signaling plays an important role in influencing the composition and functions of exosomes derived by MCs depending on their activation status.