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Institute
- Graduate School of Life Sciences (764)
- Theodor-Boveri-Institut für Biowissenschaften (475)
- Physikalisches Institut (206)
- Institut für Informatik (139)
- Institut für Theoretische Physik und Astrophysik (121)
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- Julius-von-Sachs-Institut für Biowissenschaften (88)
Schriftenreihe
Sonstige beteiligte Institutionen
- Fraunhofer-Institut für Silicatforschung ISC (5)
- Helmholtz Institute for RNA-based Infection Research (HIRI) (5)
- Technische Hochschule Nürnberg Georg Simon Ohm (3)
- Deutsches Zentrum für Luft- und Raumfahrt (DLR), Institut für Raumfahrtsysteme (2)
- EMBL Heidelberg (2)
- Institut für Tierökologie und Tropenbiologie (2)
- Lehrkrankenhaus II. Medizinische Klinik Klinikum Coburg (2)
- Rudolf Virchow Center for Integrative and Translational Bioimaging, University of Würzburg (2)
- Universität Belgrad, Serbien (2)
- Universitätsklinikum Münster (2)
- Universitätsklinikum Würzburg (2)
- Zentrum für Infektionsforschung (ZINF) Würzburg (2)
- Zentrum für Telematik e.V. (2)
- ACC GmbH Analytical Clinical Concepts (1)
- ATLAS Collaboration (1)
- Adam Opel AG (1)
- Airbus Defence and Space GmbH (1)
- Anthropology Department University of Tennessee, Knoxville (1)
- BMBF (1)
- Bio-Imaging Center Würzburg (1)
- Biomedical Center Munich, Department of Physiological Chemistry, Ludwig-Maximilians-Universität München (1)
- Biomedizinische NMR Forschungs GmbH am Max-Planck-Institut fuer biophysikalische Chemie (1)
- Boehringer Ingelheim Pharma GmbH & Co. KG (1)
- Bundesinstitut für Arzneimittel und Medizinprodukte (1)
- CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - the development agency of the Brazilian Federal Government (1)
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- Carl-Ludwig-Institut für Physiologie, Universität Leipzig (1)
- Center for Nanosystems Chemistry (1)
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- Department of Biochemistry (1)
- Department of Cellular Therapies, University of Navarra, Pamplona, Spain (1)
- Department of Mathematical Analysis, Faculty of Mathematics and Physics, Charles University in Prague (1)
- Department of Veterinary Sciences, Experimental Parasitology, Ludwig-Maximilians-Universität München (1)
- Department of X-ray Microscopy, University of Würzburg, Würzburg, Germany (1)
- Deutscher Akademischer Austauschdienst (DAAD) (1)
- Deutsches Krebsforschungszentrum Heidelberg (1)
- Deutsches Zentrum für Luft & Raumfahrt (DLR) (1)
- Deutsches Zentrum für Luft- und Raumfahrt (1)
- Deutsches Zentrum für Luft- und Raumfahrt (DLR) (1)
- Deutsches Zentrum für Luft- und Raumfahrt e.V. (1)
- EMBL Mouse Biology Unit, Monterotondo, Italien (1)
- ESPCI Paris (1)
- European Molecular Biology Laboratory, Heidelberg, Germany (1)
- European Space Agency (1)
- Evangelisches Studienwerk e.V. (1)
- Experimental Physics V, University of Wuerzburg (1)
- Expertenkreis zur Erarbeitung eines Stufenplans zur Diagnose und Therapie von Angsterkrankungen (1)
- Fachgebiet für Populationsgenomik bei Nutztieren, Universität Hohenheim (1)
- Forschungsverbund ForChange des Bayrischen Kultusministeriums (1)
- Fraunhofer ISC (1)
- Fraunhofer Insitut für Silicatforschung ISC (1)
- Fraunhofer Institute for Silicate Research ISC in Würzburg (1)
- Fraunhofer-Institut für Chemische Technologie (ICT) (1)
- Fraunhofer-Institute for Silicate Research ISC (1)
- Friedrich-Schiller-Universität Jena (1)
- GEOMAR Helmholtz-Zentrum für Ozeanforschung Kiel (1)
- Harvard Medical School, Boston, USA (1)
- Helmholtz Center for RNA-based Infection Research (1)
- Helmholtz Institut für RNA-basierte Infektionsforschung (1)
- Helmholtz Institute for RNA-based Infection Research (1)
- Helmholtz-Institut für RNA-basierte Infektionsforschung (1)
- IBMP - Institut für Biomedizinische und Pharmazeutische Forschung in Nürnberg-Heroldsberg (1)
- Institut Ruđer Bošković, Zagreb, Croatia (1)
- Institut für Hygiene und Mikrobiologie (1)
- Institut für Medizinische Mikrobiologie und Hygiene der Eberhard-Karls-Universität Tübingen (1)
- Institut für Organische Chemie, RWTH Aachen (1)
- Institut für klinische Neurobiologie (1)
- Institute for Biochemistry I, University of Cologne (1)
- Institute of Transformative Bio-Molecules, Nagoya University, Nagoya, Japan (1)
- Instituto de Higiene, Universidad de la República, Montevideo, Uruguay (1)
- Instituto de Hygiene Montevideo, Uruguay (1)
- Interdisziplinäre Zentrum für Klinische Forschung (IZKF) (1)
- Interdisziplinäres Zentrum für Klinische Forschung (IZKF) Würzburg (1)
- Joslin Diabetes Center, Harvard Medical School (1)
- Julius-von-Sachs-Institut für Biowissenschaften Lehrstuhl für Botanik II - Ökophysiologie und Vegetationsökologie (1)
- Klinik für Psychiatrie und Psychotherapie, Universität Würzburg (1)
- Klinik und Poliklinik für Anästhesiologie, Intensivmedizin, Notfallmedizin und Schmerztherapie des Universitätsklinikums Würzburg (1)
- Klinische Mikrobiologie am Universitätsklinikum Erlangen (1)
- König-Ludwig-Haus, Orthopedic Clinic, Würzburg (1)
- LMU München (1)
- Lehrstuhl für Bioinformatik (1)
- Lehrstuhl für Fernerkundung der Uni Würzburg, in Kooperation mit dem Deutschen Fernerkundungsdatenzentrum (DFD) des Deutschen Zentrums für Luft- und Raumfahrt (DLR) (1)
- Lehrstuhl für Tierökologie und Tropenbiologie, Universität Würzburg (1)
- Leibniz-Institut für Analytische Wissenschaften - ISAS - e.V. (1)
- MRB Research Center for Magnetic-Resonance-Bavaria (1)
- Maastricht University, Department of Psychiatry and Neuropsychology (1)
- Maastricht University, Maastricht, the Netherlands (1)
- Mathematical Institute, Czech Academy of Sciences (1)
- Max Delbrück Center for Molecular Medicine (1)
- Max-Delbrück-Center für molekulare Medizin, Berlin (1)
- Medizinische Klinik und Poliklinik 1, Abteilung Kardiologie (1)
- Medizinische Klinik und Poliklinik 1, Abteilung Nephrologie (1)
- Medizinische Universität Innsbruck (1)
- Michigan State University (1)
- National Park "Bavarian Forest" (1)
- New York Blood Center (1)
- Novartis AG (1)
- Novartis Pharma AG, Switzerland (1)
- OTH Regensburg (1)
- Opel Automobile GmbH (1)
- Ostbayerische Technische Hochschule Regensburg (1)
- Paul-Scherrer-Institut, Villigen, CH (1)
- Pharmakologie, Universität Bonn (1)
- Pharmazie, Universität Mailand (1)
- Politecnico di Milano (1)
- Queensland University of Technology (1)
- Research Center for Infectious Diseases, University of Würzburg (1)
- Roche Diagnostics GmbH Penzberg (1)
- Rudolf-Virchow-Zentrum für Experimentelle Biomedizin der Universität Würzburg (1)
- Sahin Lab, F.M. Kirby Neurobiology Center Boston Children’s Hospital, Department of Neurology, Harvard Medical School (1)
- Siemens Corporate Technology Munich (1)
- Spanish National Center for Biotechnology (CNB-CSIC) (1)
- Spectral Service AG (1)
- Synchrotron SOLEIL, Gif-sur-Yvette (1)
- Tanzania Food and Drugs Authority (1)
- Technische Universität Darmstadt (1)
- Technische Universität Dresden (1)
- The project was founded by IZKF (1)
- Tropenmedizin, Klinikum Würzburg Mitte gGmbH, Missioklinik (1)
- University of Applied Sciences Aschaffenburg (1)
- University of Applied Sciences and Arts Western Switzerland, Fribourg (1)
- University of Birmingham, Medical School, Institute of Biomedical Research (1)
- University of Leipzig, Faculty of Life Sciences, Institute for Biology (1)
- University of Stellenbosch, Division of Medical Virology (1)
- University of Twente (The Netherlands) (1)
- Universität Bielefeld (1)
- Universität Duisburg-Essen, Institut für Molekularbiologie, AG Becker-Flegler (1)
- Universität Würzburg GmbH für Bildungs- und Campusdienstleistungen (1)
- Universitätsklinikum Würzburg (UKW) (1)
- Université Paris I Panthéon-Sorbonne (1)
- Université d'Abomey-Calavi, Benin (1)
- Université de Bordeaux, Bordeaux, France (1)
- Université de Montréal (1)
- Université de Paris-Saclay (1)
- ZINF Research Center for Infectious Diseases (1)
- Zentrum für Infektionsforschung ZINF Würzburg (1)
ResearcherID
- B-1911-2015 (1)
- B-4606-2017 (1)
- C-2593-2016 (1)
- D-1250-2010 (1)
- I-5818-2014 (1)
- J-8841-2015 (1)
- M-1240-2017 (1)
- N-2030-2015 (1)
- N-3741-2015 (1)
EU-Project number / Contract (GA) number
- 311781 (1)
- 320377 (1)
- EU (FP7/ 2007-2013) (1)
In the scope of climate warming and the increase in frequency and intensity of severe heat waves in Central Europe, identification of temperate tree species that are suited to cope with these environmental changes is gaining increasing importance. A number of tree physiological characteristics are associated with drought-stress resistance and survival following severe heat, but recent studies have shown the importance of plant hydraulic and anatomical traits for predicting drought-induced tree mortality, such as vessel diameter, and their potential to predict species distribution in a changing climate.
A compilation of large global datasets is required to determine traits related to drought-induced embolism and test whether embolism resistance can be determined solely by anatomical traits. However, most measurements of plant hydraulic traits are labour-intense and prone to measurement artefacts. A fast, accurate and widely applicable technique is necessary for estimating xylem embolism resistance (e.g., water potential at 50% loss of conductivity, P50), in order to improve forecasts of future forest changes. These traits and their combination must have evolved following the selective pressure of the environmental conditions in which each species occurs. Describing these environmental-trait relationships can be useful to assess potential responses to environmental change and mitigation strategies for tree species, as future warmer temperatures may be compounded by drier conditions.
RNA viruses rely entirely on the host machinery for their protein synthesis and harbor non-canonical translation mechanisms, such as alternative initiation and programmed –1 ribosomal frameshifting (–1PRF), to suit their specific needs. On the other hand, host cells have developed a variety of defensive strategies to safeguard their translational apparatus and at times transiently shut down global translation. An infection can lead to substantial translational remodeling in cells and translational control is critical during antiviral response. Due to their sheer diversity, this control is likely unique to each RNA virus and the intricacies of post-transcriptional regulation are unclear in certain viral species.
Here, we explored different aspects of translational regulation in virus-infected cells in detail. Using ribosome profiling, we extensively characterized the translational landscape in HIV-1 infected T cells, uncovering novel features of gene regulation in both host and virus. Additionally, we show that substantial pausing occurs prior to the frameshift site indicating complex regulatory mechanisms involving upstream viral RNA elements that can act as cis- regulators of frameshifting.
We also characterized the mechanistic details of trans- modulation of frameshifting by host- and virus-encoded proteins. Host antiviral protein ZAP-S binds to the SARS-CoV-2 frameshift site and destabilizes the stimulatory structure, leading to frameshift inhibition. On the other hand, EMCV 2A protein stabilizes the viral frameshift site, thereby, activating EMCV frameshifting. While both proteins were shown to be antagonistic in their mechanism, they interact with the host translational machinery. Furthermore, we showed that frameshifting can be regulated not just by proteins, but also by small molecules. High-throughput screening of natural and synthetic compounds identified two potent frameshift inhibitors that also impeded viral replication, namely trichangion and compound 25. Together, this work largely enhances our understanding of gene regulation mechanisms in virus-infected cells and further validates the druggability of viral –1 PRF site.
The demand for LIB with enhanced energy densities leads to increased utilization of the space within the confinements of the battery housing or to the use of electrode material with increased intrinsic specific energy densities. Both requirements result in more stress on the battery electrodes and separator during cycling or aging. However, the effect of mechanical strain on the cell’s electrochemistry and thus the performance of batteries is rather unexplored compared to the impact of current or temperature, for example. The objective of this thesis was to give a better understanding of the electrochemical and mechanical interplay in current- and next-generation lithium based battery cells. Therefore, the thesis was structured into the investigations on SoA and next-generation LIBs. For SoA LIBs, the investigations of the interplay started at laboratory scale. Here, the expansion of various electrodes and also the impact of mechanical pressure and its distribution on the performance of the cells were
studied. The investigations at laboratory scale was followed by an examination of the electrochemical and mechanical interactions on large format commercial LIBs which are used in BEVs. Accordingly, the effect of bracing and its effect on the performance was studied in an aging and post-mortem study. To gain a deeper understanding of the mechanical changes in LIBs, an ultrasonic study was performed for pouch cells. Here, the mechanical changes were further investigated in dependence of SoC and SoH. The effects of the mechanical stress on the performance for next-generation batteries were studied at laboratory scale. In the beginning, the expansion of next-generation anode materials such as silicon and lithium was compared with today’s anode materials. Furthermore, the effect of mechanical pressure and electrolyte on the irreversible dilation and performance was investigated for lithium metal cells. Overall, it was shown that pressure has a significant effect on the performance of today’s and also future LIBs. The interplay of the electrochemical and mechanical effects inside a LIB has a considerable impact on the lifetime, capacity fading and impedance increase of the batteries.
Ownership and usage of personal voice assistant devices like Amazon Echo or Google Home have increased drastically over the last decade since their market launch. This thesis builds upon existing computers are social actors (CASA) and media equation research that is concerned with humans displaying social reactions usually exclusive to human-human interaction when interacting with media and technological devices. CASA research has been conducted with a variety of technological devices such as desktop computers, smartphones, embodied virtual agents, and robots. However, despite their increasing popularity, little empirical work has been done to examine social reactions towards these personal stand-alone voice assistant devices, also referred to as smart speakers. Thus, this dissertation aims to adopt the CASA approach to empirically evaluate social responses to smart speakers. With this goal in mind, four laboratory experiments with a total of 407 participants have been conducted for this thesis. Results show that participants display a wide range of social reactions when interacting with voice assistants. This includes the utilization of politeness strategies such as the interviewer-bias, which led to participants giving better evaluations directly to a smart speaker device compared to a separate computer. Participants also displayed prosocial behavior toward a smart speaker after interdependence and thus a team affiliation had been induced. In a third study, participants applied gender stereotypes to a smart speaker not only in self-reports but also exhibited conformal behavior patterns based on the voice the device used. In a fourth and final study, participants followed the rule of reciprocity and provided help to a smart speaker device that helped them in a prior interaction. This effect was also moderated by subjects’ personalities, indicating that individual differences are relevant for CASA research. Consequently, this thesis provides strong empirical support for a voice assistants are social actors paradigm. This doctoral dissertation demonstrates the power and utility of this research paradigm for media psychological research and shows how considering voice assistant devices as social actors lead to a more profound understanding of voice-based technology. The findings discussed in this thesis also have implications for these devices that need to be carefully considered both in future research as well as in practical design.
Most medicines are taken orally. To enter the systemic circulation, they dissolve in the intestinal fluid, cross the epithelial barrier, and pass through the liver. Intestinal absorption is driven by the unique features of the gastrointestinal tract, including the bile colloids formed in the lumen and the mucus layer covering the intestinal epithelium. Neglecting this multifaceted environment can lead to poor drug development decisions, especially for poorly water-soluble drugs that interact with bile and mucus. However, there is a lack of a rationale nexus of molecular interactions between oral medicines and gastrointestinal components with drug bioavailability. Against this background, this thesis aims to develop biopharmaceutical strategies to optimize the presentation of oral therapeutics to the intestinal epithelial barrier.
In Chapter 1, the dynamics of bile colloids upon solubilization of the poorly-water soluble drug Perphenazine was studied. Perphenazine impacted molecular arrangement, structure, binding thermodynamics, and induced a morphological transition from vesicles to worm-like micelles. Despite these dynamics, the bile colloids ensured stable relative amounts of free drug substance. The chapter was published in Langmuir.
Chapter 2 examined the impact of pharmaceutical polymeric excipients on bile-mediated drug solubilization. Perphenazine and Imatinib were introduced as model compounds interacting with bile, whereas Metoprolol did not. Some polymers altered the arrangement and geometry of bile colloids, thereby affecting the molecularly soluble amount of those drugs interacting with bile. These insights into the bile-drug-excipient interplay provide a blueprint to optimizing formulations leveraging bile solubilization. The chapter was published in Journal of Controlled Release.
Chapter 3 deals with the impact of bile on porcine intestinal mucus. Mucus exposed to bile solution changed transiently, it stiffened, and the overall diffusion rate increased. The bile-induced changes eased the transport of the bile-interacting drug substance Fluphenazine, whereas Metoprolol was unaffected. This dichotomous pattern was linked to bioavailability in rats and generalized based on two previously published data sets. The outcomes point to a bile-mucus interaction relevant to drug delivery. The chapter is submitted.
The Appendix provides a guide for biopharmaceutical characterization of drug substances by nuclear magnetic resonance spectroscopy aiming at establishing a predictive algorithm.
In summary, this thesis deciphers bile-driven mechanisms shaping intestinal drug absorption. Based on these molecular insights, pharmaceuticals can be developed along a biopharmaceutical optimization, ultimately leading to better oral drugs of tomorrow.
Academic education is seen as an important place for the development of professionalism of (future) adult educators. Since adult education academia, research, and practice is closely intertwined with global and international de- velopments, there is a need for adult education programmes to prepare their students for these interconnections. This can be examined in the context of international teaching and learning settings that integrate international, inter- cultural, or global perspectives into teaching and learning and are part of the internationalisation efforts of higher education. The focus of this international and comparative study is on how international teaching and learning settings contribute to the academic professionalisation in adult education in three mas- ter’s programmes with a focus on adult education at the University of Würzburg (Germany), University of Belgrade (Serbia) and University of Florence (Italy). International teaching and learning settings are examined on the structural and individual level of academic professionalisation. The aim is to explore the provision of international teaching and learning settings in the master’s pro- grammes on the one hand, and to analyse the contribution of international teach- ing and learning settings to the development of students’ professionalism on the other. For this purpose, three focus group interviews with programme heads, (academic) staff, and students as well as 22 guided interviews with graduates of the three master’s programmes at the three university locations are collected and analysed in an international and comparative study design. The study reveals similarities and differences in the forms, framework con- ditions, and goals of international teaching and learning settings between the three master’s programmes. Overarching contexts that guide the internationalisation of the master’s programmes become apparent (e.g. education and higher education policy, internationalisation of the university, programme structure). The triangulation of the interview data of the graduates shows that the interna- tional environment, the structural arrangement, and the practical relevance of the international teaching and learning settings support the development of the graduates’ professionalism. The results underline the relevance of international teaching and learning settings for the development of professionalism in adult education and point to the requirement for a systematic and comprehensive in- ternationalisation of adult education programmes.
The current study presents a new a group of Demotic ostraca in the belongings of the Cairo Museum. A large part of this group stem from Medinet Habu in the western bank of modern Luxor in Upper Egypt and was discovered in the beginning of the thirties of the last century by the Chicago Oriental Institute (recently renamed as Institute for the Study of Ancient Cultures ‘ISAC’). A small portion of the collection under consideration come from other Upper Egyptian provenances including Gebelein, Edfu, Kom Ombo, and possibly elsewhere in Thebes. The main goal of the present dissertation is to decipher, translate, and provide a philological, paleographical, and cultural analysis of the group of texts in question. The results of this study are spread over two main parts, the first of which is dedicated to the main and largest part of the collection, i.e. ostraca from Medinet Habu, while the second is concerned with ostraca from other places. The first part comprises of five sections beginning with receipts of money and in-kind payments including some receipts for the payments of the different capitation charges in the Ptolemaic and Roman Periods, a few for land-related payments, as well as others related to different Ptolemaic monopolies or trades such as a receipt for the price of oil, one for the linen tax, in addition to a unique receipt for the rarely attested fish tax. The second section includes accounts and lists of different kinds be it monetary, in-kind, agriculture, or any other type of lists or accounts that record different everyday transactions. The following section presents a relatively different type of lists, namely lists of personal names. The fourth section incorporates a variety of texts of different concerns, e.g. texts of religious nature, letters, temples oaths, or other private documents. Unidentified texts occupy the fifth and final section of the first part. The second part of the study, which comprises texts that originate from different Upper Egyptian localities, includes three sections, i.e. receipts, accounts, and lists of names.
Within this thesis, three main approaches for the assessment and investigation of altered hemodynamics like wall shear stress, oscillatory shear index and the arterial pulse wave velocity in atherosclerosis development and progression were conducted:
1. The establishment of a fast method for the simultaneous assessment of 3D WSS and PWV in the complete murine aortic arch via high-resolution 4D-flow MRI
2. The utilization of serial in vivo measurements in atherosclerotic mouse models using high-resolution 4D-flow MRI, which were divided into studies describing altered hemodynamics in late and early atherosclerosis
3. The development of tissue-engineered artery models for the controllable application and variation of hemodynamic and biologic parameters, divided in native artery models and biofabricated artery models, aiming for the investigation of the relationship between atherogenesis and hemodynamics
Chapter 2 describes the establishment of a method for the simultaneous measurement of 3D WSS and PWV in the murine aortic arch at, using ultra high-field MRI at 17.6T [16], based on the previously published method for fast, self-navigated wall shear stress measurements in the murine aortic arch using radial 4D-phase contrast MRI at 17.6 T [4]. This work is based on the collective work of Dr. Patrick Winter, who developed the method and the author of this thesis, Kristina Andelovic, who performed the experiments and statistical analyses. As the method described in this chapter is basis for the following in vivo studies and undividable into the sub-parts of the contributors without losing important information, this chapter was not split into the single parts to provide fundamental information about the measurement and analysis methods and therefore better understandability for the following studies. The main challenge in this chapter was to overcome the issue of the need for a high spatial resolution to determine the velocity gradients at the vascular wall for the WSS quantification and a high temporal resolution for the assessment of the PWV without prolonging the acquisition time due to the need for two separate measurements. Moreover, for a full coverage of the hemodynamics in the murine aortic arch, a 3D measurement is needed, which was achieved by utilization of retrospective navigation and radial trajectories, enabling a highly flexible reconstruction framework to either reconstruct images at lower spatial resolution and higher frame rates for the acquisition of the PWV or higher spatial resolution and lower frame rates for the acquisition of the 3D WSS in a reasonable measurement time of only 35 minutes. This enabled the in vivo assessment of all relevant hemodynamic parameters related to atherosclerosis development and progression in one experimental session. This method was validated in healthy wild type and atherosclerotic Apoe-/- mice, indicating no differences in robustness between pathological and healthy mice.
The heterogeneous distribution of plaque development and arterial stiffening in atherosclerosis [10, 12], however, points out the importance of local PWV measurements. Therefore, future studies should focus on the 3D acquisition of the local PWV in the murine aortic arch based on the presented method, in order to enable spatially resolved correlations of local arterial stiffness with other hemodynamic parameters and plaque composition.
In Chapter 3, the previously established methods were used for the investigation of changing aortic hemodynamics during ageing and atherosclerosis in healthy wild type and atherosclerotic Apoe-/- mice using the previously established methods [4, 16] based on high-resolution 4D-flow MRI. In this work, serial measurements of healthy and atherosclerotic mice were conducted to track all changes in hemodynamics in the complete aortic arch over time. Moreover, spatially resolved 2D projection maps of WSS and OSI of the complete aortic arch were generated. This important feature allowed for the pixel-wise statistical analysis of inter- and intragroup hemodynamic changes over time and most importantly – at a glance. The study revealed converse differences of local hemodynamic profiles in healthy WT and atherosclerotic Apoe−/− mice, with decreasing longWSS and increasing OSI, while showing constant PWV in healthy mice and increasing longWSS and decreasing OSI, while showing increased PWV in diseased mice. Moreover, spatially resolved correlations between WSS, PWV, plaque and vessel wall characteristics were enabled, giving detailed insights into coherences between hemodynamics and plaque composition. Here, the circWSS was identified as a potential marker of plaque size and composition in advanced atherosclerosis. Moreover, correlations with PWV values identified the maximum radStrain could serve as a potential marker for vascular elasticity. This study demonstrated the feasibility and utility of high-resolution 4D flow MRI to spatially resolve, visualize and analyze statistical differences in all relevant hemodynamic parameters over time and between healthy and diseased mice, which could significantly improve our understanding of plaque progression towards vulnerability. In future studies the relation of vascular elasticity and radial strain should be further investigated and validated with local PWV measurements and CFD.
Moreover, the 2D histological datasets were not reflecting the 3D properties and regional characteristics of the atherosclerotic plaques. Therefore, future studies will include 3D plaque volume and composition analysis like morphological measurements with MRI or light-sheet microscopy to further improve the analysis of the relationship between hemodynamics and atherosclerosis.
Chapter 4 aimed at the description and investigation of hemodynamics in early stages of atherosclerosis. Moreover, this study included measurements of hemodynamics at baseline levels in healthy WT and atherosclerotic mouse models. Due to the lack of hemodynamic-related studies in Ldlr-/- mice, which are the most used mouse models in atherosclerosis research together with the Apoe-/- mouse model, this model was included in this study to describe changing hemodynamics in the aortic arch at baseline levels and during early atherosclerosis development and progression for the first time. In this study, distinct differences in aortic geometries of these mouse models at baseline levels were described for the first time, which result in significantly different flow- and WSS profiles in the Ldlr-/- mouse model. Further basal characterization of different parameters revealed only characteristic differences in lipid profiles, proving that the geometry is highly influencing the local WSS in these models. Most interestingly, calculation of the atherogenic index of plasma revealed a significantly higher risk in Ldlr-/- mice with ongoing atherosclerosis development, but significantly greater plaque areas in the aortic arch of Apoe-/- mice. Due to the given basal WSS and OSI profile in these two mouse models – two parameters highly influencing plaque development and progression – there is evidence that the regional plaque development differs between these mouse models during very early atherogenesis.
Therefore, future studies should focus on the spatiotemporal evaluation of plaque development and composition in the three defined aortic regions using morphological measurements with MRI or 3D histological analyses like LSFM. Moreover, this study offers an excellent basis for future studies incorporating CFD simulations, analyzing the different measured parameter combinations (e.g., aortic geometry of the Ldlr-/- mouse with the lipid profile of the Apoe-/- mouse), simulating the resulting plaque development and composition. This could help to understand the complex interplay between altered hemodynamics, serum lipids and atherosclerosis and significantly improve our basic understanding of key factors initiating atherosclerosis development.
Chapter 5 describes the establishment of a tissue-engineered artery model, which is based on native, decellularized porcine carotid artery scaffolds, cultured in a MRI-suitable bioreactor-system [23] for the investigation of hemodynamic-related atherosclerosis development in a controllable manner, using the previously established methods for WSS and PWV assessment [4, 16]. This in vitro artery model aimed for the reduction of animal experiments, while simultaneously offering a simplified, but completely controllable physical and biological environment. For this, a very fast and gentle decellularization protocol was established in a first step, which resulted in porcine carotid artery scaffolds showing complete acellularity while maintaining the extracellular matrix composition, overall ultrastructure and mechanical strength of native arteries. Moreover, a good cellular adhesion and proliferation was achieved, which was evaluated with isolated human blood outgrowth endothelial cells. Most importantly, an MRI-suitable artery chamber was designed for the simultaneous cultivation and assessment of high-resolution 4D hemodynamics in the described artery models. Using high-resolution 4D-flow MRI, the bioreactor system was proven to be suitable to quantify the volume flow, the two components of the WSS and the radStrain as well as the PWV in artery models, with obtained values being comparable to values found in literature for in vivo measurements. Moreover, the identification of first atherosclerotic processes like intimal thickening is achievable by three-dimensional assessment of the vessel wall morphology in the in vitro models. However, one limitation is the lack of a medial smooth muscle cell layer due to the dense ECM. Here, the utilization of the laser-cutting technology for the generation of holes and / or pits on a microscale, eventually enabling seeding of the media with SMCs showed promising results in a first try and should be further investigated in future studies. Therefore, the proposed artery model possesses all relevant components for the extension to an atherosclerosis model which may pave the way towards a significant improvement of our understanding of the key mechanisms in atherogenesis.
Chapter 6 describes the development of an easy-to-prepare, low cost and fully customizable artery model based on biomaterials. Here, thermoresponsive sacrificial scaffolds, processed with the technique of MEW were used for the creation of variable, biomimetic shapes to mimic the geometric properties of the aortic arch, consisting of both, bifurcations and curvatures. After embedding the sacrificial scaffold into a gelatin-hydrogel containing SMCs, it was crosslinked with bacterial transglutaminase before dissolution and flushing of the sacrificial scaffold. The hereby generated channel was subsequently seeded with ECs, resulting in an easy-to-prepare, fast and low-cost artery model. In contrast to the native artery model, this model is therefore more variable in size and shape and offers the possibility to include smooth muscle cells from the beginning. Moreover, a custom-built and highly adaptable perfusion chamber was designed specifically for the scaffold structure, which enabled a one-step creation and simultaneously offering the possibility for dynamic cultivation of the artery models, making it an excellent basis for the development of in vitro disease test systems for e.g., flow-related atherosclerosis research. Due to time constraints, the extension to an atherosclerosis model could not be achieved within the scope of this thesis. Therefore, future studies will focus on the development and validation of an in vitro atherosclerosis model based on the proposed bi- and three-layered artery models.
In conclusion, this thesis paved the way for a fast acquisition and detailed analyses of changing hemodynamics during atherosclerosis development and progression, including spatially resolved analyses of all relevant hemodynamic parameters over time and in between different groups. Moreover, to reduce animal experiments, while gaining control over various parameters influencing atherosclerosis development, promising artery models were established, which have the potential to serve as a new platform for basic atherosclerosis research.
Drug Discovery based on Oxidative Stress and HDAC6 for Treatment of Neurodegenerative Diseases
(2024)
Most antioxidants reported so far only achieved limited success in AD clinical trials. Growing evidences suggest that merely targeting oxidative stress will not be sufficient to fight AD. While multi-target directed ligands could synergistically modulate different steps in the neurodegenerative process, offering a promising potential for treatment of this complex disease.
Fifteen target compounds have been designed by merging melatonin and ferulic acid into the cap group of a tertiary amide HDAC6 inhibitor. Compound 10b was screened as the best hybrid molecule exhibit potent HDAC6 inhibition and potent antioxidant capacity. Compound 10b also alleviated LPS-induced microglia inflammation and led to a switch from neurotoxic M1 to the neuroprotective M2 microglial phenotype. Moreover, compound 10b show pronounced attenuation of spatial working memory and long-term memory damage in an in vivo AD mouse model. Compound 10b can be a potentially effective drug candidate for treatment of AD and its druggability worth to be further studied.
We have designed ten novel neuroprotectants by hybridizing with several common antioxidants, including ferulic acid, melatonin, lipoic acid, and trolox. The trolox hybrid compound exhibited the most potent neuroprotective effects in multiple neuroprotection assays. Besides, we identified the synergistic effects between trolox and vitamin K derivative, and our trolox hybrid compound showed comparable neuroprotection with the mixture of trolox and vitamin K derivative.
We have designed and synthesized 24 quinone derivatives based on five kinds of different quinones including ubiquinone, 2,3,5-trimethyl-1,4-benzoquinone, memoquin, thymoquinone, and anthraquinone. Trimethylbenzoquinone and thymoquinone derivatives showed more potent neuroprotection than other quinones in oxytosis assay. Therefore, trimethylbenzoquinone and thymoquinone derivatives can be used as lead compounds for further mechanism study and drug discovery for treatment of neurodegenerative disease.
We designed a series of photoswitchable HDAC inhibitors, which could be effective molecular tools due to the high spatial and temporal resolution. In total 23 target compounds were synthesized and photophysicochemically characterized. Azoquinoline-based compounds possess more thermally stable cis-isomers in buffer solution, which were further tested in enzyme-based HDAC inhibition assay. However, none of those tested compounds show significant differences in activities between trans-isomers and corresponding cis-isomers.
Ecophysiological adaptations of the cuticular water permeability within the Solanaceae family
(2024)
The cuticle, a complex lipidic layer synthesized by epidermal cells, covers and protects primary organs of all land plants. Its main function is to avoid plant desiccation by limiting non-stomatal water loss. The cuticular properties vary widely among plant species. So far, most of the cuticle-related studies have focused on a limited number of species, and studies addressing phylogenetically related plant species are rare. Moreover, comparative studies among organs from the same plant species are still scarce.
Thus, this study focus on organ-specificities of the cuticle within and between plant species of the Solanaceae family. Twenty-seven plant species of ten genera, including cultivated and non- cultivated species, were investigated to identify potential cuticular similarities. Structural, chemical and functional traits of fully expanded leaves, inflated fruiting calyces, and ripe fruits were analyzed.
The surface morphology was investigated by scanning electron microscopy. Leaves were mainly amphistomatic and covered by an epicuticular wax film. The diversity and distribution of trichomes varied among species. Only the leaves of S. grandiflora were glabrous. Plant species of the Leptostemonum subgenus had numerous prickles and non-glandular stellate trichomes. Fruits were stomata-free, except for S. muricatum, and a wax film covered their surface. Last, lenticel- like structures and remaining scars of broken trichomes were found on the surface of some Solanum fruits.
Cuticular water permeability was used as indicators of the cuticular transpiration barrier efficiency. The water permeability differed among plant species, organs and fruit types with values ranging up to one hundred-fold. The minimum leaf conductance ranged from 0.35 × 10-5 m s-1 in S. grandiflora to 31.54 × 10-5 m s-1 in S. muricatum. Cuticular permeability of fruits ranged from 0.64 × 10-5 m s-1 in S. dulcamara (fleshy berry) to 34.98 × 10-5 m s-1 in N. tabacum (capsule). Generally, the cuticular water loss of dry fruits was about to 5-fold higher than that of fleshy fruits.
Interestingly, comparisons between cultivated and non-cultivated species showed that wild species have the most efficient cuticular transpiration barrier in leaves and fruits. The average permeability of leaves and fruits of wild plant species was up to three-fold lower in comparison to the cultivated ones. Moreover, ripe fruits of P. ixocarpa and P. peruviana showed two-times lower cuticular transpiration when enclosed by the inflated fruiting calyx.
The cuticular chemical composition was examined using gas chromatography. Very-long-chain aliphatic compounds primarily composed the cuticular waxes, being mostly dominated by n- alkanes (up to 80% of the total wax load). Primary alkanols, alkanoic acids, alkyl esters and branched iso- and anteiso-alkanes were also frequently found. Although in minor amounts, sterols, pentacyclic triterpenoids, phenylmethyl esters, coumaric acid esters, and tocopherols were identified in the cuticular waxes. Cuticular wax coverages highly varied in solanaceous (62- fold variation). The cuticular wax load of fruits ranged from 0.55 μg cm−2 (Nicandra physalodes) to 33.99 μg cm−2 (S. pennellii), whereas the wax amount of leaves varied from 0.90 μg cm−2 (N. physalodes) to 28.42 μg cm−2 (S. burchellii). Finally, the wax load of inflated fruiting calyces ranged from 0.56 μg cm−2 in P. peruviana to 2.00 μg cm−2 in N. physalodes.
For the first time, a comparative study on the efficiency of the cuticular transpiration barrier in different plant organs of closely related plant species was conducted. Altogether, the cuticular chemical variability found in solanaceous species highlight species-, and organ-specific wax biosynthesis. These chemical variabilities might relate to the waterproofing properties of the plant cuticle, thereby influencing leaf and fruit performances. Additionally, the high cuticular water permeabilities of cultivated plant species suggest a potential existence of a trade-off between fruit organoleptic properties and the efficiency of the cuticular transpiration barrier. Last, the high cuticular water loss of the solanaceous dry fruits might be a physiological adaptation favouring seed dispersion.
In aqueous environment, hydrophobic interactions play an important role for DNA. The introduction of modifications based on hydrophobic aromatic moieties offers additional ways for controlling recognition and reactivity of functional groups in DNA. Modifications are introduced through an artificial backbone or in the form of an extension of the nucleobases, resulting in additional properties of the DNA.
This dissertation focuses on the use of hydrophobic units for the functionalization of DNA.
In the first part of the work, the tolane (i. e. diphenylacetylene) motif was used in combination with the acyclic backbone of GNA and BuNA to generate recognition units in the DNA context. Fluorination of the aromatic rings in the tolane moiety provided the basis for a supramolecular language based on arene-fluoroarene interactions. The specific recognition was investigated by thermodynamic, kinetic and NMR spectroscopic methods.
In the second part of the work, deoxyuridine derivatives with a hydrophobic aromatic modification were prepared and incorporated into DNA duplexes. The irradiation with UV light led to a [2+2] cycloaddition reaction between two modified nucleosides in the DNA. This reaction product was structurally characterized and the reaction was used in various biochemical and nanotechnological DNA applications.
The detection of smallest mechanical loads plays an increasingly important role in many areas of advancing automation and manufacturing technology, but also in everyday life. In this doctoral thesis, various microparticle systems were developed that are able to indicate mechanical shear stress via simple mechanisms. Using a toolbox approach, these systems can be spray-dried from various nanoscale primary particles (silica and iron oxide) to micrometer-sized units, so-called supraparticles. By varying the different building blocks and in combination with different dyes, a new class of mechanochromic shear stress indicators was developed by constructing hierarchically structured core-shell supraparticles that can indicate mechanical stress via an easily detectable color change. Three different mechanisms can be distinguished. If a signal becomes visible only by a mechanical load, it is a turn-on indicator. In the opposite case, the turn-off indicator, the signal is switched off by a mechanical load. In the third mechanism, the color-change indicator, the color changes as a result of a mechanical load. In principle, these indicators can be used in two different ways. First, they can be incorporated into a coating as an additive. These coatings can be applied to a wide range of products, including food packaging, medical devices, and generally any sensitive surface where mechanical stress, such as scratches, is difficult to detect but can have serious consequences. Second, these shear stress indicators can also be used directly in powder form and for example then applied in 3D-printing or in ball mills. A total of six different shear stress indicators were developed, three of which were used as additives in coatings and three were applied in powder form. Depending on their composition, these indicators were readout by fluorescence, UV-Vis or Magnetic Particle Spectroscopy. The development of these novel shear stress indicator supraparticles were successfully combined molecular chemistry with the world of nano-objects to develop macroscopic systems that can enable smart and communicating materials to indicate mechanical stress in a variety of applications.
Lung cancer is the main cause of cancer-related deaths worldwide. Despite the availability of several targeted therapies and immunotherapies in the clinics, the prognosis for lung cancer remains poor. A major problem for the low benefit of these therapies is intrinsic and acquired resistance, asking for pre-clinical models for closer investigation of predictive biomarkers for refined personalized medicine and testing of possible combination therapies as well as novel therapeutic approaches to break resistances.
One third of all lung adenocarcinoma harbor mutations in the KRAS gene, of which 39 % are transitions from glycine to cysteine in codon 12 (KRASG12C). Being considered “undruggable” in previous decades, KRASG12C-inhibitors now paved the way into the standard-of-care for lung adenocarcinoma treatment in the clinics. Still, the overall response rates as well as overall survival of patients treated with KRASG12C-inhibitors are sobering. Therefore, 3D KRASG12C-biomarker in vitro models were developed based on a decellularized porcine jejunum (SISmuc) using commercial and PDX-derived cell lines and characterized in regards of epithelial-mesenchymal-transition (EMT), stemness, proliferation, invasion and c-MYC expression as well as the sensitivity towards KRASG12C-inhibiton. The phenotype of lung tumors harboring KRAS mutations together with a c-MYC overexpression described in the literature regarding invasion and proliferation for in vivo models was well represented in the SISmuc models. A higher resistance towards targeted therapies was validated in the 3D models compared to 2D cultures, while reduced viability after treatment with combination therapies were exclusively observed in the 3D models. In the test system neither EMT, stemness nor the c-MYC expression were directly predictive for drug sensitivity. Testing of a panel of combination therapies, a sensitizing effect of the aurora kinase A (AURKA) inhibitor alisertib for the KRASG12C-inhibitor ARS-1620 directly correlating with the level of c-MYC expression in the corresponding 3D models was observed. Thereby, the capability of SISmuc tumor models as an in vitro test system for patient stratification was demonstrated, holding the possibility to reduce animal experiments.
Besides targeted therapies the treatment of NSCLC with oncolytic viruses (OVs) is a promising approach. However, a lack of in vitro models to test novel OVs limits the transfer from bench to bedside. In this study, 3D NSCLC models based on the SISmuc were evaluated for their capability to perform efficacy and risk assessment of oncolytic viruses (OVs) in a pre-clinical setting. Hereby, the infection of cocultures of tumor cells and fibroblasts on the SISmuc with provided viruses demonstrated that in contrast to a wildtype herpes simplex virus 1 (HSV-1) based OV, the attenuated version of the OV exhibited specificity for NSCLC cells with a more advanced and highly proliferative phenotype, while fibroblasts were no longer permissive for infection. This approach introduced SISmuc tumor models as novel test system for in vitro validation of OVs.
Finally, a workflow for validating the efficacy of anti-cancer therapies in 3D tumor spheroids was established for the transfer to an automated platform based on a two-arm-robot system. In a proof-of-concept process, H358 spheroids were characterized and treated with the KRASG12C-inhibitor ARS-1620. A time- and dose-dependent reduction of the spheroid area after treatment was defined together with a live/dead-staining as easy-to-perform and cost-effective assays for automated drug testing that can be readily performed in situ in an automated system.
Theory and simulation of ultrafast autodetachment dynamics and nonradiative relaxation in molecules
(2024)
In this thesis, theoretical approaches for the simulation of electron detachment processes in molecules following vibrational or electronic excitation are developed and applied. These approaches are based on the quantum-classical surface-hopping methodology, in which nuclear motion is treated classically as an ensemble of trajectories in the potential of quantum-mechanically described electronic degrees of freedom.
After priming in Peyer's patches (PPs) and mesenteric lymph nodes (mLN) T- cells infiltrate the intestine through lymphatic draining and homing through the bloodstream. However, we found that in mouse models of acute graft-versus-host disease (GvHD), a subset of alloreactive T-cells directly migrates from PPs to the adjacent intestinal lamina propria (LP), bypassing the normal lymphatic drainage and vascular trafficking routes. Notably, this direct migration occurred in irradiated and unirradiated GvHD models, indicating that irradiation is not a prerequisite for this observed behavior.
Next, we established a method termed serial intravascular staining (SIVS) in mouse models to systematically investigate the trafficking and migration of donor T- cells in the early stages of acute GvHD initiation. We found that the direct migration of T-cells from PPs to LP resulted in faster recruitment of cells after allogeneic hematopoietic cell transplantation (allo-HCT). These directly migrating T-cells were found to be in an activated and proliferative state, exhibiting a TH1/TH17-like phenotype and producing cytokines such as IFN-γ and TNF-α. Furthermore, we observed that the directly migrating alloreactive T-cells expressed specific integrins (α4+, αE+) and chemokine receptors (CxCR3+, CCR5+, and CCR9+). Surprisingly, blocking these integrins and chemokine-coupled receptors did not hinder the direct migration of T- cells from PPs to LP, suggesting the involvement of alternative mechanisms. Previous experiments ruled out the involvement of S1PR1 and topographical features of macrophages, leading us to hypothesize that mediators of cytoskeleton reorganization, such as Coro1a, Dock2, or Cdc42, may play a role in this unique migration process.
Additionally, we observed that directly migrating T-cells created a local inflammatory microenvironment, which attracts circulating T-cells. Histological analysis confirmed that alloreactive PPs-derived T-cells and bloodborne T-cells colocalized. We employed two experimental approaches, including either photoconversion of T-cells in PPs or direct transfer of activated T-cells into the vasculature, to demonstrate this colocalization. We hypothesize that cytokines released by migrating T-cells, such as IFN-γ and TNF-α, may play a role in recruiting T-cells from the vasculature, as inhibiting chemokine-coupled receptors did not impair recruitment.
The unicellular pathogen Trypanosoma brucei is the causative agent of African
trypanosomiasis, an endemic disease prevalent in sub-Saharan Africa. Trypanosoma brucei alternates between a mammalian host and the tsetse fly vector. The extracellular parasite survives in the mammalian bloodstream by periodically exchanging their ˈvariant surface glycoproteinˈ (VSG) coat to evade the host immune response. This antigenic variation is achieved through monoallelic expression of one VSG variant from subtelomeric ˈbloodstream
form expression sitesˈ (BES) at a given timepoint. During the differentiation from the bloodstream form (BSF) to the procyclic form (PCF) in the tsetse fly midgut, the stage specific surface protein is transcriptionally silenced and replaced by procyclins. Due to their subtelomeric localization on the chromosomes, VSG transcription and silencing is partly regulated by homologues of the mammalian telomere complex such as TbTRF, TbTIF2 and TbRAP1 as well as by ˈtelomere-associated proteinsˈ (TelAPs) like TelAP1. To gain more insights into transcription regulation of VSG genes, the identification and characterization of other TelAPs is critical and has not yet been achieved. In a previous study, two biochemical approaches were used to identify other novel TelAPs. By using ˈco-immunoprecipitationˈ (co-IP) to enrich possible interaction partners of TbTRF and by affinity chromatography using telomeric repeat oligonucleotides, a listing of TelAP candidates has been conducted. With this approach TelAP1 was identified as a novel component of the telomere complex, involved in the kinetics of transcriptional BES silencing during BSF to PCF differentiation. To gain further insights into the telomere complex composition, other previously enriched proteins were characterized through a screening process using RNA interference to deplete potential candidates. VSG expression profile changes and overall proteomic changes after depletion were analyzed by mass spectrometry. With this method, one can gain insights into the functions of the proteins and their involvement in VSG expression site regulation. To validate the interaction of proteins enriched by co-IP with TbTRF and TelAP1 and to identify novel interaction proteins, I performed reciprocal affinity purifications of the four most promising candidates (TelAP2, TelAP3, PPL2 and PolIE) and additionally confirmed colocalization of two candidates with TbTRF via immunofluorescence (TelAP2, TelAP3). TelAP3 colocalizes with TbTRF and potentially interacts with TbTRF, TbTIF2, TelAP1 and TelAP2, as well as with two translesion polymerases PPL2 and PolIE in BSF. PPL2 and PolIE seem to be in close contact to each other at the telomeric ends and fulfill different roles as only PolIE is involved in VSG regulation while PPL2 is not. TelAP2 was previously characterized to be associated with telomeres by partially colocalizing with TbTRF and cells show a VSG derepression phenotype when the protein was depleted. Here I show that TelAP2 interacts with the telomere-binding proteins TbTRF and TbTIF2 as well as with the telomere-associated protein TelAP1 in BSF and that TelAP2 depletion results in a loss of TelAP1 colocalization with TbTRF in BSF.
In conclusion, this study demonstrates that characterizing potential TelAPs is effective in gaining insights into the telomeric complex's composition and its role in VSG regulation in Trypanosoma brucei. Understanding these interactions could potentially lead to new therapeutic targets for combatting African trypanosomiasis.
Biofabrication technologies must address numerous parameters and conditions to reconstruct tissue complexity in vitro. A critical challenge is vascularization, especially for large constructs exceeding diffusion limits. This requires the creation of artificial vascular structures, a task demanding the convergence and integration of multiple engineering approaches. This doctoral dissertation aims to achieve two primary objectives: firstly, to implement and refine engineering methods for creating artificial microvascular structures using Melt Electrowriting (MEW)-assisted sacrificial templating, and secondly, to deepen the understanding of the critical factors influencing the printability of bioink formulations in 3D extrusion bioprinting.
In the first part of this dissertation, two innovative sacrificial templating techniques using MEW are explored. Utilizing a carbohydrate glass as a fugitive material, a pioneering advancement in the processing of sugars with MEW with a resolution under 100 microns was made. Furthermore, by introducing the “print-and-fuse” strategy as a groundbreaking method, biomimetic branching microchannels embedded in hydrogel matrices were fabricated, which can then be endothelialized to mirror in vivo vascular conditions.
The second part of the dissertation explores extrusion bioprinting. By introducing a simple binary bioink formulation, the correlation between physical properties and printability was showcased. In the next step, employing state-of-the-art machine-learning approaches revealed a deeper understanding of the correlations between bioink properties and printability in an extended library of hydrogel formulations.
This dissertation offers in-depth insights into two key biofabrication technologies. Future work could merge these into hybrid methods for the fabrication of vascularized constructs, combining MEW's precision with fine-tuned bioink properties in automated extrusion bioprinting.
Adoptive immunotherapy using chimeric antigen receptor (CAR)-modified T cells is an effective treatment for hematological malignancies that are refractory to conventional chemotherapy. To address a wider variety of cancer entities, there is a need to identify and characterize additional target antigens for CAR-T cell therapy. The two members of the receptor tyrosine kinase-like orphan receptor family, ROR1 and ROR2, have been found to be overexpressed on cancer cells and to correlate with aggressive cancer phenotypes. Recently, ROR1-specific CAR-T cells have entered testing in phase I clinical trials, encouraging us to assess the suitability of ROR2 as a novel target for CAR-T cell therapy. To study the therapeutic potential of targeting ROR2 in solid and hematological malignancies, we selected two representative cancer entities with high unmet medical need: renal cell carcinoma and multiple myeloma.
Our data show that ROR2 is commonly expressed on primary samples and cell lines of clear cell renal cell carcinoma and multiple myeloma. To study the efficacy of ROR2-specific CAR T cell therapy, we designed two CAR constructs with 10-fold binding affinity differences for the same epitope of ROR2. We found both cell products to exhibit antigen-specific anti-tumor reactivity in vitro, including tumor cell lysis, secretion of the effector cytokines interleukin-2 (IL-2) and interferon-gamma (IFNγ), and T cell proliferation. In vivo studies revealed ROR2 specific CAR-T cells to confer durable responses, significant survival benefits and long-term persistence of CAR-expressing T cells. Overall, there was a trend towards more potent anti-tumor efficacy upon treatment with T cells that expressed the CAR with higher affinity for ROR2, both in vitro and in vivo.
We performed a preclinical safety and toxicology assessment comprising analyses of ROR2 expression in healthy human and murine tissues, cross-reactivity, and adoptive T cell transfer in immunodeficient mice. We found ROR2 expression to be conserved in mice, and low-level expression was detectable in the male and female reproductive system as well as parts of the gastrointestinal tract. CAR-T cells targeting human ROR2 were found to elicit similarly potent reactivity upon recognition of murine ROR2. In vivo analyses showed transient tissue-specific enrichment and activation of ROR2-specific CAR-T cells in organs with high blood circulation, such as lung, liver, or spleen, without evidence for clinical toxicity or tissue damage as determined by histological analyses.
Furthermore, we humanized the CAR binding domain of ROR2-specific CAR-T cells to mitigate the risk of adverse immune reactions and concomitant CAR-T cell rejection. Functional analyses confirmed that humanized CARs retained their specificity and functionality against ROR2-positive tumor cells in vitro.
In summary, we show that ROR2 is a prevalent target in RCC and MM, which can be addressed effectively with ROR2-specific CAR-T cells in preclinical models. Our preliminary toxicity studies suggest a favorable safety profile for ROR2-specific CAR-T cells. These findings support the potential to develop ROR2-specific CAR-T cells clinically to obtain cell products with broad utility.
Plasmonic nanostructures are considered promising candidates for essential components of integrated quantum technologies because of their ability to efficiently localize broad-band electromagnetic fields on the nanoscale. The resulting local near field can be understood as a spatial superposition of spectrally different plasmon-polariton modes due to the spectrally broad optical excitation, and thus can be described as a classical wave packet. Since plasmon polaritons, in turn, can transmit and receive non-classical light states, the exciting question arises to what extent they have to be described as quantum mechanical wave packets, i.e. as a superposition of different quantum states.
But how to probe, characterize and eventually manipulate the quantum state of such plasmon polaritons? Up to now, probing at room temperatures relied completely on analyzing quantum optical properties of the corresponding in-going and out-going far-field photon modes. However, these methods so far only allow a rather indirect investigation of the plasmon-polariton quantum state by means of transfer into photons. Moreover, these indirect methods lack spatial resolution and therefore do not provide on-site access to the plasmon-polariton quantum state. However, since the spectroscopic method of coherent two-dimensional (2D) nanoscopy offers the capability to follow the plasmon-
polariton quantum state both in Hilbert space and in space and time domain a complete characterization of the plasmon polariton is possible.
In this thesis a versatile coherent 2D nanoscopy setup is presented combining spectral tunability and femtosecond time resolution with spatial resolution on the nanometer scale due to the detection of optically excited nonlinear emitted electrons via photoemission electron microscopy (PEEM). Optical excitation by amplitude- and phase-shaped, systematically-modified and interferometric-stable multipulse sequences is realized, and characterized via Fourier-transform spectral interferometry (FTSI). This linear technique enables efficient data acquisition in parallel to a simultaneously performed experiment. The full electric-field reconstruction of every generated multipulse sequence is used to analyze the effect of non-ideal pulse sequences on the two-dimensional spectral data of population-based multidimensional spectroscopy methods like, e.g., the coherent 2D nanoscopy applied in this thesis. Investigation of the spatially-resolved nonlinear electron emission yield from plasmonic gold nanoresonators by coherent 2D nanoscopy requires a quasi-particle treatment of the addressed plasmon-polariton mode and development of a quantum model to adequately describe the plasmon-assisted multi-quantum electron emission from nanostructures. Good agreement between simulated and experimental data enables to connect certain spectral features to superpositions of non-adjacent plasmon-polariton quantum states, i.e, non-adjacent occupation-number states of the underlying quantized, harmonic oscillator, thus direct probing of the plasmon-polariton quantum wave packet at the location of the nanostructure.
This is a necessary step to locally control and manipulate the plasmon-polariton quantum state and thus of general interest for the realization of nanoscale quantum optical devices.
Biological systems are in dynamic interaction. Many responses reside in the core concepts of biological systems interplay (competition and cooperation). In infection situation, the competition between a bacterial system and a host is shaped by many stressors at spatial and temporal determinants. Reactive chemical species are universal stressors against all biological systems since they potentially damage the basic requirements of these systems (nucleic acids, proteins, carbohydrates, and lipids). Either produced endogenously or exogenously, reactive chemical species affect the survival of pathogens including the gram-positive
Staphylococcus aureus (S. aureus). Therefore, bacteria developed strategies to overcome the toxicity of reactive species.
S. aureus is a widely found opportunistic pathogen. In its niche, S. aureus is in permanent contact with surrounding microbes and host factors. Deciphering the deterministic factors
in these interactions could facilitate pinpointing novel bacterial targets. Identifying
the aforementioned targets is crucial to develop new strategies not only to kill the pathogenic organisms but also to enhance the normal flora to minimize the pathogenicity and virulence of potential pathogens. Moreover, targeting S. aureus stress response can be used
to overcome bacterial resistance against host-derived factors. In this study, I identify a novel
S. aureus stress response factor against reactive electrophilic, oxygen, and hypochlorite species to better understand its resilience as a pathogen.
Although bacterial stress response is an active research field, gene function is a current bottleneck in characterizing the understudied bacterial strategies to mediate stress conditions. I aimed at understanding the function of a novel protein family integrated
in many defense systems of several biological systems.
In bacteria, fungi, and plants, old yellow enzymes (OYEs) are widely found. Since the first isolation of the yellow flavoprotein, OYEs are used as biocatalysts for decades to reduce activated C=C bonds in α,β-unsaturated carbonyl compounds. The promiscuity
of the enzymatic catalysis is advantageous for industrial applications.
However, the physiological function of OYEs, especially in bacteria, is still puzzling.
Moreover, the relevance of the OYEs in infection conditions remained enigmatic.
Here, I show that there are two groups of OYEs (OYE flavin oxidoreductase, OfrA and OfrB) that are encoded in staphylococci and some firmicutes. OfrA (SAUSA300_0859) is more conserved than OfrB (SAUSA300_0322) in staphylococci and is a part of the staphylococcal core genome.
A reporter system was established to report for ofrA in S. aureus background.
The results showed that ofrA is induced under electrophilic, oxidative, and hypochlorite stress. OfrA protects S. aureus against quinone, methylglyoxal, hydrogen peroxide,
and hypochlorite stress. Additionally, the results provide evidence that OfrA supports
thiol-dependent redox homeostasis. At the host-pathogen interface, OfrA promotes S. aureus fitness in murine macrophage cell line. In whole human blood, OfrA is involved in S. aureus survival indicating a potential clinical relevance to bacteraemia.
In addition, ofrA mutation affects the production of the virulence factor staphyloxanthin via the upper mevalonate pathway. In summary, decoding OfrA function and its proposed mechanism of action in S. aureus shed the light on a conserved stress response within multiple organisms.