Pressure-dependent performance of CEN-specified Condensation Particle Counters

One of the most important parameters to quantify an aerosol is the particle number concentration. Condensation Particle Counters (CPCs) are commonly used to measure the aerosol number concentration in the nanometer range. To compare the data from different measurement stations and campaigns it is important to harmonize the instrument specifications, which is why the Technical Specification CEN/TS 16976:2016 was introduced for CPCs. There, the parameters of the CEN-CPC are specified for standard pressure and temperature. However, CEN-CPCs are used in various surroundings, on high mountains or on airplanes, where they are exposed to low-pressure conditions. Here, we present the pressure-dependent performance (including the concentration linearity and counting efficiency) of two different models of CEN-CPCs, the Grimm 5410 CEN and the TSI 3772-CEN. We found that their performance at 1000 hPa and 750 hPa was in accordance with the CEN-technical-specifications. Below 500 hPa, the performance decreased for both CPC-models, but the decrease was different for the two models. To gain insight into the performance of the two CPC-models, we performed a simulation study. This study included simulations of the saturation profiles and calculations of internal particle losses within the CPCs. The simulations reproduced the overall performance decrease with decreasing pressure and reveal that the internal structure of the CPC has a significant influence on the performance. We anticipate our publication to provide a deeper understanding of the counting efficiency of CPCs and their pressure dependence. Our findings might be a starting point for new standards that include the pressure-dependent performance or they could help for designing new CPCs.</p

The site of allergen expression in hematopoietic cells determines the degree and quality of tolerance induced through molecular chimerism

The transplantation of allergens (e.g. Phl p 5 or Bet v 1) expressed on BM cells as membrane-anchored full-length proteins leads to permanent tolerance at the T-cell, B-cell, and effector-cell levels. Since the exposure of complete allergens bears the risk of inducing anaphylaxis, we investigated here whether expression of Phl p 5 in the cytoplasm (rather than on the cell surface) is sufficient for tolerance induction. Transplantation of BALB/c BM retrovirally transduced to express Phl p 5 in the cytoplasm led to stable and durable molecular chimerism in syngeneic recipients (∼20% chimerism at 6 months). Chimeras showed allergen-specific T-cell hyporesponsiveness. Further, Phl p 5-specific TH1-dependent humoral responses were tolerized in several chimeras. Surprisingly, Phl p 5-specific IgE and IgG1 levels were significantly reduced but still detectable in sera of chimeric mice, indicating incomplete B-cell tolerance. No Phl p 5-specific sIgM developed in cytoplasmic chimeras, which is in marked contrast to mice transplanted with BM expressing membrane-anchored Phl p 5. Thus, the expression site of the allergen substantially influences the degree and quality of tolerance achieved with molecular chimerism in IgE-mediated allergy

Cell Therapy for Prophylactic Tolerance in Immunoglobulin E-mediated Allergy

AbstractBackgroundTherapeutic strategies for the prophylaxis of IgE-mediated allergy remain an unmet medical need. Cell therapy is an emerging approach with high potential for preventing and treating immunological diseases.We aimed to develop a cell-based therapy inducing permanent allergen-specific immunological tolerance for preventing IgE-mediated allergy.MethodsWild-type mice were treated with allergen-expressing bone marrow cells under a short course of tolerogenic immunosuppression (mTOR inhibition and costimulation blockade). Bone marrow was retrieved from a novel transgenic mouse ubiquitously expressing the major grass pollen allergen Phl p 5 as a membrane-anchored protein (BALB/c-Tg[Phlp5-GFP], here mPhl p 5). After transplantation recipients were IgE-sensitized at multiple time points with Phl p 5 and control allergen.ResultsMice treated with mPhl p 5 bone marrow did not develop Phl p 5-specific IgE (or other isotypes) despite repeated administration of the allergen, while mounting and maintaining a strong humoral response towards the control allergen. Notably, Phl p 5-specific T cell responses and allergic airway inflammation were also completely prevented. Interestingly allergen-specific B cell tolerance was maintained independent of Treg functions indicating deletional tolerance as underlying mechanism.ConclusionThis proof-of-concept study demonstrates that allergen-specific immunological tolerance preventing occurrence of allergy can be established through a cell-based therapy employing allergen-expressing leukocytes

Impact of Cerebral Microbleeds in Stroke Patients with Atrial Fibrillation

OBJECTIVES: Cerebral microbleeds are associated with the risks of ischemic stroke and intracranial hemorrhage, causing clinical dilemmas for antithrombotic treatment decisions. We aimed to evaluate the risks of intracranial hemorrhage and ischemic stroke associated with microbleeds in patients with atrial fibrillation treated with Vitamin K antagonists, direct oral anticoagulants, antiplatelets, and combination therapy (i.e. concurrent oral anticoagulant and antiplatelet) METHODS: We included patients with documented atrial fibrillation from the pooled individual patient data analysis by the Microbleeds International Collaborative Network. Risks of subsequent intracranial hemorrhage and ischemic stroke were compared between patients with and without microbleeds, stratified by antithrombotic use. RESULTS: A total of 7,839 patients were included. The presence of microbleeds was associated with an increased relative risk of intracranial hemorrhage (aHR 2.74, 95% confidence interval 1.76 - 4.26) and ischemic stroke (aHR 1.29, 95% confidence interval 1.04 - 1.59). For the entire cohort, the absolute incidence of ischemic stroke was higher than intracranial hemorrhage regardless of microbleeds burden. However, for the subgroup of patients taking combination of anticoagulant and antiplatelet therapy, the absolute risk of intracranial hemorrhage exceeded that of ischemic stroke in those with 2-4 microbleeds (25 vs 12 per 1,000 patient-years) and ≥11 microbleeds (94 vs 48 per 1,000 patient-years). INTERPRETATION: Patients with atrial fibrillation and high burden of microbleeds receiving combination therapy have a tendency of higher rate of intracranial hemorrhage than ischemic stroke, with potential for net harm. Further studies are needed to help optimize stroke preventive strategies in this high-risk group. This article is protected by copyright. All rights reserved

Prevention of IgE-mediated allergy by molecular chimerism and costimulation blockade in a murine model

Die IgE–mediierte oder Soforttyp Allergie ist eine Erkrankung mit steigender Tendenz. Zurzeit ist die einzige kausale Therapieform die spezifische Immunotherapie. Allerdings bestehen auch Risiken, wie etwa Neusensibilisierungen oder Anaphylaxie. Stabile und lebenslange Toleranzinduktion wäre eine ideale Prophylaxe und Therapie. Um dieses Ziel zu erreichen wurden zwei verschiedene Ansätze untersucht, Kostimulationsblockade und molekularer Chimärismus. Blockade von Kostimulationssignalen mittels spezifischer Antikörper hat in der Transplantation zu vielversprechenden Ergebnissen geführt. Diese Arbeit setzt sich mit der Kostimulationsblockade in der Allergieprävention und Therapie auseinander. Bei der TH2 Immunantwort werden unter anderem die Zahl und Funktion der Gedächtniszellen durch das Kostimulationsmolekül OX40L reguliert. Welche Rolle OX40L im murinen Allergie-Modell spielt, wurde durch Blockierung dieser Interaktion analysiert. Es wurde auch eine mögliche synergistische Funktion von OX40L mit, CD28 (CTLA4Ig) und CD40L (anti-CD40L) untersucht. Blockierung von OX40L hatte keinen Einfluss auf die allergen-spezifischen Antikörperproduktion oder die T-Zell Antwort. Wogegen die Kombination von anti-CD40L/CTLA4Ig zu einer supprimierten T Zell Antwort und signifikant verzögerter humorale allergische Sensibilisierung führte. Eine andere Strategie, Toleranz zu induzieren ist der molekulare Chiärismus. Molekularer Chimärismus wurde schon erfolgreich in einem Allergie-Mausmodell mit dem Allergen Phl p 5 gezeigt. Da Allergene strukturell und funktionell unterschiedliche Proteine sind, wurde diese Methode mit Bet v 1, dem Hauptbirkenpollenallergen, umgesetzt. Ein Bet v 1-GFP Konstrukt wurde in hämatopoetische Stammzellen eingeschleust und anschließend in vorbehandelte, syngene Mäuse transplantiert. Durch erfolgreiches Engraftment von allergen-exprimiernden Blutzellen im Empfänger wurde Langzeittoleranz gegen dieses Allergen erzielt. Die behandelten Mäuse waren tolerant gegen wiederholte Immunisierungen. Über den gesamten Verlauf entwickelte sich keine allergen-spezifische T-Zell Antwort und auch kein allergen-spezifisches IgE. Darüber hinaus konnte keine Effektorzellaktivität durch das Serum induziert werden. Diese Ergebnisse zeigen, dass OX40L-Blockade keine Auswirkung auf die TH2 Antwort in unserem Modell hat. Wogegen die Blockade von CD40L/CD28 zu einer stark verzögerten Immunantwort führt. Stabile Toleranz auf zellulärer und humoraler Ebene gegenüber einem Allergen konnte nur mit molekularem Chimärismus erreicht werden. Es konnte gezeigt werden, dass molekularer Chimärismus in der Allergie auch mit strukturell und funktional unterschiedlichen Allergenen erfolgreich angewendet werden kann.IgE-mediated allergy is a hypersensitivity disorder whose prevalence is still increasing. IgE is produced by plasma cells after class switching which requires T cell help, costimulation and TH2 cytokines. To date, immunotherapy is the only causative treatment but, next to limited effectiveness, it harbors risks such as anaphylaxis or additional sensitization. Therefore, development of strategies for tolerance induction towards allergens remains a desirable goal. Here we examined costimulation blockade and molecular chimerism as two potential solutions. Costimulation blockade prolongs graft survival in transplantation but its potential in the treatment of allergy still has to be fully investigated. Recently, OX40L was suggested to play an important role in TH2-mediated and memory T cell responses. We therefore analyzed the tolerizing potential of blocking OX40L alone or in addition to CD28 and CD40L blockade in a well-described model of IgE-mediated allergy using clinically relevant major allergens. Blocking OX40L alone had no therapeutic effect. Combination of anti-CD40L/CTLA4Ig significantly delayed production of allergen-specific antibodies. T cell response was suppressed even after established antibody production. Additional anti-OX40L treatment to anti-CD40L/CTLA4Ig showed no synergistic effect. In the second part of my work we examined molecular chimerism as a promising concept in tolerance induction, which is brought about through the transplantation of autologous hematopoietic cells genetically modified in vitro to express a disease-causing antigen. Recently, our group has published a proof of principle study showing robust long-term tolerance towards the grass pollen allergen, Phl p 5, through molecular chimerism. Since allergens are structurally and functionally different, we investigated whether molecular chimerism establishes tolerance towards the major birch pollen allergen, Bet v 1. VSV-Bet v 1-GFP transduced BMC were transplanted into preconditioned recipients. Engraftment of allergen-expressing stem cells was supported by inhibition of CD26 and tracked by the reportergene GFP. Molecular chimerism was detectable throughout long-term follow-up, preventing sensitization even after repeated challenges with Bet v 1. Production of Bet v 1-specific antibodies was completely abrogated throughout follow-up. Moreover, Bet v 1-specific T cell proliferation was suppressed. Our findings show that combined costimulation blockade including OX40L blockade at the time of immunization with an allergen delays but does not prevent the humoral allergic response. In contrast, molecular chimerism induced robust long-term tolerance. Chimerism is a promising strategy to achieve life-long tolerance on the B-, T cell and effector cell levels towards a wide range of allergens

Journal of Immunology Research / CTLA4Ig Improves Murine iTreg Induction via TGF and Suppressor Function In Vitro

Blockade of the CD28:CD80/86 costimulatory pathway has been shown to be potent in blocking T cell activation in vitro and in vivo. The costimulation blocker CTLA4Ig has been approved for the treatment of autoimmune diseases and transplant rejection. The therapeutic application of regulatory T cells (Tregs) has recently gained much attention for its potential of improving allograft survival. However, neither costimulation blockade with CTLA4Ig nor Treg therapy induces robust tolerance on its own. Combining CTLA4Ig with Treg therapy would be an attractive approach for minimizing immunosuppression or for possibly achieving tolerance. However, since the CD28 pathway is more complex than initially thought, the question arose whether blocking CD80/86 would inadvertently impact immunological tolerance by interfering with Treg generation and function. We therefore wanted to investigate the compatibility of CTLA4Ig with regulatory T cells by evaluating direct effects of CTLA4Ig on murine Treg generation and function in vitro. For generation of polyclonal-induced Tregs, we utilized an APC-free in vitro system and added titrated doses of CTLA4Ig at different time points. Phenotypical characterization by flow cytometry and functional characterization in suppressor assays did not reveal negative effects by CTLA4Ig. The costimulation blocker CTLA4Ig does not impair but rather improves murine iTreg generation and suppressor function in vitro.(VLID)470938

Antibiotic Abscess Penetration: Fosfomycin Levels Measured in Pus and Simulated Concentration-Time Profiles

The present study was performed to evaluate the ability of fosfomycin, a broad-spectrum antibiotic, to penetrate into abscess fluid. Twelve patients scheduled for surgical or computer tomography-guided abscess drainage received a single intravenous dose of 8 g of fosfomycin. The fosfomycin concentrations in plasma over time and in pus upon drainage were determined. A pharmacokinetic model was developed to estimate the concentration-time profile of fosfomycin in pus. Individual fosfomycin concentrations in abscess fluid at drainage varied substantially, ranging from below the limit of detection up to 168 mg/liter. The fosfomycin concentrations in pus of the study population correlated neither with plasma levels nor with the individual ratios of abscess surface area to volume. This finding was attributed to highly variable abscess permeability. The average concentration in pus was calculated to be 182 ± 64 mg/liter at steady state, exceeding the MIC(50/90)s of several bacterial species which are commonly involved in abscess formation, such as streptococci, staphylococci, and Escherichia coli. Hereby, the exceptionally long mean half-life of fosfomycin of 32 ± 39 h in abscess fluid may favor its antimicrobial effect because fosfomycin exerts time-dependent killing. After an initial loading dose of 10 to 12 g, fosfomycin should be administered at doses of 8 g three times per day to reach sufficient concentrations in abscess fluid and plasma. Applying this dosing regimen, fosfomycin levels in abscess fluid are expected to be effective after multiple doses in most patients