Due to the limitations of silicon (Si), there is a paradigm shift in research focusing on wide-bandgap-based (WBG) materials. SiC power semiconductors exhibit superiority in terms of switching speed, higher breakdown electric field, and high working temperature, slowly becoming a global solution in harsh medium-voltage (MV) high-power environments. However, to utilize the SiC MOSFET device to achieve those next-generation, high-density, high-efficiency power electronics converters, one must solve a plethora of challenges. For the MV SiC MOSFET device, a high-performance gate-driver (GD) is a key component required to maximize the beneficial SiC MOSFET characteristics. GD units must overcome associated challenges of electro-magnetic interference (EMI) with regards to common-mode (CM) currents and cross-talk, low driving loop inductance required for fast switching, and device short-circuit (SC) protection. Developed GDs (for 1.2 kV, and 10 kV devices) are able to sustain dv/dt higher than 100 V/ns, have less than 5 nH gate loop inductance, and SC protection, turning off the device within 1.5 us. Even with the introduction of SiC MOSFETs, power devices remain the most reliability-critical component in the converter, due to large junction temperature (Tj) fluctuations causing accelerated wear-out. Real-time (online) measurement of the Tj can help improve long-term reliability by enabling active thermal control, monitoring, and prognostics. An online Tj estimation is accomplished by generating integrated intelligence on the GD level. The developed Tj sensor exhibits a maximum error less than 5 degrees Celsius, having excellent repeatability of 1.2 degrees Celsius. Additionally, degradation monitoring and an aging compensation scheme are discussed, in order to maintain the accuracy of the sensor throughout the device's lifetime. Since ultra high-voltage SiC MOSFET devices (20 kV) are impractical, the modular multilevel converter (MMC) emerged as a prospective topology to achieve MV power conversion. If the kernal part of the power-cell (main constitutive part of the MMC converter) is an SiC MOSFET, the design is able to achieve very high-density and high-efficiency. To ensure a successful operation of the power-cell, a systematic design and assessment methodology (DAM) is explored, based on the 10 kV SiC MOSFET power-cell. It simultaneously addresses challenges of high-voltage insulation, high dv/dt and EMI, component and system protections, as well as thermal management. The developed power-cell achieved high-power density of 11.9 kW/l, with measured peak efficiency of n=99.3 %@10 kHz. It successfully operated at Vdc=6 kV, I=84 A, fsw>5 kHz, Tj<150 degrees Celsius and had high switching speeds over 100 V/ns. Lastly, to achieve high-power density and high-efficiency on the MV converter level, challenges of high-voltage insulation, high-bandwidth control, EMI, and thermal management must be solved. Novel switching cycle control (SCC) and integrated capacitor blocked-transistor (ICBT) control methodologies were developed, overcoming the drawbacks of conventional MMC control. These novel types of control enable extreme reduction in passive component size, increase the efficiency, and can operate in dc/dc, dc/ac, mode, potentially opening the modular converter to applications in which it was not previously used. In order to explore the aforementioned benefits, a modular, scalable, 2-cell per arm, prototype MV converter based on the developed power-cell is constructed. The converter successfully operated at Vdc=12 kV, I=28 A, fsw=10 kHz, with high switching speeds, exhibiting high transient immunity in both SCC and ICBT. / Doctor of Philosophy / In medium-voltage applications, such as an electric grid interface in highly populated areas, a ship dc system, a motor drive, renewable energy, etc., land and space can be very limited and expensive. This requires the attributes of high-density, high-efficiency, and reliable distribution by a power electronics converter, whose central piece is the semiconductor device. With the recent breakthrough of SiC devices, these characteristics are obtainable, due to SiC inherent superiority over conventional Si devices. However, to achieve them, several challenges must be overcome and are tackled by this dissertation. Firstly, as a key component required to maximize the beneficial SiC MOSFET characteristics, it is of utmost importance that the high-performance gate-driver be immune to interference issues caused by fast switching and be able to protect the device against a short-circuit, thus increasing the reliability of the system. Secondly, to prevent accelerated degradation of the semiconductor devices due to high-temperature fluctuations, real-time (online) measurement of the Tj is developed on the gate-driver to help improve long-term reliability. Thirdly, to achieve medium-voltage high-power density, high-efficiency modular power conversion, a converter block (power-cell) is developed that simultaneously addresses the challenges of high-voltage insulation, high interference, component and system protections, and thermal management. Lastly, a full-scale medium-voltage modular converter is developed, exploiting the advantages of the fast commutation speed and high switching frequency offered by SiC, meanwhile exhibiting exceptional power density and efficiency.

Junction flows originate from the interaction between a fluid moving over a wall with an obstacle mounted on the same surface. Understanding the physics of such flows is of great interest to engineers responsible for the design of systems consisting of wall-body junctions. From aerodynamics to turbomachinery and electronics to bridge hydraulics, a number of phenomena (drag, heat transfer, scouring) are driven by the behavior of the most prominent feature of junction flows: the horseshoe vortex system (HVS). Focusing on turbulent flows, the complex dynamics of the HVS is established through its unsteadiness and non-uniformity. The fundamentals of this dynamically-rich phenomenon have been described within the body of a rapidly-expanding literature. Nevertheless, important aspects remain inadequately understood and call for further scrutiny. This study emphasized three of them, by investigating the effects of: model scale, wall roughness, and bed geometry. High-resolution experiments were carried out using Particle Image Velocimetry (PIV). Statistical analyses, vortex identification schemes, and Proper Orthogonal decomposition were employed to extract additional information from the large PIV datasets. The time-averaged topology of junction flows developing over a smooth and impermeable wall was independent of the flow Reynolds number, Re (parameter that expresses the effects of scale). On the contrary, time-resolved analysis revealed a trend of increasing vorticity, momentum, and eruptions of near-wall fluid with Re. New insights on the modal dynamics of the HVS were also documented in a modified flow mechanism. Wall roughness (modeled with a permeable layer of crushed stones) diffused turbulence and vorticity throughout the domain. This effect manifested with high levels of intermittency and spatial irregularity for the HVS. Energetic flow structures were also identified away from the typical footprint of the HVS. Finally, a novel implementation of PIV allowed for unique velocity measurements over an erodible bed. It was demonstrated that, during the initial stages of scouring, the downflow at the face of the obstacle becomes the dominant flow characteristic in the absence of the HVS. Notwithstanding modeling limitations, the physical insight contributed here could be used to enhance the design of systems with similar flow and geometrical characteristics. / Ph. D.

Curdlan, a (1,3)-linked linear homopolysaccharide composed of beta-D-glucan, is produced by the bacterium Alcaligenes faecalis var. myxogenes. Several strategies to synthesize chemically modified curdlan derivatives have been reported, but there have been few reports of regioselective functionalization at specific positions of the curdlan backbone, especially of aminated curdlan derivatives which have remarkable potential in biomedical and pharmaceutical applications. We demonstrate herein the design, synthesis and characterization of a family of regioselectively aminated curdlan derivatives including 6-deoxy-6-(bromo/azido/amino/amido/ammonium) curdlans starting from 6-bromo/azido-6-deoxycurdlan. A key reaction that enabled the whole synthesis of new curdlan derivatives at C-6 described in this dissertation was the highly selective bromination of curdlan. The resultant 6-bromo-6-deoxycurdlan, prepared with high regioselectivity, was treated with trialkylamines or heterocyclic amines to produce a range of water-soluble curdlan ammonium salts. The bromide was then nucleophilically displaced by sodium azide to produce the versatile precursor 6-azido-6-deoxycurdlan. Its water solubility was enhanced either by the incorporation of hydrophilic trioxadecanoate esters into O-2/4 positions or by the borohydride reduction to afford 6-amino-6-deoxycurdlan. The iminophosphorane intermediate generated during Staudinger reactions was further investigated for subsequent syntheses: i) 6-amino or 6-amido-6-deoxycurdlan by in situ reaction with water or excess carboxylic anhydride, ii) 6-monoalkylamino curdlan by reductive amination using aldehydes and sodium cyanoborohydride, and iii) 6-dialkylamino-/tri-alkylammoniocurdlans by reacting with methyl iodide. Such derivatives could have properties useful for a range of biomedical applications, including interactions with proteins, encapsulation of drugs, and formulation with genes or other biological compounds. / Ph. D.

No / Transposable elements (TEs), such as endogenous retroviruses (ERVs), are common in the genomes of vertebrates. ERVs result from retroviral infections of germ-line cells, and once integrated into host DNA they become part of the host's heritable genetic material. ERVs have been ascribed positive effects on host physiology such as the generation of novel, adaptive genetic variation and resistance to infection, as well as negative effects as agents of tumorigenesis and disease. The avian leukosis virus subgroup E family (ALVE) of endogenous viruses of chickens has been used as a model system for studying the effects of ERVs on host physiology, and approximately 30 distinct ALVE proviruses have been described in the Gallus gallus genome. In this report we describe the development of a software tool, which we call Vermillion, and the use of this tool in combination with targeted next-generation sequencing (NGS) to increase the number of known proviruses belonging to the ALVE family of ERVs in the chicken genome by 4-fold, including expanding the number of known ALVE elements on chromosome 1 (Gga1) from the current 9 to a total of 40. Although we focused on the discovery of ALVE elements in chickens, with appropriate selection of target sequences Vermillion can be used to develop profiles of other families of ERVs and TEs in chickens as well as in species other than the chicken. / Financial support was provided by the EW GROUP, as well as grants from the Canada Foundation for Innovation, Canada Research Chairs Program, and the Natural Sciences and Engineering Council of Canada to RGB, and Canada Institutes of Health Research funding to MGIL and CJM.

Eine einzigartige experimentelle Beobachtung, welche die Basis für eine ganzheitliche, neurowissentschafliche Theorie für Gedächtnis darstellen könnte, sind sharp wave-ripples (SWRs). SWRs werden in lokalen Neuronennetzwerken erzeugt und sind wichtig für Gedächtniskonsolidierung; SWRs sind charakteristische Ereignisse der lokalen Feldpotentiale im Hippocampus des Säugetiers, die in Phasen von Schlaf und Ruhe vorkommen. Eine SWR besteht aus einer sharp wave, einer ≈ 100 ms langen Auslenkung des Feldpotentials, welche mit ripples, 110–250 Hz Oszillationen, überlagert ist. Jüngste Experimente bekräftigen die Theorie, dass ripples in Netzwerken inhibitorischer Interneurone (INT-INT) erzeugt werden, die aus parvalbumin-positive basket cells (PV+BCs) bestehen. PV+BCs sind untereinander über rekurrente inhibitorische Synapsen und Gap Junctions (GJs) gekoppelt. In dieser Arbeit untersuche ich die spezifische Funktion von interneuronalen Gap Junctions in ripples. Im Hauptteil dieser Arbeit demonstriere ich, dass GJs in INT-INT Netzwerken die neuronale Synchronität und die Feuerrate während ripples erhöhen, die ripple-Frequenz sich hingegen nur leicht verändert. Zusätzlich zeige ich, dass diese rippleunterstützenden Effekte nur dann auftreten, wenn die GJ-Transmission schnell genug ist (≈< 0.5 ms), was wiederum somanahe Kopplung voraussetzt (≈< 100 µm). Darüber hinaus zeige ich, dass GJs die oszillatorische Stärke der ripples erhöhen und so die minimale für ripples notwendige Netzwerkgröße verringern. Abschließend zeige ich, dass ausschließlich mit Gap Junctions gekoppelte INT-INT Netzwerke zwar mit ripple Frequenz oszillieren können, aber wahrscheinlich nicht der Erzeuger von experimentell beobachteten ripple-artigen Oszillationen sind. Zusammengenommen zeigen meine Resultate, dass schnelle Gap Junction-Kopplung von Interneuronen die Entstehung von ripples begünstigt und somit SWRs unterstützt, welche einen wichtigen Beitrag zur Bildung unserers Gedächtnisses leisten. / A unique experimental observation that opens ways for a holistic, bottom-up theory for memory generation are sharp-wave ripples (SWRs). SWRs are generated in local neuronal networks and are important for memory consolidation. SWRs are prominent features of the extracellular field potentials in the mammalian hippocampus that occur during rest and sleep; they are characterized by sharp waves, ≈ 100 ms long voltage deflections, that are accompanied by ripples, i.e., 110–250 Hz oscillations. Recent experiments support the view that ripples are clocked by recurrent networks of inhibitory interneurons (INT-INT), which are likely constituted by networks of parvalbumin-positive basket cells (PV+BCs). PV+BCs are not only recurrently coupled by inhibition but also by gap junctions (GJs). In this thesis, I investigate the specific function of interneuronal GJs in hippocampal ripples. Consequently, I simulate INT-INT networks and demonstrate that gap junctions increase the neuronal synchrony and firing rates during ripple oscillations, while the ripple frequency is only affected mildly. I further show that GJs only have these supporting effects on ripples when they are sufficiently fast (≈< 0.5 ms), which requires proximal GJ coupling (≈< 100 µm). Additionally, I find that gap junctions increase the oscillatory power of ripple oscillations and by this means reduce the minimal network size required for INT-INT networks to generate ripple oscillations. Finally, I demonstrate that exclusively GJ-coupled INT-INT networks can oscillate at ripple frequency, however, are unlikely the generator of experimentally observed ripple-like oscillations. In sum, my results show that fast interneuronal gap junction coupling promotes the emergence of ripples and hereby supports SWRs, which are important for the formation of memory.

Die Tight Junction (TJ) reguliert den parazellulären Transport von Ionen, Wasser und Soluten an Epithelien und Endothelien und ist von entscheidender Bedeutung für die Aufrechterhaltung der Funktion von Organen und Geweben. Obwohl Claudin-3 zu den zuerst identifizierten und ubiquitär exprimierten Komponenten der TJ gehört, konnte seine spezifische Funktion bislang nicht geklärt werden. Für die funktionelle Charakterisierung des humanen Claudin-3-Proteins wurden stabile Überexpressionsklone der lecken Nierenepithel-Zelllinie MDCK II generiert. Die Überexpression von Claudin-3 führte zu einer deutlichen Änderung des TJ-Strangmusters sowie zu einer starken Zunahme des transepithelialen Widerstandes und einer verminderten Permeabilität für Ionen und Moleküle der Größe 332 Da und 4000 Da. Der parazelluläre Durchtritt von Wasser war unverändert. Claudin-3 konnte eindeutig als abdichtende Komponente der TJ identifiziert werden. Anhand des endothelialen Zellkulturmodells HUVEC wurden Expression und Regulation von Claudin-3 und anderen TJ-Proteinen unter dem Einfluss mechanischer Strömungsverhältnisse und des Sauerstoffpartialdrucks analysiert. Die Behandlung mit fehlender Wandschubspannung führte zur Hochregulation der abdichtenden TJ-Proteine Occludin, Claudin-3, Claudin-5 und Claudin-11, nicht aber Claudin-23. Die Regulation der einzelnen TJ-Komponenten wurde durch unterschiedliche Signalwege vermittelt, wobei der verstärkten Proteinexpression jeweils eine Hochregulation auf mRNA-Ebene zugrunde lag. Die kombinierte Behandlung mit fehlender Wandschubspannung und Hypoxie resultierte in einer sehr stark erhöhten Expression von Claudin-3. Durch die Hochregulation abdichtender TJ-Komponenten unter Bedingungen fehlender Wandschubspannung und Hypoxie, wie sie in verschiedenen physiologischen und pathologischen Situationen auftreten, könnte einem unerwünschten Durchtritt von Substanzen aus dem Blut in das umliegende Gewebe vorgebeugt werden. / The tight junction (TJ) regulates the paracellular transport of ions, water and solutes in epithelia and endothelia and is of particular importance for a correct function of organs and tissues. Although claudin-3 is one of the first identified and ubiquitously expressed TJ components, its specific function was unsolved as yet. For functional characterization, human claudin-3 was stably overexpressed in the leaky epithelial cell line MDCK II. Overexpression of claudin-3 led to a marked alteration of TJ meshwork pattern, a strong increase in transepithelial resistance and a decrease in permeability for ions and paracellular tracers (332 or 4000 Da). Paracellular water transport was not affected. It was proved that claudin-3 acts as a „tightening“ TJ component. The endothelial cell culture model HUVEC was used for analysis of expression and regulation of claudin-3 and several other TJ proteins under different conditions of wall shear stress and oxygen saturation. Treatment with lacking wall shear stress led to an upregulation of the “tightening” TJ proteins occludin, claudin-3, claudin-5, and claudin-11, but not claudin-23. Upregulation of all proteins was due to increased mRNA levels. Apparently, different signaling pathways were involved in regulation of particular TJ components. Combined treatment with lacking shear stress and hypoxia resulted in drastically increased claudin-3 expression. Upregulation of tightening TJ components under lacking shear stress and hypoxic conditions as occuring in different physiological or pathological situations would limit the passage of solutes from the blood into the surrounding tissue.

Le syndrome de Hutchinson-Gilford, ou progéria, est une pathologie génétique rare qui se caractérise par des symptômes assimilés à un vieillissement prématuré. Les mutations à l'origine de la progéria affectent le gène LMNA, codant la lamine A, qui joue un rôle majeur dans la formation, la maintenance et la résistance du noyau. Ces mutations activent l'utilisation de sites 5' alternatif ou cryptique d'épissage présents dans l'exon 11 du pré-ARNm LMNA en amont du site normalement utilisé. Nous avons révélé un effet des mutations sur la structure secondaire de l'ARN aux alentours des mutations, qui permet l'augmentation de l'utilisation des sites d'épissage mutants. De plus, nous avons montré l'implication de plusieurs protéines SR (SRSF1, SRSF5 et SRSF6) dans la régulation de l'utilisation des différents sites d'épissage. D'autre part, il a déjà été observé que les noyaux des cellules des patients atteints de progéria contiennent des granules de stress, les nSB, situés dans les régions péricentromériques des chromosomes et contenant des ARN dits satellite III et des facteurs d'épissage. Des nSB similaires sont formés dans les cellules saines suite à divers stress, comme le stress thermique. Il est possible que ces nSB séquestrent ces facteurs d'épissage afin de réguler le profil d'épissage alternatif des cellules pendant la régénération après un stress. Nous avons purifié les protéines associées aux ARN satellite III in vitro afin de trouver de nouveaux composants des nSB et analysé, par emploi de puces jonction-exon, le transcriptome de cellules soumises à un choc thermique, pour mieux comprendre à terme comment la formation des nSB peut affecter l'épissage alternatif / The Hutchinson-Gilford syndrome, also called progeria, is a rare genetic disease, characterized by symptoms that can be assimilated to accelerated natural ageing. Mutations that cause progeria affect the LMNA gene, which codes the lamin A that plays a major role in the shaping, maintenance and resistance of the nucleus. These mutations lead to the activation of alternative or cryptic 5' splice sites located within the exon 11 of LMNA pre-mRNA upstream from the normal 5' splice site. Our work revealed an effect of the mutations on the 2D RNA structure of the splice sites, which contributes to the increased use of the mutant sites. On top of it, we showed the impact of several SR proteins, (SRSF1, SRSF5 and SRSF6) on the regulation of the use of the exon 11 5' splice sites. On the other hand, it was previously observed that cells from progeria patients contain nuclear stress bodies (nSB), located in chromosomal pericentromeric regions and containing satellite III RNAs and several splicing regulatory proteins. Similar bodies are formed in healthy cells submitted to various stresses such as heat shock. A work hypothesis is that those nSBs sequester splicing factors in order to regulate the global alternative splicing profile in cells during the recovery period after stress. We purified proteins associated with satellite III RNAs in vitro, to find new components of the nSBs, and analyzed the transcriptome of cells subjected to heat shock using exon junction microarrays, in order to eventually understand how nSB formation can affect alternative splicing

Die tight junction schränken die Diffusion durch den parazellulären Raum in Epithel- und Endothelzellschichten für viele Moleküle stark ein. Dadurch behindern sie die Aufnahme von wasserlöslichen Medikamenten in das dahinterliegende Gewebe. Zwei Proteine, die am tight junction Aufbau mitwirken, sind Zonula Occludens Protein 1 (ZO-1) und Occludin. Eine Öffnung der tight junctions stellt eine Möglichkeit für die Verabreichung von Medikamenten dar. Deshalb wurden die tight junction Proteine ZO-1 und Occludin auf ihre Funktion, Struktur und Regulation untersucht. Für die Interaktion beider Proteine gab es ein Modell, welches eine Oligomerisierung der Bindungspartner als Voraussetzung ihrer Interaktion über helikale Wechselwirkungen vorhersagte. Die Annahmen aus dem Modell der Interaktion von ZO-1 und Occludin konnten experimentell bestätigt werden. Für den C-Terminus von Occludin wurde darüber hinaus eine Interaktion über Disulfidbrücken nachgewiesen. Diese Interaktion könnte in der Zelle von pathologischer Bedeutung bei Schlaganfall und Ischchämie sein. Beide Erkrankungen verursachen eine Öffnung der tight junction im Zusammenhang mit oxidativem Stress. ZO-1 bindet über PDZ Domänen eine Vielzahl von tight junction Proteinen, die an der Abdichtung des parazellulären Raums beteiligt sind. Deshalb wurde die Interaktion und Regulation der PDZ-Domänen aus ZO-1 untersucht. Eine Phosphorylierung der PDZ durch die Proteinkinase C alpha sowie eine Interaktion mit den Phosphatasen 2A und 4 konnte nachgewiesen werden. In vitro konnte gezeigt werden, dass die Phosphorylierung der PDZ-Domänen die Bindung an Membranproteine der tight junction beeinflusst. Diese Arbeit leistet einen Beitrag, die Mechanismen, die zum Verschluss des parazellulären Spaltes führen, aufzuklären. Damit zeigt sie Ansatzpunkte für eine pharmakologische Beeinflussung der Permeabilität der tight junction auf. / Tight junctions restrict diffusion through the paracellular gap in endothelia and epithelia. Thereby they constrain the uptake of water soluble drugs to the tissue. Zonula occludens protein 1 (ZO-1) and occludin are some of proteins involved in tight junction assembly. The opening of tight junctions is a possibility to apply drugs. Therefore the structure, function and regulation of ZO-1 and occludin is characterised. In previous studies, a model predicted the interaction of occludin and ZO-1 through helices. It was proposed that the interaction is mediated by oligomers of ZO-1 and Occludin. This author´s experimental research supports these hypotheses. Furthermore, occludin is shown to self assemble via disulfide bridges. This interaction could be of importance during stroke and ischemia. Both diseases cause the opening of tight junctions in combination with oxidative stress. In addition, this author investigated the interaction and regulation of the PDZ domains of ZO-1. It was shown that the PDZ domains are phosphorylated by protein kinase C alpha and interact with protein phosphatases 2A and 4. Phosphorylation led to a reduction in affinity of PDZ to membrane proteins in vitro. This thesis contributes to the understanding of the mechanisms which are involved in the sealing of the paracellular gap.

No description available.

Da vorliegende Studien unserer Arbeitsgruppe auf eine Beteiligung von Connexinen an der oralen Plattenepithelkarzinogenese hindeuten, war es Ziel dieser Arbeit in-vivo am Hamstermodell die Expression der Connexine 26, 43 und 45 im Verlauf der Karzinogenese zu untersuchen. Die durch 9,10-Dimethyl-1,2-Benzanthrazen (DMBA) induzierten Karzinome wurden makroskopisch, histologisch, sowie auf ihren Inflammationsgrad hin untersucht, um die Effektivität des Tiermodells zu prüfen. Weiterhin erfolgte sie Genexpressionsanalyse mittels RNA-Isolation, PCR-Analyse und statistischer Auswertung. Es konnte insgesamt gezeigt werden, dass die Applikation von DMBA in Abhängigkeit von Behandlungsdauer makroskopisch, histologisch und inflammatorisch einen deutlichen karzinogenen Effekt hatte und, dass das histopathologische Grading signifikant mit der Länge der Behandlungsdauer korrelierte. Im Rahmen der Genexpressionsanalyse konnte im Verlauf der DMBA-Behandlung für Connexin 26 und 45 eine signifikante Überexpression beschrieben werden. Connexin 43 zeigte sich als nicht differentiell exprimiert. Ein Zusammenhang zwischen histologischem Grading und Genexpression war nicht nachvollziehbar.