Pathogenesis, immunology, and immune-targeted management of the multisystem inflammatory syndrome in children (MIS-C) or pediatric inflammatory multisystem syndrome (PIMS): EAACI Position Paper.

Multisystem inflammatory syndrome in children (MIS-C) is a rare, but severe complication of coronavirus disease 2019 (COVID-19). It develops approximately 4 weeks after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and involves hyperinflammation with multisystem injury, commonly progressing to shock. The exact pathomechanism of MIS-C is not known, but immunological dysregulation leading to cytokine storm plays a central role. In response to the emergence of MIS-C, the European Academy of Allergy and Clinical Immunology (EAACI) established a task force (TF) within the Immunology Section in May 2021. With the use of an online Delphi process, TF formulated clinical statements regarding immunological background of MIS-C, diagnosis, treatment, follow-up, and the role of COVID-19 vaccinations. MIS-C case definition is broad, and diagnosis is made based on clinical presentation. The immunological mechanism leading to MIS-C is unclear and depends on activating multiple pathways leading to hyperinflammation. Current management of MIS-C relies on supportive care in combination with immunosuppressive and/or immunomodulatory agents. The most frequently used agents are systemic steroids and intravenous immunoglobulin. Despite good overall short-term outcome, MIS-C patients should be followed-up at regular intervals after discharge, focusing on cardiac disease, organ damage, and inflammatory activity. COVID-19 vaccination is a safe and effective measure to prevent MIS-C. In anticipation of further research, we propose a convenient and clinically practical algorithm for managing MIS-C developed by the Immunology Section of the EAACI.

Loss of regulatory capacity in Treg cells following rhinovirus infection.

BACKGROUND: Respiratory infections with rhinoviruses (RV) are strongly associated with development and exacerbations of asthma, and they pose an additional health risk for subjects with allergy. OBJECTIVE: How RV infections and chronic allergic diseases are linked and what role RV plays in the breaking of tolerance in regulatory T (Treg) cells is unknown. Therefore, this study aims to investigate the effects of RV on Treg cells. METHODS: Treg cells were isolated from subjects with asthma and controls after experimental infection with the RV-A16 (RV16) and analyzed with next-generation sequencing. Additionally, suppression assays, quantitative PCR assays, and protein quantifications were performed with Treg cells after in vitro RV16 infection. RESULTS: RV16 induced a strong antiviral response in Treg cells from subjects with asthma and controls, including the upregulation of IFI44L, MX1, ISG15, IRF7, and STAT1. In subjects with asthma, the inflammatory response was exaggerated and showed a dysregulated immune response compared with that in the controls. Furthermore, subjects with asthma failed to upregulate several immunosuppressive molecules such as CTLA4 and CD69, and they upregulated the inflammasome-related genes PYCARD and AIM2. Additionally, RV16 reduced the suppressive capacity of Treg cells from healthy subjects and subjects with asthma in vitro and increased TH2 cell-type cytokine production. CONCLUSIONS: Treg cells from healthy subjects and subjects with asthma displayed an antiviral response after RV infection and showed reduced suppressive capacity. These data suggest that Treg cell function might be altered or impaired during RV infections, which might play an important role in the association between RV and the development of asthma and asthma exacerbations.

Trained immunity and tolerance in innate lymphoid cells, monocytes, and dendritic cells during allergen-specific immunotherapy.

BACKGROUND: Despite the efficacy of allergen-specific immunotherapy (AIT), the role of trained immunity and tolerance in this process has not been elucidated. OBJECTIVE: Here, we have performed a comprehensive longitudinal analysis of the systemic innate immune cell repertoire during the course of AIT. METHODS: Patients with allergy received standard preseasonal subcutaneous AIT with allergoids to birch and/or grass. Healthy controls were monitored without any intervention. Flow cytometry of innate lymphoid cell (ILC), natural killer cell, monocyte cell, and dendritic cell (DC) subsets was performed at baseline, 3 months (birch season), 6 months (grass seasons), and 12 months after the therapy in patients or at similar seasonal time points in controls. Additional analyses were performed in the third-year birch and grass season. RESULTS: We observed a durable decrease in group 2 ILCs and an increase of group 1 ILCs after AIT, with dynamic changes in their composition. We found that an expansion of CD127+CD25++ clusters caused observed shifts in the heterogeneity of group 1 ILCs. In addition, we observed development of CD127+CD25++c-Kit+ group 3 ILC clusters. Moreover, we found an increase in the number of intermediate monocytes in parallel with a reduction in nonclassical monocytes during the first year after AIT. Classical and intermediate monocytes presented significant heterogeneity in patients with allergy, but AIT reduced the HLA-DR++ clusters. Finally, an increase in plasmacytoid DCs and CD141+ myeloid DCs was observed in individuals with allergy, whereas the number of CD1c+ myeloid DCs was reduced during the first year of AIT. CONCLUSION: AIT induces changes in the composition and heterogeneity of circulating innate immune cells and brings them to the level observed in healthy individuals. Monitoring of ILCs, monocytes, and DCs during AIT might serve as a novel biomarker strategy.

Monocarbonyl Analogs of Curcumin Based on the Pseudopelletierine Scaffold: Synthesis and Anti-Inflammatory Activity.

Curcumin (CUR) is a natural compound that exhibits anti-inflammatory, anti-bacterial, and other biological properties. However, its application as an effective drug is problematic due to its poor oral bioavailability, solubility in water, and poor absorption from the gastrointestinal tract. The aim of this work is to synthesize monocarbonyl analogs of CUR based on the 9-methyl-9-azabicyclo[3.2.1]nonan-3-one (pseudopelletierine, granatanone) scaffold to improve its bioavailability. Granatane is a homologue of tropane, whose structure is present in numerous naturally occurring alkaloids, e.g., l-cocaine and l-scopolamine. In this study, ten new pseudopelletierine-derived monocarbonyl analogs of CUR were successfully synthesized and characterized by spectral methods and X-ray crystallography. Additionally, in vitro test of the cytotoxicity and anti-inflammatory properties of the synthesized compounds were performed.

Optimization of Novel Human Acellular Dermal Dressing Sterilization for Routine Use in Clinical Practice.

Gamma rays and electrons with kinetic energy up to 10 MeV are routinely used to sterilize biomaterials. To date, the effects of irradiation upon human acellular dermal matrices (hADMs) remain to be fully elucidated. The optimal irradiation dosage remains a critical parameter affecting the final product structure and, by extension, its therapeutic potential. ADM slides were prepared by various digestion methods. The influence of various doses of radiation sterilization using a high-energy electron beam on the structure of collagen, the formation of free radicals and immune responses to non-irradiated (native) and irradiated hADM was investigated. The study of the structure changes was carried out using the following methods: immunohistology, immunoblotting, and electron paramagnetic resonance (EPR) spectroscopy. It was shown that radiation sterilization did not change the architecture and three-dimensional structure of hADM; however, it significantly influenced the degradation of collagen fibers and induced the production of free radicals in a dose-dependent manner. More importantly, the observed effects did not disrupt the therapeutic potential of the new transplants. Therefore, radiation sterilization at a dose of 35kGy can ensure high sterility of the dressing while maintaining its therapeutic potential.

Immunology of COVID-19: Mechanisms, clinical outcome, diagnostics, and perspectives-A report of the European Academy of Allergy and Clinical Immunology (EAACI).

With the worldwide spread of the novel severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) resulting in declaration of a pandemic by the World Health Organization (WHO) on March 11, 2020, the SARS-CoV-2-induced coronavirus disease-19 (COVID-19) has become one of the main challenges of our times. The high infection rate and the severe disease course led to major safety and social restriction measures worldwide. There is an urgent need of unbiased expert knowledge guiding the development of efficient treatment and prevention strategies. This report summarizes current immunological data on mechanisms associated with the SARS-CoV-2 infection and COVID-19 development and progression to the most severe forms. We characterize the differences between adequate innate and adaptive immune response in mild disease and the deep immune dysfunction in the severe multiorgan disease. The similarities of the human immune response to SARS-CoV-2 and the SARS-CoV and MERS-CoV are underlined. We also summarize known and potential SARS-CoV-2 receptors on epithelial barriers, immune cells, endothelium and clinically involved organs such as lung, gut, kidney, cardiovascular, and neuronal system. Finally, we discuss the known and potential mechanisms underlying the involvement of comorbidities, gender, and age in development of COVID-19. Consequently, we highlight the knowledge gaps and urgent research requirements to provide a quick roadmap for ongoing and needed COVID-19 studies.

Laundry detergents and detergent residue after rinsing directly disrupt tight junction barrier integrity in human bronchial epithelial cells.

BACKGROUND: Defects in the epithelial barrier have recently been associated with asthma and other allergies. The influence of laundry detergents on human bronchial epithelial cells (HBECs) and their barrier function remain unknown. OBJECTIVE: We investigated the effects of laundry detergents on cytotoxicity, barrier function, the transcriptome, and the epigenome in HBECs. METHODS: Air-liquid interface cultures of primary HBECs from healthy control subjects, patients with asthma, and patients with chronic obstructive pulmonary disease were exposed to laundry detergents and detergent residue after rinsing. Cytotoxicity and epithelial barrier function were evaluated. RNA sequencing, Assay for Transposase Accessible Chromatin with high-throughput sequencing, and DNA methylation arrays were used for checking the transcriptome and epigenome. RESULTS: Laundry detergents and rinse residue showed dose-dependent toxic effects on HBECs, with irregular cell shape and leakage of lactate dehydrogenase after 24 hours of exposure. A disrupted epithelial barrier function was found with decreased transepithelial electrical resistance, increased paracellular flux, and stratified tight junction (TJ) immunostaining in HBECs exposed to laundry detergent at 1:25,000 dilutions or rinse residue at further 1:10 dilutions. RNA sequencing analysis showed that lipid metabolism, apoptosis progress, and epithelially derived alarmin-related gene expression were upregulated, whereas cell adhesion-related gene expression was downregulated by laundry detergent at 1:50,000 dilutions after 24 hours of exposure without substantially affecting chromatin accessibility and DNA methylation. CONCLUSION: Our data demonstrate that laundry detergents, even at a very high dilution, and rinse residue show significant cell-toxic and directly disruptive effects on the TJ barrier integrity of HBECs without affecting the epigenome and TJ gene expression.

Tight junction, mucin, and inflammasome-related molecules are differentially expressed in eosinophilic, mixed, and neutrophilic experimental asthma in mice.

BACKGROUND: Asthma is a chronic respiratory disease with marked clinical and pathophysiological heterogeneity. Specific pathways are thought to be involved in the pathomechanisms of different inflammatory phenotypes of asthma; however, direct in vivo comparison has not been performed. METHODS: We developed mouse models representing three different phenotypes of allergic airway inflammation-eosinophilic, mixed, and neutrophilic asthma via different methods of house dust mite sensitization and challenge. Transcriptomic analysis of the lungs, followed by the RT-PCR, western blot, and confocal microscopy, was performed. Primary human bronchial epithelial cells cultured in air-liquid interface were used to study the mechanisms revealed in the in vivo models. RESULTS: By whole-genome transcriptome profiling of the lung, we found that airway tight junction (TJ), mucin, and inflammasome-related genes are differentially expressed in these distinct phenotypes. Further analysis of proteins from these families revealed that Zo-1 and Cldn18 were downregulated in all phenotypes, while increased Cldn4 expression was characteristic for neutrophilic airway inflammation. Mucins Clca1 (Gob5) and Muc5ac were upregulated in eosinophilic and even more in neutrophilic phenotype. Increased expression of inflammasome-related molecules such as Nlrp3, Nlrc4, Casp-1, and IL-1ß was characteristic for neutrophilic asthma. In addition, we showed that inflammasome/Th17/neutrophilic axis cytokine-IL-1ß-may transiently impair epithelial barrier function, while IL-1ß and IL-17 increase mucin expressions in primary human bronchial epithelial cells. CONCLUSION: Our findings suggest that differential expression of TJ, mucin, and inflammasome-related molecules in distinct inflammatory phenotypes of asthma may be linked to pathophysiology and might reflect the differences observed in the clinic.

Selected commensal bacteria change profiles of Helicobacter pylori-induced T cells via dendritic cell modulation.

BACKGROUND: The mechanisms of downregulation of protective immunity against Helicobacter pylori (Hp) infection strongly depend on dendritic cell (DC)-induced T-lymphocyte differentiation pattern. Lactic acid bacteria (LAB) strains can modulate Hp-induced immunoresponse by changes in DC activation profiles. Here, we want to find out if the LAB-pulsed DCs will change Hp-induced T-cell responsiveness patterns. MATERIALS AND METHODS: The naive peripheral CD4+ T cells were co-cultured with Hp CagA + pulsed monocyte-derived DCs (DC/CD4+ T cell) in the presence/absence of the feces-derived probiotics: antagonistic or non-antagonistic to Hp (Lactobacillus rhamnosus 900, Lr, Lactobacillus paracasei 915, Lp, respectively), as assessed by the agar slab method. The regulatory T-cell (Treg) population was assessed by flow cytometry, and IFN-γ, IL-12p70, IL-10, and IL-17A levels were evaluated by ELISA method. RESULTS: The Hp-pulsed DC/CD4+ T-cell co-cultures were characterized by high IL-10, decreased IL-12p70 and IFN-γ levels, and elevated Treg population. In contrast, Lr-pulsed DC/CD4+ T-cell co-cultures expressed low IL-10, high IL-12p70 and IFN-γ levels and declined Treg population; this responsiveness pattern was not changed by Hp. The responsiveness pattern of the Lp/Hp-pulsed DC/CD4+ T-cell co-cultures did not differ from those pulsed with Hp alone. CONCLUSION: In contrast to Lp, Lr probiotic strain overcomes Hp-mediated immune profile in the DC/T-cell co-cultures toward Th1 pattern and limited generation of Tregs in vitro. Lr may therefore be used as a component of anti-Hp treatment.

Gastric cancer increases transmigratory potential of peripheral blood monocytes by upregulation of ß1- and ß2-integrins.

INTRODUCTION: Immune responses within the tumor depend on the ability of leukocytes to migrate from peripheral circulation into the local microenvironment. This process is controlled by mechanisms that guide leukocytes to the side of inflammation, allowing them to cross vascular endothelial barrier. Monocytes/macrophages are the predominant population of leukocyte infiltrate of many tumors, including, gastric cancer. However, to date mechanisms that control monocyte trafficking to the side of tumor growth are not fully elucidated. AIM OF THE STUDY: It this study we aimed to evaluate transmigratory potential of peripheral blood monocytes from gastric cancer patients. MATERIAL AND METHODS: By using multicolor flow cytometry we assessed expression of ß1- and ß2-integrins on peripheral blood monocytes from gastric cancer patients. RESULTS: We found increased frequencies of VLA-4 and VLA-6 expressing monocytes and increased expression of analyzed ß2-integrins in gastric cancer patients when compared to age matched controls. CONCLUSIONS: In summary, this study revealed that gastric cancer increases transmigratory potential of peripheral blood monocytes.

Interleukins (from IL-1 to IL-38), interferons, transforming growth factor ß, and TNF-α: Receptors, functions, and roles in diseases.

There have been extensive developments on cellular and molecular mechanisms of immune regulation in allergy, asthma, autoimmune diseases, tumor development, organ transplantation, and chronic infections during the last few years. Better understanding the functions, reciprocal regulation, and counterbalance of subsets of immune and inflammatory cells that interact through interleukins, interferons, TNF-α, and TGF-ß offer opportunities for immune interventions and novel treatment modalities in the era of development of biological immune response modifiers particularly targeting these molecules or their receptors. More than 60 cytokines have been designated as interleukins since the initial discoveries of monocyte and lymphocyte interleukins (called IL-1 and IL-2, respectively). Studies of transgenic or gene-deficient mice with altered expression of these cytokines or their receptors and analyses of mutations and polymorphisms in human genes that encode these products have provided essential information about their functions. Here we review recent developments on IL-1 to IL-38, TNF-α, TGF-ß, and interferons. We highlight recent advances during the last few years in this area and extensively discuss their cellular sources, targets, receptors, signaling pathways, and roles in immune regulation in patients with allergy and asthma and other inflammatory diseases.

MicroRNA modulators of epigenetic regulation, the tumor microenvironment and the immune system in lung cancer.

Cancer is an exceedingly complex disease that is orchestrated and driven by a combination of multiple aberrantly regulated processes. The nature and depth of involvement of individual events vary between cancer types, and in lung cancer, the deregulation of the epigenetic machinery, the tumor microenvironment and the immune system appear to be especially relevant. The contribution of microRNAs to carcinogenesis and cancer progression is well established with many reports and investigations describing the involvement of microRNAs in lung cancer, however most of these studies have concentrated on single microRNA-target relations and have not adequately addressed the complexity of their interactions. In this review, we focus, in part, on the role of microRNAs in the epigenetic regulation of lung cancer where they act as active molecules modulating enzymes that take part in methylation-mediated silencing and chromatin remodeling. Additionally, we highlight their contribution in controlling and modulating the tumor microenvironment and finally, we describe their role in the critical alteration of essential molecules that influence the immune system in lung cancer development and progression.

Differentiation of morphotic elements in human blood using optical coherence tomography and a microfluidic setup.

We demonstrate a novel optical method for the detection and differentiation between erythrocytes and leukocytes that uses amplitude and phase information provided by optical coherence tomography (OCT). Biological cells can introduce significant phase modulation with substantial scattering anisotropy and dominant forward-scattered light. Such physical properties may favor the use of a trans-illumination imaging technique. However, an epi-illumination mode may be more practical and robust in many applications. This study describes a new way of measuring the phase modulation introduced by flowing microobjects. The novel part of this invention is that it uses the backscattered signal from the substrate located below the flowing/moving objects. The identification of cells is based on phase-sensitive OCT signals. To differentiate single cells, a custom-designed microfluidic device with a highly scattering substrate is introduced. The microchannels are molded in polydimethylsiloxane (PDMS) mixed with titanium dioxide (TiO2) to ensure high scattering properties. The statistical parameters of the measured signal depend on the cells' features, such as their size, shape, and internal structure.

Vitamin D3 Treatment Decreases Frequencies of CD16-Positive and TNF-α-Secreting Monocytes in Asthmatic Patients.

BACKGROUND: Previously, we demonstrated that glucocorticoid (GC) treatment of asthmatic patients resulted in decreasing frequencies of monocyte subsets expressing CD16 and capable of releasing TNF-α. Here, we wished to analyze whether the active form of vitamin D, i.e. vitamin D3, referred to as 1α,25-dihydroxyvitamin D3 [1,25-(OH)2D3] can exert GC-like proapoptotic effects on CD16-positive monocytes and thus decrease the proinflammatory potential of these cells. Finally, we set out to investigate whether the addition of 1,25-(OH)2D3 would facilitate the use of lower doses of GC without decreasing their anti-inflammatory properties. METHODS: Peripheral blood mononuclear cells collected from healthy individuals and asthmatic patients were cultured with 1,25-(OH)2D3 and/or varying doses of GC in the presence or absence of caspase inhibition. The cells were either directly stained for extracellular markers or prestimulated with lipopolysaccharide for the assessment of intracellular cytokine production and then analyzed by flow cytometry. RESULTS: We found that 1,25-(OH)2D3 alone (and in combination with GC) decreased the frequency of CD14++CD16+ and CD14+CD16++ monocytes from asthmatic patients and significantly diminished TNF-α production by the monocytes. With regard to the CD14+CD16++ subset, the monocyte-depleting effects of 1,25-(OH)2D3 were abrogated in the presence of pan-caspase inhibitor, suggesting a proapoptotic mechanism of 1,25-(OH)2D3 action. Interestingly, we found that a combined treatment of 1,25-(OH)2D3 and GC allowed for a 5-fold reduction of the GC dose while maintaining their anti-inflammatory effects. CONCLUSIONS: This study has revealed novel immunomodulatory properties of 1,25-(OH)2D3 directed against monocyte subsets capable of TNF-α production. In addition, our data suggest that the introduction of 1,25-(OH)2D3 to anti-inflammatory therapy would possibly allow for the use of lower doses of GC.

Synthesis and anti-inflammatory activity of new 1,2,4-triazole derivatives.

The series of new 1,2,4-triazole derivatives with methacrylic acid moiety were synthesized and characterized by NMR and IR spectroscopy as well as X-ray crystallography. The influence of newly synthesized compounds on the inflammation on the level of cytokine production and the proliferation of human peripheral blood mononuclear cells (PBMC) were experimentally evaluated. Obtained triazoles showed antiproliferative activity and diverse effects on cytokine production. Two compounds demonstrated potentially anti-inflammatory activity and comparable effects with ibuprofen.

Collaborating with the enemy: function of macrophages in the development of neoplastic disease.

Due to the profile of released mediators (such as cytokines, chemokines, growth factors, etc.), neoplastic cells modulate the activity of immune system, directly affecting its components both locally and peripherally. This is reflected by the limited antineoplastic activity of the immune system (immunosuppressive effect), induction of tolerance to neoplastic antigens, and the promotion of processes associated with the proliferation of neoplastic tissue. Most of these responses are macrophages dependent, since these cells show proangiogenic properties, attenuate the adaptive response (anergization of naïve T lymphocytes, induction of Treg cell formation, polarization of immune response towards Th2, etc.), and support invasion and metastases formation. Tumor-associated macrophages (TAMs), a predominant component of leukocytic infiltrate, "cooperate" with the neoplastic tissue, leading to the intensified proliferation and the immune escape of the latter. This paper characterizes the function of macrophages in the development of neoplastic disease.

Mesenchymal stem cells under epigenetic control - the role of epigenetic machinery in fate decision and functional properties.

Mesenchymal stem cells (mesenchymal stromal cells, MSC) are multipotent stem cells that can differentiate into cells of at least three mesodermal lineages, namely adipocytes, osteoblasts, and chondrocytes, and have potent immunomodulatory properties. Epigenetic modifications are critical regulators of gene expression and cellular differentiation of mesenchymal stem cells (MSCs). Epigenetic machinery controls MSC differentiation through direct modifications to DNA and histones. Understanding the role of epigenetic machinery in MSC is crucial for the development of effective cell-based therapies for degenerative and inflammatory diseases. In this review, we summarize the current understanding of the role of epigenetic control of MSC differentiation and immunomodulatory properties.