Role of activation of the coagulation system in the pathogenesis of urticariaTaro Yasuma,1 Corina N. D’Alessandro-Gabazza,1 Tetsu Kobayashi,2 John Morser,3 Esteban C Gabazza.1*1Department of Immunology,2Department of Pulmonary and Critical Care Medicine, Mie University Faculty and Graduate School of Medicine, and Mie University Hospital, Edobashi 2-174, Tsu, Mie 514-8507, Japan.3Division of Hematology, Stanford University School of Medicine, 291 Campus Drive, Stanford, CA 94305, United States*Correspondence: Esteban C Gabazza, MD, PhD, Department of Immunology, Mie University Graduate School of Medicine, Edobashi 2-174, Postal Code 514-8507, Tsu-city, Mie, Japan. Tel.: +81 59 231 5017; fax: +81 59 231 5225.E-mail:email@example.comWord count: 590
Background The same dosing schedule, 1000 SQ-U times three, with one-month intervals, have been evaluated in most trials of intralymphatic immunotherapy (ILIT) for the treatment of allergic rhinitis (AR). The present studies evaluated if a dose escalation in ILIT can enhance the clinical and immunological effects, without compromising safety. Methods Two randomized double-blind placebo-controlled trials of ILIT for grass pollen induced AR were performed. The first included 29 patients that had recently ended 3 years of SCIT and the second contained 39 not previously vaccinated patients. An up-dosage of 1000-3000-10 000 (5000 + 5000 with 30 minutes apart) SQ-U with one month in between was evaluated. Results Doses up to 10 000 SQ-U was safe after recent SCIT. The combined symptom-medication scores (CSMS) were reduced by 31% and the grass specific IgG4 levels in blood were doubled. In ILIT de novo, the two first patients that received active treatment developed serious adverse reactions at 5000 SQ-U. A modified up-dosing schedule; 1000-3000-3000 SQ-U appeared to be safe but failed to improve the CSMS. Flow cytometry analyses showed increased activation of lymph node derived dendritic but not T-cells. Quality of life and nasal provocation response did not improve in any study. Conclusion ILIT in high doses after SCIT appears to further reduce grass pollen induced seasonal symptoms and may be considered as an add-on treatment for patients that do not reach full symptom control after SCIT. Up-dosing schedules de novo with three monthly injections that exceeds 3 000 SQ-U should be avoided.
Food Allergy (FA) is now one of the most common chronic diseases of childhood often lasting throughout life and leading to significant worldwide healthcare burden. The precise mechanisms responsible for the development of this inflammatory condition are largely unknown; however, a multifactorial aetiology involving both environmental and genetic contributions is well accepted. A precise understanding of the pathogenesis of FA is an essential first step to developing comprehensive prevention strategies that could mitigate this epidemic. As it is frequently preceded by atopic dermatitis and can be prevented by early antigen introduction, the development of FA is likely facilitated by the improper initial presentation of antigen to the developing immune system. Primary oral exposure of antigens allowing for presentation via a well-developed mucosal immune system, rather than through a disrupted skin epidermal barrier, is essential to prevent FA. In this review, we present the data supporting the necessity of 1) an intact epidermal barrier to prevent epicutaneous antigen presentation, 2) the presence of specific commensal bacteria to maintain an intact mucosal immune system and 3) maternal/infant diet diversity, including vitamins and minerals, and appropriately timed allergenic food introduction to prevent FA.
Title: Notch4, uncovering an immunomodulator in allergic asthmaAuthors: Beatriz Moyaa,b, Manali Mukherjeec and Parameswaran Nairca. Department of Allergy, Hospital Universitario 12 de Octubre, Madrid, Spainb. Instituto de Investigación Sanitaria Hospital 12 de Octubre (imas12), Madrid, Spainc. Division of Respirology, Department of Medicine, McMaster University, Hamilton & Firestone Institute for Respiratory Health, St Joseph’s Healthcare, Hamilton, ON, CanadaCorrespondence to : Beatriz Moya. Allergy Department. Hospital Universitario 12 de Octubre, Madrid, SpainEmail:firstname.lastname@example.orgAcknowledgements : Dr. Mukherjee is supported by investigator award from Canadian Institutes of Health Research and Canadian Allergy, Asthma, and Immunology Foundation. She has received honorarium from AZ, GSK and her university has received grants from Methapharm Speciality Pharmaceuticals. Dr. Nair is supported by the Frederick E. Hargreave Teva Innovation Chair in Airway Diseases. He has received honoraria from AZ, Sanofi, Teva, Merck, Novartis and Equillium and his university has received research grants from AZ, Teva, Sanofi, Novartis, BI and Methapharm. The authors recognize Dr. Anna Globinska for graphical abstract design and Dr. Rodrigo Jiménez-Saiz for critical review of the manuscript.Keywords: Allergic asthma; Airway inflammation; Th2 cell; Th17 cell; Treg cell; Notch4 receptorAbbreviations: Th, T helper; UFPs, pollutant ultrafine particles; AMs, alveolar macrophages; Treg cells, regulatory T cells; ILC2s, type 2 innate lymphoid cells; GDF15, cytokine growth and differentiation factor 15; IL, interleukin; IL-6R, interleukin-6 receptor; IL-4R, interleukin-4 receptorWord count: 918/1000
Methods: Twenty-two French nursing homes were included. COVID-19 had been diagnosed with real-time reverse-transcriptase polymerase chain reaction (RT-PCR) for SARS-CoV-2. Blood S-protein IgG and nucleocapsid (N) IgG protein (N-protein IgG) were measured 21-24 days after the first jab (1,004 residents) and 6 weeks after the second (820 residents). Results: Among the 735 residents without prior COVID-19, 41.7% remained seronegative for S-protein IgG after the first jab vs 2.1% of the 270 residents with a previous positive RT-PCR (p<0.001). After the second jab, only 3% of the 586 residents without prior COVID-19 remained seronegative. However, 26.5% of them had low S-protein IgG levels (50-1050 UA/mL) vs 6.4% of the 222 residents with prior COVID-19. Residents with old infection (first wave), or seropositive for N-protein IgG at the time of vaccination, had the highest S-protein IgG levels. Residents with a prior COVID-19 infection had higher S-protein IgG levels after one dose than those without two jabs. Interpretation: A single vaccine jab is sufficient to reach immunity in residents with prior COVID-19. Most residents without prior COVID-19 are seropositive for S-protein IgG after the second jab, but around 30% have low levels of S-protein IgG. Whether residents with no or low post-vaccine immunity are at higher risk of symptomatic COVID-19 requires further analysis.
Background: Several new variants of SARS-CoV-2 have emerged since fall 2020 which have multiple mutations in the receptor binding domain (RBD) of the spike protein. It is unclear which mutations affect receptor affinity versus immune recognition. Methods: We produced RBD with single mutations (E484K, K417N or N501Y) or with all three mutations combined and tested their binding to ACE2 by biolayer interferometry (BLI). The ability of convalescent sera to recognize RBDs and block their interaction with ACE2 was tested as well. Results: We demonstrated that single mutation N501Y increased binding affinity to ACE2 but did not significantly affect its recognition by convalescent sera. In contrast, single mutation E484K had almost no impact on the binding kinetics, but essentially abolished recognition of RBD by convalescent sera. Interestingly, combining mutations E484K, K417N and N501Y resulted in a RBD with both features: enhanced receptor binding and abolished immune recognition. Conclusion: Our data demonstrate that single mutations either affect receptor affinity or immune recognition while triple mutant RBDs combine both features.
Metagenomic analysis of the conjunctival bacterial and fungal microbiome in vernal keratoconjunctivitisShort title: Microbiome in VKCText Word Count: 597To the Editor,there is increasing interest in the role of conjunctival microbiome in the healthy ocular surface (1) and in different ocular diseases. There is also enough evidence that dysregulation of resident microbial communities (dysbiosis) might be associated with allergy risk(2). Vernal keratoconjunctivitis (VKC) is a severe form of ocular allergy affecting mostly male children and young adults with typical seasonal recurrences and potentially visual impairment. We recently found and overexpression of multiple pattern recognition receptors (PRRs) in VKC suggesting the role of host-microbe interaction in VKC pathogenesis(3). To investigate the VKC-associated ocular microbiome, we applied 16S and ITS2 amplicon sequencing (online supplementary MM) in conjunctival swabs obtained from 22 VKC patients and 20 age, sex and ethnicity-matched healthy controls (HC) (Table 1S). Written informed consent was obtained from all parent/subjects enrolled. Type of VKC, allergen sensitization, disease-specific ongoing treatment and results of the Quality of Life in Children with VKC (QUICK) questionnaire were recorded. A 10-items questionnaire investigating the presence of the principal factors related to allergy development was administered to all included subjects.16S rRNA amplification was successfully carried out in 12/22 VKC and 4/20 HC samples. High-throughput amplicon sequencing produced a total of 734.157 high-quality reads (average of 45.885 reads/sample), which were clustered into 1.241 OTUs (97% sequence identity) and classified according to the Greengenes database. Compared with HC, α-diversity was significantly higher in all VKC (p=0.05), in IgE-negative patients (p=0.03), in tarsal VKC (p=0.03), in formula-fed versus breastfed children (p=0.03) and in children with history of atopic dermatitis (AD) (p=0.01). β-diversity highlighted differences in microbiota composition (Figure 1S) suggesting clusters of subjects with different conjunctival microbiomes. At the phylum level, conjunctival microbiome was dominated by Proteobacteria , Firmicutes , Actinobacteria andBacteriodetes that accounted for >90% of all reads (Figure 1). Of the 132 observed families, Moraxellaceae (W=15) showed a higher abundance in VKC than HC (Figure 2S). At the genus level, Pseudomonas , Staphylococcus , Streptococcus ,Acinetobacter , Neisseria , Haemophilus ,Prevotella , Corynebacterium , Propionibacterium andRothia accounted for >70% of sequences (Figure 3S). In VKC, Bacteroidetes and Fusobacteria met the core microbiome’s definition criteria (Table 2S), which includes different species of gram-negative bacteria able to potentially induce an LPS-driven inflammatory response as shown in experimental models(4).For the fungal microbiome, 10/22 VKC samples produced a detectable ITS2 amplicon. A total of 677,115 high-quality reads (average of 48,365 reads/sample) were clustered into 933 OTUs and taxonomically classified against the UNITE ver. 7.0 database. At the family level,Saccharomycetaceae , Malasseziaceae , Pleosporaceaeand Cladosporiaceae accounted for the majority of sequences (Figure 2). OTUs referred to Malasseziaceae were significantly increased in VKC compared with HC (W=42). Malassezia have been associated with AD inducing a mixed Th2/Th17 response(5) interacting with several PRRs and activating multiple signaling pathways. Since multiple PRRs are over-expressed in VKC(3), we suggest that glycan, phospholipid carbohydrate residues of allergens and microbes may engage innate receptors on conjunctival cells priming a type-2 response in VKC.14/22 VKC children were under topical treatments at the time of swabbing, creating a potential bias (Table 1S). However, only 5/10 patients whose swabs were not sequenced because of absence of amplicons, were under topical medication. In addition, β-diversity didn’t show differences in microbial communities considering the use and type of medications suggesting that factors other than topical eyedrops may alter the conjunctival microbiota. The main limitation was the higher difficulty to obtain amplicon products from HC, which has been already described and attributed to the physiological antimicrobial activity and to the lower microbial load of healthy subjects(6).In conclusion, the described dysbiosis in VKC highlights the role of the host-microbes interaction in VKC pathogenesis.Andrea Leonardi1Rocco Luigi Modugno1Fabiano Cavarzeran1Umberto Rosani21 Department of Neuroscience, Ophthalmology Unit, University of Padova, Padova, Italy2 Department of Biology, University of Padova, Padova, ItalyKeywords: ocular surface microbiome, vernal keratoconjunctivitis, 16S rRNA gene amplicon sequencing, ITS2 rRNA gene amplicon sequencing, core microbiomeCorresponding author:Andrea Leonardi, MDDepartment of Neuroscience, Ophthalmology Unit, University of Paduavia Giustiniani 235128 Padua, ItalyFax: +39-049-875 5168Email: email@example.comAcknowledgmentsa. Funding/Support: Supported by MIUR DOR1952345/19 and DOR2092417/20 from Italian Institute of Health.b. Conflicts of Interest:Andrea Leonardi: No Conflicts of Interest.Rocco Luigi Modugno: No Conflicts of InterestFabiano Cavarzeran: No Conflicts of InterestUmberto Rosani: No Conflicts of Interestc. Contributions to Authors in each of these areas: Andrea Leonardi (AL), Rocco Luigi Modugno (RLM), Fabiano Cavarzeran (FC), Umberto Rosani (UR)Conception and Design: AL, URAnalysis and interpretation: AL, RLM, URWriting the article: AL, RLMCritical revision of the article: URFinal approval of the article: AL, RLM, URData Collection: RLM, UR, FCProvision of materials, patients, or resources: AL, URStatistical expertise: FCObtaining funding: ALLiterature search: RLM, AL, URAdministrative, technical or logistic support: AL, FCd. Statement about Conformity with Author Information: nonee. Other Acknowledgments: none
Long Term Disruption of Cytokine Signalling Networks are Evident Following SARS-CoV-2 InfectionSinead Ahearn-Ford1, Nonhlanhla Lunjani1, Brian McSharry1,2, John MacSharry1,2,3, Liam Fanning1,3, Gerard Murphy4, Cormac Everard4, Aoife Barry4, Aimee McGreal4, Sultan Mohamed al Lawati4, Susan Lapthorne4, Colin Sherlock4, Anna McKeogh4, Arthur Jackson4, Eamonn Faller4, Mary Horgan3,4, Corinna Sadlier4, Liam O’Mahony1,2,3*1APC Microbiome Ireland, University College Cork, Cork, Ireland2School of Microbiology, University College Cork, Cork, Ireland3Department of Medicine, University College Cork, Cork, Ireland4 Department of Infectious Diseases, Cork University Hospital, Cork, Ireland*Corresponding author – firstname.lastname@example.orgTo the Editor,The current pandemic caused by the SARS-CoV-2 virus has so far infected more than 130 million people worldwide, resulting in approximately 3 million deaths. While the current clinical and public health priorities are designed to limit severe acute and fatal episodes of the disease, and to quickly roll out vaccines to the general population, it has become apparent that there may also be significant detrimental long-term effects following SARS-CoV-2 infection that impact daily functioning and quality of life1. The mechanisms underpinning the post-acute sequelae of SARS-CoV-2 infection’s long-lasting symptoms can include direct effects of the infection (e.g. endothelial damage, lung fibrosis) or indirect effects associated with changes in the microbiome or abnormalities in inflammatory and immune signalling pathways stimulated by the infection2,3.In order to examine the potential long-term immune changes that occur following elimination of the primary infection, we examined the levels of 52 cytokines and growth factors (using MSD multiplex kits) in the serum of patients that attended follow-up post-COVID infection clinics at Cork University Hospital, Cork, Ireland (The Clinical Research Ethics Committee of the Cork Teaching Hospitals approved this study and all patients provided informed consent). All patients had been hospitalised for PCR-proven SARS-CoV-2 infection (median in-patient stay of 5.5 days, range 1 day to 24 days) during the first wave of the pandemic in Ireland (March-May 2020). 38 serum samples were obtained from 24 patients (median age 53.5 years, 11 female) at 3-9 months following hospital discharge. Clinical severity ranged from mild to critical during hospitalisation and the most common symptoms at follow-up clinics were fatigue and/or dyspnoea (supplementary Table S1). Sera obtained prior to the pandemic from 29 healthy volunteers (median age 43.2 years, 14 female) were analysed in parallel.Of the 52 analytes measured, 19 were significantly elevated in post-COVID patient sera compared to healthy controls (supplementary Table S2). These 19 mediators are illustrated as dot plots in Figure 1 and Figure 2. One group of mediators, c-reactive protein (CRP), serum amyloid A (SAA), Interleukin-1 receptor antagonist (IL-1RA), IL-6, IL-8, IL-15, IL-16, monocyte chemotactic protein (MCP)-1 and MCP-4, can be broadly categorised as being associated with ongoing inflammatory responses (Figure 1a)4. These mediators remained as elevated in samples taken 6-9 months following hospital discharge as those levels observed 3-6 months following discharge (p<0.05 versus controls, ANOVA). A second group of mediators, vascular endothelial growth factor (VEGF-A), soluble tyrosine-protein kinase receptor Tie-2 (Tie-2), soluble intercellular adhesion molecule (ICAM-1) and basic fibroblast growth factor (bFGF), can be generally associated with endothelial dysfunction, remodelling and angiogenesis (Figure 1b)5. The remaining elevated mediators are associated with patterns of lymphocyte polarisation. Elevated IL-4, macrophage-derived chemokine (MDC) and thymic stromal lymphopoietin (TSLP) sera levels indicate activation of TH2 responses (Figure 2a), while IL-17A, macrophage inflammatory protein (MIP)-3α and IL-12/23p40 indicate ongoing TH17 activity (Figure 2b). Other indicators of TH2-associated activities are just outside statistical significance (IL-5, p=0.06; supplementary Table S2). While TH1 responses are well described to be upregulated during acute infection6, the levels of these mediators (e.g. IFN-γ, IP-10) decrease following elimination of the virus and are at control levels in our cohort of post-COVID patients (supplementary Table S2).Our data suggests that there are long term immunological consequences following SARS-CoV-2 infection, at least in those that had acute symptoms severe enough to require hospitalisation. While the relatively low number of patients included in our study at this stage does not allow us to perform subgroup analysis, it is possible that these immune mediators may associate with clinically meaningful disease variables and ultimately may be of therapeutic value, if findings are replicated in future studies. Of particular interest is the elevation in TH2-associated mediators. Could this response be a component of the mucosal repair mechanisms that occur following viral damage, or does this indicate new TH2-associated pathological immune activity that might underpin an increased risk of developing allergy or asthma? Clearly the potential immune mechanisms underpinning the emerging post-COVID clinical entities will become increasingly more important to understand as the health care systems adapt to caring for large numbers of COVID-19 survivors during the coming months and years.
Progressive knowledge of allergenic structures resulted in a broad availability of allergenic molecules for diagnosis. Component resolved diagnosis allowed a better understanding of patient sensitization patterns, facilitating allergen immunotherapy decisions. In parallel to the discovery of allergenic molecules, there was a progressive development of a regulation framework that affected both in vitro diagnostics and Allergen Immunotherapy products. With a progressive understanding of underlying mechanisms associated to Allergen immunotherapy and an increasing experience of application of molecular diagnosis in daily life, we focus in analyzing the evidences of the value provided by molecular allergology in daily clinical practice, with a focus on Allergen Immunotherapy decissions.
Advances in molecular biology alongside the accelerated development of gene and cell engineering have contributed to the development of several endotype-targeted biological therapies against chronic immune-mediated allergic diseases. Conventional therapies for asthma, chronic rhinosinusitis with polyposis (CRSwNP), chronic spontaneous urticaria and atopic dermatitis (AD) are not without limitations, and as such the advent of biological therapies have provided a promising alternative treatment option. Biologicals have proven efficacious in the treatment of refractory chronic spontaneous urticaria, asthma, AD, CRSwNP and there is increasing evidence for their utility in treating food allergy. Biologicals are applied and investigated for the most urgent need: acute treatment, symptom control and reduction of steroid usage. Currently there are five approved biologicals for allergic disease management, targeted against IgE (omalizumab), type 2 (T2) cytokines and cytokine receptors (IL-4Ra; dupilumab, IL-5; mepolizumab/reslizumab, IL-5Ra; benralizumab).
Because of their selectivity, biologicals are crucial therapeutic agents in oncological, immunological, and inflammatory diseases and their use in clinical practice is broadening. Biologicals are among the most common drugs that can cause hypersensitivity reactions (HSRs), and this is primarily attributed to an explosion in new treatment options that has developed through personalized and precision medicine. Patients can develop HSRs to these agents during the first lifetime exposure or after repeated exposure. Despite its relatively high prevalence, the underlying mechanisms and optimal management of HSRs to biologicals remain incompletely explained. In this position paper, the authors provided evidence-based recommendations for the diagnosis and management of HSRs to biologicals. Additionally, the document defines unmet needs, which should be topics of future studies.
The TGF-β-Th2 axis: a new target for cancer therapy?García de Durango C1, Escribese MM2, Rosace D3.1: DKTK Research Group, Oncogenic Signalling Pathways of Colorectal Cancer, Institute of Pathology, Ludwig-Maximilians-Universität München, Munich, Germany.2: Institute of Applied Molecular Medicine (IMMA), Department of Basic Medical Sciences, Facultad de Medicina, Universidad San Pablo-CEU, CEU Universities, Urbanización Montepríncipe, 28660 Boadilla del Monte, Madrid, Spain.3: Centro De Investigación Del Cáncer and Instituto De Biología Molecular Y Celular Del Cáncer, Consejo Superior De Investigaciones Científicas (CSIC) - Universidad de Salamanca, Salamanca, Spain.Correspondence :Domenico Rosace, Centro De Investigación Del Cáncer and Instituto De Biología Molecular Y Celular Del Cáncer, Consejo Superior De Investigaciones Científicas (CSIC) - University of Salamanca, Salamanca, Spain. email@example.com
Immunoglobulin E (IgE)-mediated allergy is the most common hypersensitivity disease affecting more than 30% of the population. In genetically-predisposed subjects exposure to minute quantities of allergens leads to the production of IgE antibodies which is termed allergic sensitization and mainly occurs in early childhood. Allergen-specific IgE then binds to the high (FcRI) and low affinity receptors (FcRII, also called CD23) for IgE on effector cells and antigen-presenting cells, respectively. Subsequent and repeated allergen exposure increases allergen-specific IgE levels and, by receptor cross-linking, triggers immediate release of inflammatory mediators from mast cells and basophils whereas IgE-facilitated allergen presentation perpetuates T cell-mediated allergic inflammation. Due to engagement of receptors which are highly selective for IgE even tiny amounts of allergens can induce massive inflammation. Naturally occurring allergen-specific IgG and IgA antibodies usually recognize different epitopes on allergens compared to IgE, and do not efficiently interfere with allergen-induced inflammation. However IgG and IgA antibodies to these important IgE epitopes can be induced by allergen-specific immunotherapy or by passive immunization. These will lead to competition with IgE for binding with the allergen and prevent allergic responses. Similarly, anti-IgE treatment does the same by preventing IgE from binding to its receptor on mastcells and basophils. Here we review the complex interplay of allergen-specific IgE, IgG and IgA and the corresponding cell receptors in allergic diseases and its relevance for diagnosis, treatment and prevention of allergy.
Increase of allergic conditions has occurred at the same pace with the Great Accleration, which stands for the rapid growth rate of human activities upon Earth from 1950s. Changes of environment and lifestyle along with escalating urbanization, are acknowledged as the main underlying causes. Secondary (tertiary) prevention for better disease control has advanced considerably with innovations for oral immunotherapy and effective treatment of inflammation with corticosteroids, calcineurin inhibitors and biologic medications. Patients are less disabled than before. However, primary prevention has remained a dilemma. Factors predicting allergy and asthma risk have proven complex: risk factors increase the risk while protective factors counteract them. Interaction of human body with environmental biodiversity with micro-organisms and biogenic compounds as well as the central role of epigenetic adaptation in immune homeostasis have given new insight. Allergic diseases are good indicators of the twisted relation to environment. In various non-communicable diseases, the protective mode of the immune system indicates low-grade inflammation without apparent cause. Giving microbes, pro- and prebiotics, has shown some promise in prevention and treatment. The real-world public health programme in Finland (2008-2018) emphasized nature relatedness and protective factors for immunological resilience, instead of avoidance. The nationwide action mitigated the allergy burden, but in the lack of controls, primary preventive effect remains to be proven. The first results of controlled biodiversity interventions are promising. In the fastly urbanizing world, new approaches are called for allergy prevention, which also has a major cost saving potential.