Determinants of HCV-specific CD8+ T cell expansion

Infektionen mit dem Hepatitis C Virus (HCV) sind global ein enormes Problem. Verschiedene Studien belegen, dass während einer akuten HCV-Infektion CD8+ T Zellen eine wichtige Rolle bei der Kontrolle der Virusinfektion spielen. Allerdings sind HCV-spezifische T Zellantworten oft schwach und dysfunktional. Die Aktivierung virusspezifischer T Zellen ist ein sensitiver, komplexer Prozess, bei dem sowohl die Sequenz des präsentierten Antigens, als auch das Umfeld in dem der primäre Kontakt zwischen antigenpräsentierender und virusspezifischer Zelle stattfindet, Einfluss auf die resultierende T Zellantwort ausübt. Daher wurden in der vorliegenden Studie Faktoren der HCV-spezifischen T Zellantwort untersucht. Das Hepatitis C Virus zeichnet sich durch ein hohes Maß an genetischer Variabilität aus. Es werden sieben verschiedene Genotypen und multiple Subtypen unterschieden, aber selbst zwischen Varianten des gleichen Subtyps gibt es wichtige Sequenzunterschiede. Diese hohe Sequenzvariabilität ist ein entscheidendes Hindernis in der Entwicklung eines geeigneten Impfstoffs gegen HCV. Allerdings wird selbst nach spontaner Ausheilung einer Infektion eine protektive Immunantwort häufig nicht erreicht, da sich immundominante Epitope zwischen verschiedenen HCV Isolaten unterscheiden und diese Sequenzvarianten häufig nicht von CD8+ T Zellen erkannt werden. Im ersten Teil dieser Arbeit wurde die globale Sequenzdiversität der Genotypen 1a und 1b in einem immundominanten HLA-A*02-assoziierten CD8+ T Zell-Epitop NS31406-1415 analysiert. Dabei wurde der Einfluss der Sequenzunterschiede auf die ‚Priming‘-Kapazität von CD8+ T Zellen sowie die Induktion von kreuzreaktiven CD8+ T Zellen untersucht. Es konnte gezeigt werden, dass sich die global zirkulierenden Varianten in ihrer ‚Priming‘-Kapazität von CD8+ T Zellen unterscheiden. Diese in vitro generierten T Zellantworten unterschieden sich maßgeblich in ihrer Fähigkeit der Kreuzreaktivität. Interessanterweise stammt die Sequenzvariante mit der breitesten Kreuzreaktivität aus Isolaten, die hauptsächlich in Japan zirkulieren und somit nur selten in Europa zu finden sind. Ein Impfstoff gegen HCV sollte eine polyfunktionelle CD8+ T Zellantwort hervorrufen und ebenso kreuzreaktiv mit Sequenzvarianten des Epitops sein. In dieser Studie konnte die entscheidende Bedeutung von Sequenz-variationen in einem Antigen für die folgende CD8+ T Zellantwort gezeigt werden. Die Daten unterstreichen, wie wichtig die Auswahl der optimalen Epitopsequenz für eine Vakzine gegen HCV ist, indem eine polyfunktionelle sowie auch kreuzreaktive CD8+ T Zellantwort aktiviert wird. Im Falle einer Infektion mit dem Hepatitis C Virus findet der erste Kontakt zwischen antigenpräsentierender und virusspezifischer Zelle in der Leber statt. Die Leber spielt eine wichtige Rolle im adaptiven Immunsystem, da hier die Präsentation von Antigenen an CD8+ T Zellen Toleranz induzieren kann, indem dysfunktionale CD8+ T Zellen induziert werden oder deren Aktivierung in der Leber zu früher Apoptose führt. Ein Kennzeichen chronischer Infektion mit hepatotropen Viren ist somit die schwache Immunantwort durch CD8+ T Zellen. Jüngere Studien an Mausmodellen konnten belegen, dass Hepatozyten ebenfalls als antigenpräsentierende Zellen fungieren und demnach möglicherweise eine Rolle in der Toleranzinduktion von CD8+ T Zellen einnehmen. Im zweiten Teil der vorliegenden Arbeit wurde gezeigt, dass die Expansion virus-spezifischer CD8+ T Zellen in der Anwesenheit von primären humanen Hepatozyten drastisch reduziert ist. Die Hemmung der virusspezifischen CD8+ T Zellexpansion war unabhängig vom antigenpräsentierenden Zelltyp und war unabhängig vom antigenspezifischen Kontakt. Die Hemmung der virusspezifischen CD8+ T Zell Expansion war allerdings aufgehoben, wenn direkter Kontakt zwischen Hepatozyten und PBMC durch den Gebrauch eines Membraneinsatzes in der Kultur verhindert wurde. Dies deutet an, dass der hemmende Effekt von Hepatozyten kontaktabhängig ist. Eine Analyse der Zellkulturüberstände erwies eine erhöhte Konzentration von TGF-β in der Anwesenheit von Hepatozyten. In der vorliegenden Studie wurde demnach gezeigt, dass eine Hemmung der CD8+ T Zellantwort durch Hepatozyten möglicherweise zur Chronifizierung von Infektionen mit hepatotropen Viren beitragen kann.Infection with the hepatitis C virus still remains a global health problem. The antiviral immune response against HCV by CD8+ T cells plays a central role in viral containment, however, it has been highlighted in numerous studies that the CD8+ T cell responses during HCV infection are relatively weak and dysfunctional. Activation of virus-specific CD8+ T cells is a multi-step process, in which the sequence of the presented antigen, as well as the site of antigen presentation has important consequences for the quality of the CD8+ T cell response. Hence, in the present thesis, determinants of HCV-specific CD8+ T cell expansion were analyzed. HCV is characterized by high levels of genetic variation between genotypes and subtypes but also between isolates of the same subtype. This high sequence variation in immunodominant epitopes precludes protective immunity and represents a major obstacle to vaccine design. In this study, we analyzed the global sequence diversity of the immunodominant HLA-A*02-restricted CD8+ T cell epitope NS31406-1415 in circulating genotype 1a and 1b isolates and its impact on CD8+ T cell priming and induction of cross-reactive T cell responses. Circulating sequence variants of the NS31406-1415 epitope showed distinct priming potential and cross-reactivity patterns. Importantly, the highest degree of cross-reactivity was induced by a sequence that is rarely observed in the European population. Upon vaccination, the elicited CD8+ T cells should be polyfunctional and target multiple epitope variants. This study highlights the relevance of antigen sequence variation for subsequent CD8+ T cell responses and provides a rational for the selection of the optimal epitope sequence for a vaccine against HCV, associated with the induction of a broad cross-reactive and polyfunctional CD8+ T cell response. A hallmark of chronic infections with hepatotropic viruses such as HCV is the notoriously weak antiviral immune response by CD8+ T cells. Primary antigen encounter in HCV infections happens in the liver. It is believed that the liver-specific environment can induce tolerance either by induction of dysfunctional CD8+ T cells or by premature death of activated CD8+ T cells. Recent reports from mouse models indicate that hepatocytes can act as antigen-presenting. Direct antigen presentation by virally infected hepatocytes thus might inhibit the CD8+ T cell response. This study showed that the frequency of virus-specific CD8+ T cells after 10 days expansion was dramatically reduced in the presence of hepatocytes. This suppressive effect was independent of the cell-type that was used for antigen-presentation (hepatocyte or PBMC) and of antigen-specific contact. Expansion of CD8+ T cells in the presence of hepatocytes was almost completely restored in transwell experiments indicating that the suppressive effect is cell-contact dependent. Analysis of the cytokine profile from culture supernatants revealed increased concentrations of TGF-β in the presence of hepatocytes compared to their absence. This study gives important hints for the establishment of chronic infections by hepatotropic viruses

Systematic comparison and reconstruction of sea urchin (Echinoidea) internal anatomy: a novel approach using magnetic resonance imaging

<p>Abstract</p> <p>Background</p> <p>Traditional comparative morphological analyses and subsequent three-dimensional reconstructions suffer from a number of drawbacks. This is particularly evident in the case of soft tissue studies that are technically demanding, time-consuming, and often prone to produce artefacts. These problems can partly be overcome by employing non-invasive, destruction-free imaging techniques, in particular micro-computed tomography or magnetic resonance imaging.</p> <p>Results</p> <p>Here, we employed high-field magnetic resonance imaging techniques to gather numerous data from members of a major marine invertebrate taxon, the sea urchins (Echinoidea). For this model study, 13 of the 14 currently recognized high-ranking subtaxa (orders) of this group of animals were analyzed. Based on the acquired datasets, interactive three-dimensional models were assembled. Our analyses reveal that selected soft tissue characters can even be used for phylogenetic inferences in sea urchins, as exemplified by differences in the size and shape of the gastric caecum found in the Irregularia.</p> <p>Conclusion</p> <p>The main focus of our investigation was to explore the possibility to systematically visualize the internal anatomy of echinoids obtained from various museum collections. We show that, in contrast to classical preparative procedures, magnetic resonance imaging can give rapid, destruction-free access to morphological data from numerous specimens, thus extending the range of techniques available for comparative studies of invertebrate morphology.</p

Historical sources of ethnomusicology in contemporary debate

This anthology concerns traditional music and archives, and discusses their relationship as seen from historical and epistemological perspectives. Music recordings on wax cylinders, 78 records or magnetic tape, made in the first half of the 20th century, are regarded today as valuable sources for understanding musical processes in their social dimension and as unique cultural heritage. Most of these historical sound recordings are preserved in sound archives, now increasingly accessible in digital formats. Written by renowned experts, the articles here focus on archives, individual and collective memory, and heritage as today’s recreation of the past. Contributors discuss the role of historical sources of traditional music in contemporary research based on examples from music cultures in West Africa, Scandinavia, Turkey, and Portugal, among others. The book will appeal to musicologists and cultural anthropologists, as well as historians and sociologists, and will be of interest to anyone concerned with sound archives, libraries, universities and cultural institutions dedicated to traditional music

Vergleichende Differenzierung von Clostridium spp. mittels biochemischer und molekularbiologischer Methoden sowie MALDI-TOF Massenspektrometrie

The aim of this study was to examine whether the classical biochemical differentiation methods in relation to clostridial analysis can be replaced by newer molecular biological techniques such as real-time PCR and by MALDI-TOF MS. For this purpose, 118 potentially pathogenic Clostridium spp. isolates were compared at the Bavarian Health and Food Safety Authority using the above mentioned methods. On the basis of the obtained results a recommendation concerning the general approach to the detection of Clostridium spp. in the food industry has been elaborated

Analysis of EEC Regulation 2092/91 in relation to other national and international organic standards

This Deliverable 3.2 report presents an analysis of differences between EEC Regulation 2092/91 and other organic standards and their implementation, using a specially developed database (www.organicrules.org). It further reports on database development. The work was carried out as part of the “EEC 2092/91 (Organic) Revision” STREP project (No. SSPE-CT- 2004-502397) within the EU 6th Framework Programme. The main objective was to identify differences in organic standards in relation to Regulation (EEC) 2092/91 and to analyse selected national governmental and private organic standards with the aim of identifying specific areas in the (EEC) 2092/91 where revision in terms of harmonisation, regionalisation or simplification may be possible

Rock Mechanical Laboratory Testing of Thebes Limestone Formation (Member I), Valley of the Kings, Luxor, Egypt

The Thebes Limestone Formation of Lower Eocene age is one of the most extensive rock units in Egypt. It is of importance to the apogee of the ancient Egyptian civilization, particularly in Luxor (South-Central Egypt), where the rock formation hosts the Theban Necropolis, a group of funerary chambers and temples from the New Kingdom Egyptian era (3500-3000 BP). In this work, we investigated the petrophysical and rock mechanical properties (e.g., rock strength, critical crack stress thresholds) through laboratory tests on eleven rock blocks collected from one area within the Theban Necropolis known as the Valley of the Kings (KV). The blocks belong to Member I of the Thebes Limestone Formation, including six blocks of marly limestone, three blocks of micritic limestone, one block of argillaceous limestone from the Upper Esna Shale Formation, and one block of silicified limestone of unknown origin. Special attention was given to the orientation of bedding planes in the samples: tests were conducted in parallel (PA) and perpendicular (PE) configurations with respect to bedding planes. We found that the marly limestone had an average unconfined compressive strength (UCS) of 30 MPa and 39 MPa for the PA and PE tests, respectively. Similarly, the micritic limestone tests showed an average UCS of 24 MPa for the PA orientation and 58 MPa for the PE orientation. The critical crack thresholds were the first ever reported for Member I, as measured with strain gauge readings. The average crack initiation (CI) stress thresholds for the marly limestone (PA: 14 MPa) and the micritic limestone (PA: 11 MPa; PE: 24 MPa) fall within the typical ratio of CI to UCS (0.36-0.52). The micritic limestone had an average Young's modulus (E) of 19.5 GPa and 10.3 GPa for PA and PE, respectively. The Poisson's ratios were 0.2 for PA and 0.1 for PE on average. Both marly and micritic limestone can be characterised by a transverse isotropic strength behaviour with respect to bedding planes. The failure strength for intact anisotropic rocks depends on the orientation of the applied force, which must be considered when assessing the stability of tombs and cliffs in the KV and will be used to understand and improve the preservation of this UNESCO World Heritage site

Digital three-dimensional imaging techniques provide new analytical pathways for malacological research

Author Posting. © BioOne Complete, 2019. This article is posted here by permission of BioOne Complete for personal use, not for redistribution. The definitive version was published in Ziegler, A., Bock, C., Ketten, D. R., Mair, R. W., Mueller, S., Nagelmann, N., Pracht, E. D., & Schroeder, L. Digital three-dimensional imaging techniques provide new analytical pathways for malacological research. American Malacological Bulletin, 36(2), (2018):248-273, doi:10.4003/006.036.0205.Research on molluscan specimens is increasingly being carried out using high-throughput molecular techniques. Due to their efficiency, these technologies have effectively resulted in a strong bias towards genotypic analyses. Therefore, the future large-scale correlation of such data with the phenotype will require a significant increase in the output of morphological studies. Three-dimensional (3D) scanning techniques such as magnetic resonance imaging (MRI) or computed tomography (CT) can achieve this goal as they permit rapidly obtaining digital data non-destructively or even entirely non-invasively from living, fixed, and fossil samples. With a large number of species and a relatively complex morphology, the Mollusca would profit from a more widespread application of digital 3D imaging techniques. In order to provide an overview of the capacity of various MRI and CT techniques to visualize internal and external structures of molluscs, more than twenty specimens ranging in size from a few millimeters to well over one meter were scanned in vivo as well as ex vivo. The results show that all major molluscan organ systems can be successfully visualized using both MRI and CT. The choice of a suitable imaging technique depends primarily on the specimen's life condition, its size, the required resolution, and possible invasiveness of the approach. Apart from visual examples derived from more than two dozen scans, the present article provides guidelines and best practices for digital 3D imaging of a broad range of molluscan taxa. Furthermore, a comprehensive overview of studies that previously have employed MRI or CT techniques in malacological research is given.We would like to express our gratitude to Adam J. Baldinger, Thomas Bartolomaeus, Patrick Beckers, Rüdiger Bieler, Roger T. Hanlon, Carsten Lüter, Iliana Ruiz-Cooley, Tom Schiøtte, Andreas Schmidt-Rhaesa, and Sid Staubach for help with specimen collection or for providing access to museum material. Cornelius Faber, Julia Koch, Tony Stöcker, and W. Caroline West kindly facilitated use of scanning systems. We would also like to thank Julie Arruda, Scott Cramer, Jörg Döpfert, Charlotte Eymann, Bastian Maus, Malte Ogurreck, Christina L. Sagorny, Gillian Trombke, and Christopher Witte for support with data acquisition and analysis. We are particularly grateful to Elizabeth K. Shea for inviting the present contribution and for her extensive commentary on the manuscript. We also thank two anonymous reviewers for their helpful criticisms. Funding for this study was provided by the Ocean Life Institute, the Office of Naval Research, the Seaver Institute, and the Deutsche Forschungsgemeinschaft (INST 217/849-1 FUGG)

Forschungsbericht zum Thema "Arbeitnehmerüberlassung": Endbericht zum 29. Mai 2009

"Das Institut für Arbeitsmarkt- und Berufsforschung der Bundesagentur für Arbeit (IAB) hat für das Bundesministerium für Arbeit und Soziales ein Forschungsvorhaben zum Thema 'Arbeitnehmerüberlassung' durchgeführt. Anlass für das Forschungsvorhaben war der Auftrag des Bundeskabinetts aus der Kabinettklausur in Meseberg im Jahr 2007 an den damaligen Bundesarbeitsminister, die Entwicklung in der Zeitarbeit zu analysieren und zu prüfen. Die Ergebnisse des Forschungsvorhabens machen deutlich, dass der Einsatz von Zeitarbeitnehmerinnen und Zeitarbeitnehmern ein Weg der Flexibilisierung des Personaleinsatzes ist, der in den vergangenen Jahren an Bedeutung gewonnen hat. Sie zeigen auch, dass die Mehrheit der Zeitarbeitnehmerinnen und Zeitarbeitnehmer unmittelbar vor der Tätigkeit in der Zeitarbeit ohne Beschäftigung war. Darüber hinaus liefert das Forschungsvorhaben keine Anhaltspunkte für eine systematische Verdrängung von Stammbelegschaften durch Zeitarbeitnehmerinnen und Zeitarbeitnehmer. Die Ergebnisse des Forschungsvorhabens sind in den Elften Bericht der Bundesregierung über Erfahrungen bei der Anwendung des Arbeitnehmerüberlassungsgesetzes eingeflossen." (Autorenreferat, IAB-Doku