T-Zell-Dysfunktion im Pankreaskarzinommodell: Rolle der IL-18- und IL-1- Rezeptor-Signaltransduktion bei der Induktion intratumoraler T-Zell-Dysfunktion

Das Pankreaskarzinom zeichnet sich durch geringe Immunogenität und geringe Antigenität aus. Aus diesem Grund haben sich Therapieansätze auf Basis immuntherapeutischer Strategien bisher nicht im klinischen Behandlungsalgorithmus durchsetzen können. Zwar ist die Infiltration mit zytotoxischen CD8+-T-Zellen (CTLs) bei Pankreaskarzinompatienten prognostisch günstig, allerdings zeigen intratumorale CTLs reduzierte zytotoxische Effektorfunktionen sowie gesteigerte Expression koinhibitorischen Rezeptoren, wie PD-1 und TIM-3. Aufgrund dieser T-Zell- Dysfunktion sind die CTLs nicht in der Lage Pankreaskarzinomzellen adäquat zu bekämpfen. Die NLRP3-vermittelten Zytokine IL-1β und IL-18 können auf die Plastizität der CTLs einwirken und die intratumorale Dysfunktion dieser Zellen steigern. Diese Arbeit untersucht die Rolle der NLRP3-vermittelten IL-1-Rezeptor (IL-1R)- und IL-18- Rezeptor (IL-18R)-Signaltransduktion für Plastizität und Effektorfunktion von zytotoxischen T-Zellen während der Immunantwort in einem murinen Pankreaskarzinommodell. Das Ziel dieser Arbeit ist es a) den Einfluss von IL-1β und IL-18 auf Differenzierung und zytotoxische Effektorfunktion von CTLs zu untersuchen, b) die Rolle der IL-1R- und IL- 18R-Signaltransduktion in intratumoralen CTLs und ihre Auswirkung auf deren Dysfunktion zu analysieren, c) den Einfluss von IL1β und IL-18 auf Tumorzellen zu untersuchen sowie d) zu zeigen wie intratumorale NLRP3-Expression die Immunantwort adoptiv transferierter und endogener CTLs beeinflusst. Diese Arbeit zeigt, dass IL-18R-Signaltranduktion, und in einem geringeren Maße die Signaltransduktion über IL-1R, einen immunsuppressiven Effekt auf intratumorale CTLs hat und die Entwicklung eines dysfunktionalen Zustands fördert. Die IL-18R- Signaltransduktion induziert den dysfunktionalen Zustand dabei über die Stimulation des IL-2-Rezeptors und dem damit verbundenen IL-2/Stat5-Signalweg. IL-18R- defiziente intratumorale T-Zellen zeigen hingegen im Vergleich zu intratumoralen Wildtyp (WT) T-Zellen eine verminderte Expression von Pdcd1, Havcr2, Lag3, Tigit, Eomes und Prdm1 mit einer gleichzeitig gesteigerten Expression von Tcf7 und Lef1, was auf eine erhöhte stemness-like memory-Funktion dieser Zellen hinweist. Des Weiteren wird gezeigt, dass die Signaltransduktion NLPR3-abgeleiteter Zytokine einerseits wichtig für die Aktivierung von CTLs ist, andererseits aber auch die Induktion eines dysfunktionalen Phänotyps befördert. Hierbei wirken pleiotrope Effekte auf die intratumoralen CTLs. Zusammenfassend liefert diese Arbeit mechanistische Einblicke in die Induktion intratumoraler T-Zell-Dysfunktion durch IL-1R- und IL-18R-Signaltransduktion und ist daher relevant für die Entwicklung

Lil3 assembles as chlorophyll-binding protein complex during deetiolation

AbstractDark-grown angiosperm seedlings are etiolated and devoid of chlorophyll. Deetiolation is triggered by light leading to chlorophyll dependent accumulation of the photosynthetic machinery. The transfer of chlorophyll to the chlorophyll-binding proteins is still unclear. We demonstrate here that upon illumination of dark-grown barley seedlings, two new pigment-binding protein complexes are de novo accumulated. Pigments bound to both complexes are identified as chlorophyll a and protochlorophyll a. By auto-fluorescence tracking and mass spectrometry, we show that exclusively Lil3 is the pigment-binding complex subunit in both complexes

The proteome of the heterocyst cell wall in Anabaena sp. PCC 7120

Anabaena sp. PCC 7120 is a filamentous cyanobacterium that serves as a model to analyze prokaryotic cell differentiation, evolutionary development of plastids, and the regulation of nitrogen fixation. The cell wall is the cellular structure in contact with the surrounding medium. To understand the dynamics of the cell wall proteome during cell differentiation, the cell wall from Anabaena heterocysts was enriched and analyzed. In line with the recently proposed continuity of the outer membrane along the Anabaena filament, most of the proteins identified in the heterocyst cell-wall fraction are also present in the cell wall of vegetative cells, even though the lipid content of both membranes is different

Impact of COVID-19 policy responses on live-in care workers in Austria, Germany, and Switzerland

Context: The measures taken to counter the COVID-19 pandemic restricted the circular migration of live-in care workers between their countries of origin and the elderly persons’ households. Objective: In this comparative policy analysis, the impact of COVID-19 related policy measures for transnationally organised live-in care in Austria, Germany, and Switzerland is investigated. Method: Policy measures and media debates were analysed and inquiries with care workers, representatives of care agencies, unions, and activist groups were carried out between March and June 2020. Findings: In accordance with their institutionalisation of live-in care, Austria, Germany, and Switzerland responded differently to the challenges the pandemic posed to live-in care arrangements. However, all three countries focused on extending care workers’ rotas and re-establishing transnational mobility. These priorities subordinated the interests of care workers to those of care recipients. Furthermore, the measures remained short-term solutions that failed to acknowledge the fundamental flaws and inequalities of a care model that relies primarily on female migrant workers and wage differentials within Europe. Limitations: This policy comparison is based on an in-depth analysis of COVID-19 related policies, supplemented by inquiries among stakeholders with whom research had been done prior to the pandemic. More in-depth interviews are required to further substantiate the findings concerning their perspectives and gain insight into the longer-term effects of the pandemic. Implications: The pandemic has brought the flaws of the live-in care model to the fore. Countries need to rethink their fragile care policies, which build on social inequality and uninhibited transnational mobility

Sheduling approach for Microfactories with setup times.

International audienceIn this paper we consider microfactories for manipulation and assembly. These microfactories are composed of several cells containing microrobotic systems capable of a high level of repeatability. The assembly plan of the production is a pipeline of tasks that are performed by the cells. Our aim is to manage the production flow in the case where the cells can be reconfigured to perform different task types. Each cell is in charge of several consecutive tasks. A setup time is necessary to switch from the processing of one task type to another, and multiple intermediate results may be stored temporarily in storage areas to avoid switching the task type after the processing of each product. In this context we assess the optimized use of these storage areas, called buffers, and its impact on the production throughput

Heterotrimeric G-protein subunit Gαi2 contributes to agonist-sensitive apoptosis and degranulation in murine platelets

Gαi2, a heterotrimeric G-protein subunit, regulates various cell functions including ion channel activity, cell differentiation, proliferation and apoptosis. Platelet-expressed Gαi2 is decisive for the extent of tissue injury following ischemia/reperfusion. However, it is not known whether Gαi2 plays a role in the regulation of platelet apoptosis, which is characterized by caspase activation, cell shrinkage and cell membrane scrambling with phosphatidylserine (PS) translocation to the platelet surface. Stimulators of platelet apoptosis include thrombin and collagen-related peptide (CoRP), which are further known to enhance degranulation and activation of αII bβ3-integrin and caspases. Using FACS analysis, we examined the impact of agonist treatment on activation and apoptosis in platelets drawn from mice lacking Gαi2 and their wild-type (WT) littermates. As a result, treatment with either thrombin (0.01 U/mL) or CoRP (2 μg/mL or 5 μg/mL) significantly upregulated PS-exposure and significantly decreased forward scatter, reflecting cell size, in both genotypes. Exposure to CoRP triggered a significant increase in active caspase 3, ceramide formation, surface P-selectin, and αII bβ3-integrin activation. These molecular alterations were significantly less pronounced in Gαi2-deficient platelets as compared to WT platelets. In conclusion, our data highlight a previously unreported role of Gαi2 signaling in governing platelet activation and apoptosis.Fil: Cao, Hang. Universität Tübingen; AlemaniaFil: Qadri, Syed M.. Canadian Blood Services; Canadá. McMaster University; CanadáFil: Lang, Elisabeth. Heinrich-heine-universität Düsseldorf; AlemaniaFil: Pelzl, Lisann. Universität Tübingen; AlemaniaFil: Umbach, Anja T.. Universität Tübingen; AlemaniaFil: Leiss, Veronika. Universität Tübingen; AlemaniaFil: Birnbaumer, Lutz. National Institutes of Health; Estados Unidos. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Instituto de Investigaciones Biomédicas. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas; ArgentinaFil: Nürnberg, Bernd. Universität Tübingen; AlemaniaFil: Pieske, Burkert. Berlin Institute of Health; Alemania. Universitätsmedizin Berlin; AlemaniaFil: Voelkl, Jakob. Berlin Institute of Health; Alemania. Universitätsmedizin Berlin; AlemaniaFil: Gawaz, Meinrad. Universität Tübingen; AlemaniaFil: Bissinger, Rosi. Universität Tübingen; AlemaniaFil: Lang, Florian. Universität Tübingen; Alemania. Heinrich-heine-universität Düsseldorf; Alemani

Detection of Microorganisms Onboard the International Space Station Using an Electronic Nose

Abstract We report on the detection of microorganisms onboard the International Space Station (ISS) using an electronic nose we named the E-Nose. The E-Nose, containing an array of ten different metal oxide gas sensors, was trained on Earth to detect the four most abundant microorganisms that are known to exist onboard the ISS. To assess its performance in space, the E-Nose was brought to the ISS and three measurement campaigns were carried out in three different locations inside the ISS during a 5-month mission. At the end of this mission, all investigated locations were wiped with swabs, and the swabs and odor sensor signal data were sent back to Earth for an in-depth analysis in earthbound laboratories. The in-space measurements were compared with an odor database containing four organisms, but a consensus odor could not be identified. Microbiological results could not provide clues to the smell that was measured. The yeast Rhodotorula mucilaginosa was identified in the literature as the most probable candidate for the unknown odor. Further investigations showed that the smell of Rhodotorula mucilaginosa matches very well with the data obtained inside the ISS. Finally, Rhodotorula mucilaginosa DNA was identified in swabs taken from the sleeping cabin of the astronaut, which confirms the assumption that the yeast Rhodotorula mucilaginosa was actually measured in space by the E-Nose

The endocannabinoid, anandamide, induces cannabinoid receptor-independent cell death in renal proximal tubule cells

Background: The endocannabinoid (EC) system is well characterized in the central nervous system but scarcely studied in peripheral organs. In this paper, we newly identify the effect of the EC anandamide (AEA) upon renal proximal tubule cells. Methods: Measurement of lactate dehydrogenase (LDH) release after treatment of primary renal proximal tubule cells (RPTEC) and renal carcinoma cell line (Caki-1) with AEA, arachidonic acid (AA), ethanolamide (EtAm), EC receptor CB1 antagonist (AM251), CB2 receptor antagonist (SR144528), TRPV1 receptor antagonist (capsazepine), degradation enzyme fatty acid amide hydrolase (FAAH) antagonist (URB597), antioxidants GSH-EE; Trolox, GSH depletor BSO, membrane cholesterol depletor (MCD), apoptosis inhibitor zVAD, necroptosis inhibitor Nec-1 or ferroptosis inhibitor Fer-1. Western blot and qRT-PCR analysis plus determination of reactive oxygen species (ROS) via H2-DCFDA were performed. Histology for EC enzymes, N-acetylphosphatidylethanolamine hydrolyzing phospholipase D (NAPE-PLD) and FAAH, as well as the determination of physiological levels of ECs in human and rat renal tissue via liquid chromatography were conducted. Results: AEA both dose- and time-dependently induces cell death in RPTEC and Caki-1 within hours, characterized by cell blebbing, not influenced by blocking the described EC receptors by AM251, SR144528, capsaze pine or FAAH by URB597 or MCD. Cell death is mediated via ROS. There is no difference found in the histology of the enzymes FAAH and NAPE-PLD in human renal tissue with interstitial nephritis. Blocking of apoptotic, necroptotic or ferroptotic cell death does not lead to a reduction in LDH release in vitro. Conclusion: The endocannabinoid anandamide induces cell death in renal proximal tubule cell in a time- and dose-dependent manner. This pathway is mediated via ROS and is independent of cannabinoid receptors, membrane cholesterol or FAAH activity

Biotransformation of nitriles to amides using soluble and immobilized nitrile hydratase from Rhodococcus erythropolis A4

A semi-purified nitrile hydratase from Rhodococcus erythropolis A4 was applied to biotransformations of 3-oxonitriles 1a–4a, 3-hydroxy-2-methylenenitriles 5a–7a, 4-hydroxy-2-methylenenitriles 8a–9a, 3-hydroxynitriles 10a–12a and 3-acyloxynitrile 13a into amides 1b–13b. Cross-linked enzyme aggregates (CLEAs) with nitrile hydratase and amidase activities (88% and 77% of the initial activities, respectively) were prepared from cell-free extract of this microorganism and used for nitrile hydration in presence of ammonium sulfate, which selectively inhibited amidase activity. The genes nha1 and nha2 coding for α and β subunits of nitrile hydratase were cloned and sequenced