Almut Meyer-Bahlburg

Deficiency of adenosine deaminase 2 (DADA2) is an autoinflammatory disease with diverse phenotypes. We describe the genetics, phenotypes, and treatment of n = 48 DADA2 patients from Germany, Austria, and Switzerland. We report a high incidence of hematological (83%) and immunological features (85%), a comparatively low anti-tumor necrosis factor full-response rate (58%), and a high decision probability (21%) for hematopoietic cell transplantation (HCT). We establish a correlation between genetic variant ADA2 activity and patient ADA2 activity. Remarkably, lower patient ADA2 activity is predictive of neutropenia and shows a trend toward HCT decision, whereas higher patient ADA2 activity is predictive of vasculitis symptoms. Genetic variants with low residual ADA2 activity are significantly more common among patients receiving HCT. Our study corroborates previous observations connecting ADA2 activity and clinical phenotype, which up to now have been mainly based on in vitro data.

Background: Limited data on SARS-CoV-2 seroprevalence in rural areas of northern Germany necessitate comprehensive cohort studies. We aimed to evaluate the seroprevalence, silent infection (SI) rates and risk factors for infections among children and adolescents in Western Pomerania from December 2020 to August 2022. Methods: In this cross-sectional study, serum or plasma samples from children and adolescents (6 months to 17 years) were collected during routine blood draw. SARS-CoV-2 specific antibodies (S1 and nucleocapsid) and their neutralizing capacity were analyzed using commercially available enzyme-linked immunosorbent and neutralization assays. Socio-demographic data and information about SARS-CoV-2 infection or vaccination were obtained. Multivariable logistic regression was used to identify independent risk factors for SARS-CoV-2 infections and SI. Results: A total of 1,131 blood samples were included into the study. Overall, SARS-CoV-2 seroprevalence was 25.1%, strongly influenced by the pandemic course, predominant virus variants, age and approval of vaccination. SI rate was 5.4% (95%-CI [3.7%–6.8%]) among unvaccinated and undiagnosed children over the entire study period with highest rates among adolescents. Main risk factor despite the time at risk for silent infections was an infected household member (Odds ratio = 9.88, 95%-CI [4.23–22.9], p < 0.001). Factors associated with overall infections (known and silent) also include the infection of a household member (Odds ratio = 17.8, 95%-CI [10.7–29.6], p < 0.001). Conclusions: We believe that the introduction of governmental measures and systematic test strategies in schools strongly impacted on the SI rate, as we suspect that asymptomatic cases have already been identified, resulting in surprisingly low SI identified in our study.

Inflammasome activation occurs in various diseases, including rare diseases that require multicenter studies for investigation. Flow cytometric analysis of ASC speck⁺ cells in patient samples can be used to detect cell type-specific inflammasome activation. However, this requires standardized sample processing and the ability to compare data from different flow cytometers. To address this issue, we analyzed stimulated and unstimulated PBMCs from healthy donors using seven different flow cytometers. Additionally, human PBMCs were analyzed by fluorescence microscopy, imaging flow cytometry and high-content imaging (HCI). Flow cytometers differed significantly in their ability to detect ASC speck⁺ cells. Aria III, Astrios EQ, and Canto II performed best in separating ASC speck⁺ from diffuse ASC cells. Imaging flow cytometry and HCI provided additional insight into ASC speck formation based on image-based parameters. For optimal results, the ability to separate cells with diffuse ASC from ASC speck⁺ cells is decisive. Image-based parameters can also differentiate cells with diffuse ASC from ASC speck⁺ cells. For the first time, we analyzed ASC speck detection by HCI in PBMCs and demonstrated advantages of this technique, such as high-throughput, algorithm-driven image quantification and 3D-rendering. Thus, inflammasome activation by ASC speck formation can be detected by various technical methods. However, the results may vary depending on the device used.

Monitoring the seroprevalence of SARS-CoV-2 in children and adolescents can provide valuable information for effective SARS-CoV-2 surveillance, and thus guide vaccination strategies. In this study, we quantified antibodies against the spike S1 domains of several SARS-CoV-2 variants (wild-type, Alpha, Delta, and Omicron variants) as well as endemic human coronaviruses (HCoVs) in 1,309 children and adolescents screened between December 2020 and March 2023. Their antibody binding profiles were compared with those of 22 pre-pandemic samples from children and adolescents using an in-house Luminex^®-based Corona Array (CA). The primary objectives of this study were to (i) monitor SARS-CoV-2-specific antibodies in children and adolescents, (ii) evaluate whether the S1-specific antibody response can identify the infecting variant of concern (VoC), (iii) estimate the prevalence of silent infections, and (iv) test whether vaccination or infection with SARS-CoV-2 induce HCoV cross-reactive antibodies. Both SARS-CoV-2 infection and vaccination induced a robust antibody response against the S1 domain of WT and VoCs in children and adolescents. Antibodies specific for the S1 domain were able to distinguish between SARS-CoV-2 VoCs in infected children. The serologically identified VoC was typically the predominant VoC at the time of infection. Furthermore, our highly sensitive CA identified more silent SARS-CoV-2 infections than a commercial ELISA (12.1% vs. 6.3%, respectively), and provided insights into the infecting VoC. Seroconversion to endemic HCoVs occurred in early childhood, and vaccination or infection with SARS-CoV-2 did not induce HCoV S1 cross-reactive antibodies. In conclusion, the antibody response to the S1 domain of the spike protein of SARS-CoV-2 is highly specific, providing information about the infecting VoC and revealing clinically silent infections.

Zusammenfassung Schon zu Beginn der Corona-Pandemie 2020 wurde SARS-CoV‑2 als „Autoimmunvirus“ bezeichnet. Seither gibt es zahlreiche Berichte über das vermehrte Auftreten von Autoantikörpern und Autoimmunphänomenen nach SARS-CoV-2-Infektionen. Dabei können Autoantikörper einerseits den Krankheitsverlauf beeinflussen, andererseits zur Erstmanifestation von Autoimmunerkrankungen führen. Zudem wird auch für die Pathogenese von Post-COVID eine Rolle von Autoantikörpern diskutiert. Im vorliegenden Beitrag werden dazu wesentliche Aspekte und Studien aufgeführt, und es wird dargestellt, welche möglichen therapeutischen Konsequenzen sich aus den Erkenntnissen ergeben.

Objective: The formation of large intracellular protein aggregates of the inflammasome adaptor ASC is a hallmark of inflammasome activation and characteristic of autoinflammation. Inflammasome activated cells release the highly proinflammatory cytokine IL-1β in addition to ASC specks into the extracellular space. Autoinflammatory activity has been demonstrated in systemic JIA, however minimal data exist on the role of inflammasomes in other JIA subtypes. We therefore investigated, if pyroptotic cells are present in the circulation of oligo- and poly-articular JIA.Methods: Peripheral blood of JIA patients (n = 46) was investigated for ASC speck formation, a key step in inflammasome activation, by flow cytometry and immunofluorescence. Free ASC and proinflammatory cytokine levels were determined by ELISA and multiplex assay.Results: Oligo-articular JIA patients showed a significantly increased proportion of ASC speck⁺ monocytes compared to poly-articular JIA patients. In serum free ASC alone is not sufficient to assess inflammasome activity and does not correlate with ASC speck⁺ monocytes. Compared to control several cytokines were significantly elevated in samples of JIA patients. JIA serum containing antinuclear antibodies, incubated with ASC specks boosts a secondary inflammation by IL-1β production in macrophages.Conclusion: For the first time, we detect ex vivo inflammasome activation by ASC speck formation in oligo- and poly-articular JIA patients. Most notably, inflammasome activation was significantly higher in oligo- compared to poly-articular JIA patients. This data suggests that inflammasome derived autoinflammation may have a greater influence in the previously thought autoimmune oligo-articular JIA patients.

Rhinoviruses (RV) account for a significant number of asthma exacerbations, and RV species C may be associated with a severe course in vulnerable patient groups. Despite important evidence on the role of RV reported by clinicians and life scientists, there are still unanswered questions regarding their influence on asthma exacerbation in young patients. Thus, we measured the RVspecies-specific IgG titers in our German pediatric exacerbation cohort using a microarray-based technology. For this approach, human sera of patients with exacerbated asthma and wheeze, as well as healthy control subjects (n = 136) were included, and correlation analyses were performed. Concordantly with previously published results, we observed significantly higher cumulative levels of RV species A-specific IgG (p = 0.011) and RV-C-specific IgG (p = 0.051) in exacerbated asthma group compared to age-matched controls. Moreover, atopic wheezers had increased RV-specific IgG levels for species A (p = 0.0011) and species C (p = 0.0009) compared to non-atopic wheezers. Hypothesizing that bacterial infection positively correlates with immune memory against RV, we included nasopharyngeal swab results in our analyses and detected limited correlations. Interestingly, the eosinophil blood titer positively correlated with RV-specific IgG levels. With these observations, we add important observations to the existing data regarding exacerbation in pediatric and adolescent medicine. We propose that scientists and clinicians should pay more attention to the relevance of RV species in susceptible pediatric patients.

Inflammasome activation is linked to the aggregation of the adaptor protein ASC into a multiprotein complex, known as the ASC speck. Redistribution of cytosolic ASC to this complex has been widely used as a readout for inflammasome activation and precedes the downstream proteolytic release of the proinflammatory cytokines, IL-1β and IL-18. Although inflammasomes are important for many diseases such as periodic fever syndromes, COVID-19, gout, sepsis, atherosclerosis and Alzheimer’s disease, only a little knowledge exists on the precise and cell type specific occurrence of inflammasome activation in patient samples ex vivo. In this report, we provide detailed information about the optimal conditions to reliably identify inflammasome activated monocytes by ASC speck formation using a modified flow cytometric method introduced by Sester et al. in 2015. Since no protocol for optimal sample processing exists, we tested human blood samples for various conditions including anticoagulant, time and temperature, the effect of one freeze–thaw cycle for PBMC storage, and the fast generation of a positive control. We believe that this flow cytometric protocol will help researchers to perform high quality translational research in multicenter studies, and therefore provide a basis for investigating the role of the inflammasome in the pathogenesis of various diseases.

Die Klassifikation der juvenilen idiopathischen Arthritis (JIA) steht derzeit im Fokus vieler Diskussionen. In dem Übersichtsartikel stellt ein internationales Autor*innenteam aus den USA, Großbritannien, Kanada und Italien unter der Seniorautorenschaft von Alberto Martini die derzeit verwendete ILAR-Klassifikation einer möglichen zukünftigen Klassifikation basierend auf biologischen Charakteristika gegenüber. Publication Date: 19 August 2021 (online) © 2021. Thieme. All rights reserved. Georg Thieme Verlag KG Rüdigerstraße 14, 70469 Stuttgart, Germany