The hyperinflammatory spectrum: from defects in cytotoxicity to cytokine control

Cytotoxic lymphocytes kill target cells through polarized release of the content of cytotoxic granules towards the target cell. The importance of this cytotoxic pathway in immune regulation is evidenced by the severe and often fatal condition, known as hemophagocytic lymphohistiocytosis (HLH) that occurs in mice and humans with inborn errors of lymphocyte cytotoxic function. The clinical and preclinical data indicate that the damage seen in severe, virally triggered HLH is due to an overwhelming immune system reaction and not the direct effects of the virus per se. The main HLH-disease mechanism, which links impaired cytotoxicity to excessive release of pro-inflammatory cytokines is a prolongation of the synapse time between the cytotoxic effector cell and the target cell, which prompts the former to secrete larger amounts of cytokines (including interferon gamma) that activate macrophages. We and others have identified novel genetic HLH spectrum disorders. In the present update, we position these newly reported molecular causes, including CD48-haploinsufficiency and ZNFX1-deficiency, within the pathogenic pathways that lead to HLH. These genetic defects have consequences on the cellular level on a gradient model ranging from impaired lymphocyte cytotoxicity to intrinsic activation of macrophages and virally infected cells. Altogether, it is clear that target cells and macrophages may play an independent role and are not passive bystanders in the pathogenesis of HLH. Understanding these processes which lead to immune dysregulation may pave the way to novel ideas for medical intervention in HLH and virally triggered hypercytokinemia

Life-Threatening Primary Varicella Zoster Virus Infection With Hemophagocytic Lymphohistiocytosis-Like Disease in GATA2 Haploinsufficiency Accompanied by Expansion of Double Negative T-Lymphocytes

Two unrelated patients with GATA2-haploinsufficiency developed a hemophagocytic lymphohistiocytosis (HLH)-like disease during a varicella zoster virus (VZV) infection. High copy numbers of VZV were detected in the blood, and the patients were successfully treated with acyclovir and intravenous immunoglobulins. After treatment with corticosteroids for the HLH, both patients made a full recovery. Although the mechanisms leading to this disease constellation have yet to be characterized, we hypothesize that impairment of the immunoregulatory role of NK cells in GATA2-haploinsufficiency may have accentuated the patients' susceptibility to HLH. Expansion of a double negative T-lymphocytic population identified with CyTOF could be a further factor contributing to HLH in these patients. This is the first report of VZV-triggered HLH-like disease in a primary immunodeficiency and the third report of HLH in GATA2-haploinsufficiency. Since HLH was part of the presentation in one of our patients, GATA2-haploinsufficiency represents a potential differential diagnosis in patients presenting with the clinical features of HLH—especially in cases of persisting cytopenia after recovery from HLH

Genetic landscape of congenital insensitivity to pain and hereditary sensory and autonomic neuropathies

Congenital insensitivity to pain (CIP) and hereditary sensory and autonomic neuropathies (HSAN) are clinically and genetically heterogeneous disorders exclusively or predominantly affecting the sensory and autonomic neurons. Due to the rarity of the diseases and findings based mainly on single case reports or small case series, knowledge about these disorders is limited. Here, we describe the molecular workup of a large international cohort of CIP/HSAN patients including patients from normally under-represented countries. We identify 80 previously unreported pathogenic or likely pathogenic variants in a total of 73 families in the >20 known CIP/HSAN-associated genes. The data expand the spectrum of disease-relevant alterations in CIP/HSAN, including novel variants in previously rarely recognized entities such as ATL3-, FLVCR1- and NGF-associated neuropathies and previously under-recognized mutation types such as larger deletions. In silico predictions, heterologous expression studies, segregation analyses and metabolic tests helped to overcome limitations of current variant classification schemes that often fail to categorize a variant as disease-related or benign. The study sheds light on the genetic causes and disease-relevant changes within individual genes in CIP/HSAN. This is becoming increasingly important with emerging clinical trials investigating subtype or gene-specific treatment strategies

Formation of Trans-Activation Competent HIV-1 Rev:RRE Complexes Requires the Recruitment of Multiple Protein Activation Domains

The HIV-1 Rev trans-activator is a nucleocytoplasmic shuttle protein that is essential for virus replication. Rev directly binds to unspliced and incompletely spliced viral RNA via the cis-acting Rev Response Element (RRE) sequence. Subsequently, Rev oligomerizes cooperatively and interacts with the cellular nuclear export receptor CRM1. In addition to mediating nuclear RNA export, Rev also affects the stability, translation and packaging of Rev-bound viral transcripts. Although it is established that Rev function requires the multimeric assembly of Rev molecules on the RRE, relatively little is known about how many Rev monomers are sufficient to form a trans-activation competent Rev:RRE complex, or which specific activity of Rev is affected by its oligomerization. We here analyzed by functional studies how homooligomer formation of Rev affects the trans-activation capacity of this essential HIV-1 regulatory protein. In a gain-of-function approach, we fused various heterologous dimerization domains to an otherwise oligomerization-defective Rev mutant and were able to demonstrate that oligomerization of Rev is not required per se for the nuclear export of this viral trans-activator. In contrast, however, the formation of Rev oligomers on the RRE is a precondition to trans-activation by directly affecting the nuclear export of Rev-regulated mRNA. Moreover, experimental evidence is provided showing that at least two protein activation domains are required for the formation of trans-activation competent Rev:RRE complexes. The presented data further refine the model of Rev trans-activation by directly demonstrating that Rev oligomerization on the RRE, thereby recruiting at least two protein activation domains, is required for nuclear export of unspliced and incompletely spliced viral RNA

Hematopoietic stem cell transplantation for adolescents and adults with inborn errors of immunity: an EBMT IEWP study.

peer reviewedAllogeneic hematopoietic stem cell transplantation (HSCT) is the gold standard curative therapy for infants and children with many inborn errors of immunity (IEI), but adolescents and adults with IEI are rarely referred for transplant. Lack of published HSCT outcome data outside small, single-center studies and perceived high risk of transplant-related mortality have delayed the adoption of HSCT for IEI patients presenting or developing significant organ damage later in life. This large retrospective, multicenter HSCT outcome study reports on 329 IEI patients (age range, 15-62.5 years at HSCT). Patients underwent first HSCT between 2000 and 2019. Primary endpoints were overall survival (OS) and event-free survival (EFS). We also evaluated the influence of IEI-subgroup and IEI-specific risk factors at HSCT, including infections, bronchiectasis, colitis, malignancy, inflammatory lung disease, splenectomy, hepatic dysfunction, and systemic immunosuppression. At a median follow-up of 44.3 months, the estimated OS at 1 and 5 years post-HSCT for all patients was 78% and 71%, and EFS was 65% and 62%, respectively, with low rates of severe acute (8%) or extensive chronic (7%) graft-versus-host disease. On univariate analysis, OS and EFS were inferior in patients with primary antibody deficiency, bronchiectasis, prior splenectomy, hepatic comorbidity, and higher hematopoietic cell transplant comorbidity index scores. On multivariable analysis, EFS was inferior in those with a higher number of IEI-associated complications. Neither age nor donor had a significant effect on OS or EFS. We have identified age-independent risk factors for adverse outcome, providing much needed evidence to identify which patients are most likely to benefit from HSCT

MicroTCA.4 Usage in the Femtosecond-Synchronization System at European XFEL - Enabling the XFEL for Femtosecond Precision User Experiments

The optical synchronization system at the European XFEL serves as femtosecond-stable reference throughout the 3.5 km long facility. Its signal is essential for the pump-probe experiments, first by enabling the LLRF system to stabilize the the electron bunch arrival time and thereby the FEL X-ray pulse timing, and second by synchronizing the experimental laser to fs precision. Most of the electronic hardware is realized in various MTCA.4 AMC-, RTM-, and FMC-Modules, most of them specially designed for this aplication. Several digital control feedback loops are implemented on FPGAs for the laser synchronization, the fiber link stabilization, and the 32 different end-stations. This talk will give a short introduction on the system and explain the involved hardware and software components

Über den Einfluss einer Inhibition der Proteinkinase C(ε) auf die TRAIL-induzierte Apoptose in Lungenkarzinom- und Medulloblastomzellen

Lungenkrebs ist die weltweit häufigste Ursache eines krebsassoziierten Todes und nimmt weiter stetig in seiner Prävalenz zu. Das Versagen der Standard-Chemotherapie ist besonders im fortgeschrittenen Stadium für viele Todesfälle verantwortlich. Neue Therapiewege werden daher dringend benötigt. Auch die Behandlung von Medulloblastomen stellt sich gerade bei erkrankten Kindern unter 3 Jahren als schwierig dar, auch weil durch die oftmals sehr intensiven Therapieregime mit schwerwiegenden Spätfolgen gerechnet werden muss. Es muß dementsprechend ebenso für diese Erkrankung nach innovativen Behandlungsansätzen gesucht werden. TRAIL verkörpert solch einen neuen, vielversprechenden Ansatz, da es in der Lage ist selektiv in Tumorzellen eine Apoptose zu bewirken. In ersten klinischen Studien hat es bereits seine gute Verträglichkeit bewiesen. Jedoch sind in zahlreichen Tumoren Resistenzen gegenüber TRAIL gefunden worden, welche die therapeutische Nutzung einschränken könnten. Die Proteinkinase C-Familie wurde schon mehrfach mit einer TRAIL-Resistenz in Verbindung gebracht. In dieser Arbeit wurde daher die TRAIL-Resistenz von A549 Lungenkarzinom- und DAOY Medulloblastomzellen in Hinblick auf ihre Beeinflussbarkeit durch die Proteinkinase C (PKC) untersucht. Sowohl in A549, als auch in DAOY Zellen, die zuvor noch nicht in diesem Zusammenhang untersucht wurden, konnte durch eine Inhibition der PKC die TRAIL-Resistenz durchbrochen werden. NPC 15437, ein PKC-Inhibitor, der vor allem die neuen Isoformen der PKC hemmt, verstärkte sehr deutlich den TRAIL-induzierten Zelltod und den Verlust des mitochondrialen Membranpotentials in beiden Zelllinien, ohne selbst wesentlich eine Apoptose auszulösen. Durch den pan-Caspase-Inhibitor z-VAD-fmk war dieser Effekt komplett unterdrückbar. Anschließend wurde versucht die verantwortliche PKC-Isoform zu bestimmen. Die Vorbehandlung mit Phorbol-12-myristat-13-acetat (PMA), Phorbol-12,13-didecanoat (PDD), sowie 12-Deoxyphorbol-13-phenylacetat-20-acetat (dPPA) verminderte den TRAIL-sensibilisierenden Effekt von NPC 15437 signifikant um etwa die Hälfte, während die Vorbehandlung mit Phorbol-13-monoacetat (PA) und Phorbol-12-monomyristat (PMM) keine Wirkung zeigte. Gö6976 und der membrangängige PKCα/β-spezifische Peptidinhibitor myr-ψ-PKC waren zudem nicht in der Lage eine Sensibilisierung für TRAIL zu bewirken. Es ergab sich daraus die Vermutung, dass PKCε eine herausragende Funktion in der TRAIL-Resistenz der Zelllinien ausübt. Der Einsatz von gegen PKCε gerichteter siRNA bestätigte dies und der membrangängige Peptidinhibitor myr-PKCε-V1-2 erfüllte von allen getesteten Substanzen am besten die Funktion eines TRAIL-Wirkungsverstärkers. Außerdem wurde durch Western Blot-Analysen eine mögliche Rolle von cFLIP/CFLAR für die TRAIL-Resistenz von A549 und DAOY Zellen weitestgehend ausgeschlossen. Die vorliegende Arbeit stellt die Proteinkinase C(ε) als einen wichtigen Resistenzfaktor der TRAIL-induzierten Apoptose heraus und zeigt wie deren Hemmung die zukünftige Krebstherapie verbessern könnte.Resistance to tumour necrosis factor apoptosis inducing ligand (TRAIL) is worthy to be studied, since TRAIL still attracts attention as a possible future anti-cancer agent that spares normal cells. Influence of protein kinase C on TRAIL resistance is shown, but there is confusion about the responsible isoforms. In the present study we broke resistance to TRAIL in A549 lung adenocarcinoma and DAOY medulloblastoma cells by using NPC 15437, a chemical PKC inhibitor. Sensitization to TRAIL was caspase-dependent and was accompanied by substantial loss in mitochondrial membrane potential. GÖ 6976 and myr-ψPKC, a cell-permeable peptide specific for cPKCs, were not capable of sensitizing cells to TRAIL. These findings excluded classical isoforms from involvement in conferring resistance to TRAIL. By pretreatment with distinct phorbolesters we could also exclude δ and η isoforms and we therefore concluded PKC ε to be responsible for TRAIL resistance in A549 cells. We could prove the importance of PKC ε in A549 cells by largely augmenting TRAIL-response using pre-treatment with Myr-PKCε V1-2, a specific cell-permeable inhibitory peptide. Empolying PKCε-specific siRNA we could confirm the special role of this isoform in TRAIL-resistance in A549 and DAOY cells. Immunoblot analysis showed that FLIPS/L does not play a significant role in resistance to TRAIL. Our findings may help to clarify TRAIL signaling or improve a possible therapeutic use

Digitalisierung in der Sozialwirtschaft Innovative Klientelinformation und -kommunikation mit Hilfe von Smartphone Apps

Smartphone Apps boomen und viele Unternehmen in der Wirtschaftswelt benutzen Apps um in den Kontakt mit ihrer Kundschaft zu kommen oder um diese zu informieren. Die vorliegende Arbeit beschäftigt sich mit der Verwendung von Smartphone Apps in der Sozialwirtschaft für die Klientelkommunikation und -information. Dabei wird untersucht, inwiefern sozialwirtschaftliche Organisationen bereits Smartphone Apps einsetzen und welche Erfahrungen diese Organisationen mit den Apps gemacht haben. Es zeigte sich, dass Apps am meisten in der Kinder- und Jugendhilfe eingesetzt werden um die Klientel über aktuelle Angebote zu informieren. Ebenfalls werden Apps zur Beratung von Flüchtlingen und als Hilfsmittel von Organisationen in der Schuldenberatung (Budgetplanung) oder in der Suchthilfe (Konsumtagebuch) eingesetzt. Die Untersuchung über die Erfahrungen mit Apps ergab, dass diese bei der Zielgruppe beliebt sind, die Entwicklung und die Pflege der Apps jedoch hohe Kosten verursachen.Smartphone Apps have been on the rise in the economic world. Although it is an efficient tool to reach the target group of a wide range of social organizations, the sector is still reticent. The aim of this thesis is to analyse the use of smartphone apps in social organizations to inform and interact with the target group and to explore what experiences social organizations have with this tool. This qualitative study starts with a market analysis of the existing smartphone apps in social organizations in the German-speaking regions. Guided interviews were carried out to determine the experiences of organizations, which are already launched apps. This thesis showed that apps are most widely used in the youth welfare to inform the clients about upcoming activities. Also, apps are used to inform refugees or as a tool for the clients in the debt counselling (budget plan) and addiction care (consumption log-books). The experiences showed that apps are popular with the target group, but apps are cost-intensive to development and maintenance.vorgelegt von: Nicolas Matthias FelberAbweichender Titel laut Übersetzung der Verfasserin/des VerfassersWien, FH Campus Wien, Masterarb., 201