Development of a three dimensional (3D) HYBROT model with targeted in vivo like intact cellular circuitry in thick brain slices for multi-site stimulation and recording will provide a useful in vitro model to study neuronal dynamics at network level. In order to make this in vitro model feasible, we need to develop several associated technologies. These technologies include development of a thick organotypic brain slice culturing method, a three dimensional (3D) micro-fluidic multielectrode Neural Interface system (µNIS) and the associated electronic interfaces for stimulation and recording of/from tissue, development of targeted stimulation patterns for closed-loop interaction with a robotic body, and a deep-tissue non-invasive imaging system. To make progress towards this goal, I undertook two projects: (i) to develop a method to culture thick organotypic brain slices, and (ii) construct a multiphoton imaging system that allows long-term and deep-tissue imaging of two dimensional and three dimensional cultures. Organotypic brain slices preserve cytoarchitecture of the brain. Therefore, they make more a realistic reduced model for various network level investigations. However, current culturing methods are not successful for culturing thick brain slices due to limited supply of nutrients and oxygen to inner layers of the culture. We developed a forced-convection based perfusion method to culture viable 700µm thick brain slices. Multiphoton microscopy is ideal for imaging living 2D or 3D cultures at submicron resolution. We successfully fabricated a custom-designed high efficiency multiphoton microscope that has the desired flexibility to perform experiments using multiple technologies simultaneously. This microscope was used successfully for 3D and time-lapse imaging. Together these projects have contributed towards the progress of development of a 3D HYBROT. ----- 3D Hybrot: A hybrid system of a brain slice culture embodied with a robotic body.

Magnetic Resonance Imaging (MRI) is considered as a slow imaging technique. Various approaches to accelerate MRI imaging have been explored by researchers in the past decades. Earlier gradient based methods have reached the safety limit. Parallel receiving techniques achieve accelerations by reducing phase encoding steps. Among these methods, SEA Imaging achieved the highest possible acceleration by completely eliminating phase encoding. However, SEA imaging is limited to thin planar slices above the array due to the correction needed for the inherent phase cancellation caused by voxel-sized coils. A phase compensation gradient pulse is used for this correction in SEA imaging. This phase compensation is dependent on slice position and thickness as well as the orientation of the array elements, placing stringent restrictions on SEA imaging, limiting its applications. Converting the SEA system into Transmit / Receive (T/R) mode, which is the main purpose of this study, eliminates the requirement for phase compensation gradient because phase departed during transmit is refocused during receiving. Independent amplitude and phase control of RF pulse for each coil of a SEA array is achieved using a low cost scalable parallel transmit system design. The first 64-channel parallel transmitter for MRI in the world is constructed and tested. Software is also developed to control the phase and amplitude of all the 64 channels of RF excitation pulses independently through National Instruments DAQ system. The system consists of vector modulators controlled by digital controlled potentiometers, two-stage amplifiers and T/R switches on the transmit side. All these are combined with newly designed and constructed preamplifiers and the existing 64-channel parallel receivers on the receive side, leading to the only 64-channel parallel T/R system available for MRI. As a bonus, the system can be easily updated to full Transmit SENSE capability. Furthermore, simulations and images are done to synthesize transmit patterns thanks to the large channel count. Testing results show that the system is capable of 100W per channel simultaneous transmission. Using this system, transmit field can be synthesized by varying the phase and amplitude across channel without traditionally required complicated pulse sequences involving simultaneous RF and gradient fields. Curved slice excitation has conventionally been considered a difficult task for MRI, achievable only through complicated pulses sequences. Using this system and flexible array wrapped around the subject to be imaged, the system is able to excite curved slice using one shot. TR images indicate that the system is capable of high speed surface imaging at 200 frames per second following the surface of a flexible SEA array coil which has not been achieved using other methods in MRI.

The aim of this thesis was to investigate some methods for quantitative analysis of bone structure, particularly techniques which might ultimately be applied post-operatively following orthopaedic reconstruction operations. Initially it was decided to explore the efficacy of MRI in quantifying the bone structure at high resolution by comparing high resolution MRI against 'gold standards' such as Scanning Electron Microscopy (SEM) and optical histology. This basic study provided a measure of the distortions in the morphological bone parameters derived from MR images due to susceptibility artefacts and partial volume effects. The study of bone architecture was then extended to a model of advanced renal osteodystrophy in a growing rat. For this study, high-resolution micro computed tomography (microCT) was used and as a result of the high resolution images obtained, three new bone morphological parameters were introduced to characterise the bone structure. The desire to study bone architecture post-operatively in hip replacements led to a preliminary study on ex-vivo sheep acetabulae following total hip replacement, to determine the extent that the bone architecture could be investigated around the acetabulum. The motivation for studying the acetabulum was based on the high occurrence of debonding at the bone / prosthesis interface. This study demonstrated the superior nature of 3D MRI over conventional x-ray radiographs in early quantitation of fibrous membranes located between the host bone and the non-metallic implant and/or the bone cement. The presence of such fibrous membranes is strongly indicative of failure of the prosthesis. When using clinical MRI to image post-operative hip replacement, the image quality is severely affected by the presence of the metallic implant. The head of the prosthesis is shaped like a metal sphere and is located in the acetabular cup. This problem was investigated by performing simulations of MR images in the presence of the field perturbation induced by the presence of a metal sphere, with the effects of slice excitation and frequency encoding incorporated into the simulations. The simulations were compared with experimental data obtained by imaging a phantom comprising a stainless steel ball bearing immersed in agarose gel. The simulations were used to predict the effects of changing imaging parameters that influence artefact size and also to show how current metal artefact reduction techniques such as view angle tilting (VAT) work and to identify their limitations. It was shown that 2D SE and VAT imaging techniques should not be used when metallic prosthesis are present due to extreme slice distortion, whereas 3D MRI provided a method that has no slice distortion, although the effects of using a frequency encoding gradient still remain.

Diffusion-weighted (DW) magnetic resonance imaging is an important neuroimaging technique that has successful applications in diagnosis of ischemic stroke and methods based on diffusion tensor imaging (DTI). Tensor measures have been used for detecting changes in tissue microstructure and for non-invasively tracing white matter connections in vivo. The most common image acquistion strategy is to use a DW single-shot echo-planar imaging (ss-EPI) pulse sequence, which is attractive due to its robustness to motion artefacts and high imaging speed. However, this sequence has limited achievable spatial resolution and suffers from geometric distortion and blurring artefacts. Readout-segmented echo-planar imaging (rs-EPI) is a DW sequence that is capable of acquiring high-resolution images by segmenting the acquisition of k- space into multiple shots. The fast, short readouts reduce distortion and blurring and the problem of artefacts due to motion-induced phase changes between shots can be overcome with navigator techniques. The rs-EPI sequence has two main shortcomings. (i) The method is slow to produce image volumes, which is limiting for clinical scans due to patient welfare and prevents us from acquiring very many directions in DTI. (ii) The sequence (like other diffusion techniques) is far from the optimum repetition time (TR) for acquiring data with the highest possible signal-to-noise ratio (SNR) in a given time. The work in this thesis seeks to address both of these important issues using a range of approaches. In Chapter 4 a partial Fourier extension is presented, which addresses point (i) by reducing the number of readout segments acquired and estimating the missing data. This allows reductions in scan time by approximately 40% and the reliability of the images is demonstrated in comparisons with the original images. The application of a simultaneous multi-slice scheme to rs-EPI, to address points (i) and (ii), is described in Chapter 5. Using the slice-accelerated rs-EPI sequence, tractography data were compared to ss-EPI data and high-resolution trace-weighted data were acquired in clinically relevant scan times. Finally, a 3D multi-slab extension that addresses point (i) is presented in Chapter 6. A 3D sequence could also allow higher resolution in the slice direction than 2D multi-slice methods, which are limited by the difficulties in exciting thin, accurate slices. A 3D version of rs-EPI was simulated and implemented and a k-space acquisition synchronised to the cardiac cycle showed substantial improvements in image artefacts compared to a conventional k-space acquisition.

The aim of this thesis is to assess using selected marketing tools how Czech consumers perceive different formats of television advertisement used by Vodafone. To this end, Vodafone TV spots aired in the Czech Republic between 2005 and 2016 were analysed focusing mainly on TV spots aired in the past few years. An integral part of this thesis is the qualitative Focus Group research method and probe questionnaires distributed among Czech consumers. The results of this research are subsequently compared with the Multicriterial research MML TGI Median Company and also with the research results of the Nielsen Company.

La maladie d’Alzheimer (AD), pathologie neurodégénérative progressive, est caractérisée par des dépôts β-amyloïdes extracellulaires, des enchevêtrements neurofibrillaires intracellulaires de Tau et une dégénérescence neuronale. A travers les nombreux modèles transgéniques AD disponibles, les connaissances sur les peptides amyloïdes et la protéine Tau ne cessent de progresser. Mais contrairement aux cas génétiques, l’étiologie des cas sporadiques d’AD reste à ce jour idiopathique, rendant difficile d’établir une stratégie thérapeutique efficace. Au cours d’une étude sur l’implication des protéines kinases dans la pathogénèse d’AD, des collaborateurs ont fait une observation totalement inattendue, mais très intéressante: une molécule de faible poids moléculaire, serait capable d’induire une production spécifique d’Aβ1-42 sans altérer les niveaux d’Aβ1-40 dans un modèle de lignée cellulaire. Dans ce contexte, le projet de thèse portait sur l’utilisation de dérivé purique (PD1) pour développer des modèles AD induits chimiquement sur différents supports (culture primaire de neurones, culture organotypique d’hippocampe et souris) et en investiguer les mécanismes sous-jacents à l’augmentation des peptides A1-42 (issus du métabolisme de l’APP (Amyloid precursor protein)).La première partie du projet de thèse a permis de mettre en évidence dans un contexte in vitro (culture primaire de neurones et culture organotypique d’hippocampe) que PD1 à forte dose induisait une augmentation du ratio Aβ42/40 et de manière répétable. Fort de ces résultats, nous avons voulu étudier les mécanismes d’action de PD1 autour de deux hypothèses : interaction dans le métabolisme de l’APP et implication des cellules gliales. Contrairement à nos premières hypothèses, nous avons montré que PD1 aurait de potentiels effets anti-inflammatoires (i.e. IL-1β) in vitro et in vivo. La voie de signalisation de l’IL-1β étant de plus en plus incriminée dans la pathogenèse d’Alzheimer; nous nous sommes interrogés sur l’effet dual de PD1 : outil pharmacologique alzheimerigène ou candidat médicament pour le traitement d’AD? / Alzheimer’s disease (AD), a progressive neurodegenerative disorder, appears to be associated with an increase in a particular form of β-amyloid deposits, intracellular Tau tangles and neuronal degeneration. Through many available transgenic AD models, knowledge about amyloid peptides and Tau protein continues to increase. However, in contrast to the genetic cases of AD, the etiology of sporadic AD cases remains unknown, making the establishment of an effective therapeutic strategy difficult.During the course of a study on the role of protein kinase involved in AD, our collaborators made an unexpected but very interesting observation. They identified a low molecular weight compound able to induce production of Aβ1-42 while the level of the much less toxic form Aβ1-40 remained constant. This selective induction of Aβ1-42 versus Aβ1-40 was observed in a cell line model. Therefore, the overall goal of the project thesis was based on the use of purine derivative (PD1) to understand the molecular mechanisms underlying the selective production of Aβ1-42. This would allow us to establish cellular assays and a chemically-induced animal AD model relevant to studies on the treatment and prevention of AD.The first part of this project allowed us to demonstrate in vitro that PD1, at high dose, repeatedly induced an increase in Aβ42/40 ratio in primary neurons and in neuronal hippocampal slice culture (OHSCs). Based on these facts, we analyzed the amyloid profile by focusing on APP metabolism and on glial cell activity. In contrary to our hypothesis, we highlighted whether PD1 exhibits potential anti-inflammatory properties (i.e. IL-1β) both in vitro and in vivo. The IL-1β pathway is more and more linked in the AD pathogen which leads us to consider that PD1 could have a dual effect : alzheimerogenic pharmacological tool or potential drug candidate for the treatment of AD ?

This master thesis deals with calculations and simulations of multi-speed induction motors. In the first part, basic principle of these machines is described. Next, there are given three examples of pole-changing stator windings and their properties are analyzed (winding factors, magnetomotive force). Main part includes simulations and measuring of the specific motor, which is produced by company Siemens Mohelnice. This is the motor with 2/1 pole-changing in Y/YY connection. First, influence of skewed rotor slots on current and torque is analyzed. Next, parametres of motor from simulations are given and these results are compared to measured values.

This master´s thesis deals with of human factor impact on optical fibers manipulation. It describes types and methods used for splicing and placing optical fibers and optical cables. These methods are specified in the chapters; their application in different environments and conditions is accentuated. This paper presents splicing optical fibers. It emphasizes problems which can occur if proper procedures aren´t kept or if improper device is used. In the thesis there is the chapter dealing with problems which can arise during splicing optical fibers - these negative influences are evaluated with educational equipments EF-970-01 PLASTIC by the company MIKROKOM, s.r.o. Another aim of the work is to describe mechanical optical fiber splice 3MTM FibrlokTM. This kind of device was lent for testing optical fibers multiple splicing. It was tested under extreme conditions. Statistically processed values of insertion loss and splice reflectance, measured by OTDR are the results of the test. Comparing measured values, technical report ones, regarding for usage in practice, there is a possibility of 75 multiple reusing of the mechanical optical fiber splice. The prediction of magnitude insertion loss, calculated for thousands splice cycles follows. The next part of the work deals with fusion splices and contains comparison of splices made with two cleavers: CORNING LWL-TRENNGERAT S46999-M9-D12 and the later model - Fitel S325A cleaver. The result is that older S46999-M9-D12 model gained lower attenuation. The results of both previous measurements are influenced by human factor. It appears at mechanical optical fiber splices as growing fluctuate of insertion losses and as the unequal attenuation fusion splices at tested cleavers.

Gastric and esophagogastric junction cancers are heterogeneous and aggressive tumors with an unpredictable response to cytotoxic treatment. New methods allowing for the analysis of drug resistance are needed. Here, we describe a novel technique by which human tumor specimens can be cultured ex vivo, preserving parts of the natural cancer microenvironment. Using a tissue chop- per, fresh surgical tissue samples were cut in 400 μm slices and cultivated in 6-well plates for up to 6 days. The slices were processed for routine histopa- thology and immunohistochemistry. Cytokeratin stains (CK8, AE1/3) were ap- plied for determining tumor cellularity, Ki-67 for proliferation, and cleaved caspase-3 staining for apoptosis. The slices were analyzed under naive conditions and following 2–4 days in vitro exposure to 5-FU and cisplatin. The slice culture technology allowed for a good preservation of tissue morphology and tumor cell integrity during the culture period. After chemotherapy exposure, a loss of tumor cellularity and an increase in apoptosis were observed. Drug sensitivity of the tumors could be assessed. Organotypic slice cultures of gastric and es- ophagogastric junction cancers were successfully established. Cytotoxic drug effects could be monitored. They may be used to examine mechanisms of drug resistance in human tissue and may provide a unique and powerful ex vivo platform for the prediction of treatment response.

Glioblastoma multiforme (GBM) ist der nach WHO am gefährlichsten eingestufte Hirntumor astrozytären Ursprungs. Patienten versterben ohne Behandlung etwa drei bis sechs Monaten nach Diagnose, die derzeitig modernste Behandlung mit Chemo-Radiotherapie verlängert das mediane Überleben auf 12-15 Monate. Trotz intensiver Forschung gibt es zurzeit keine realistische Heilungschance. Bislang erfolgt der Großteil der Forschung an Zellkulturen oder mit Hilfe von Tiermodellen, bei denen ein Tumor künstlich erzeugt wird. Dabei ergeben sich Probleme für die Übertragung der Ergebnisse auf den Menschen. Zellkulturen werden z.B. als sogenannte Monolayer-Kulturen gehalten, was bedeutet, dass ihnen der natürliche Gewebeverband und die für Signalling-Wege wichtige extrazelluläre Matrix fehlen. Außerdem werden solche Langzeitkulturen häufig subkultiviert und mutieren dadurch in Richtung einer klonalen Linie, was zwar Ergebnisse leichter reproduzierbar macht, aber nicht die Situation im Patienten widerspiegelt. Tierversuche implizieren zwar den Gewebeverband im Körper, jedoch müssen die dafür verwendeten Nager immunsupprimiert sein, so dass sie den induzierten Tumor nicht abstoßen. Dies erzeugt wiederum ein künstliches Umfeld. In diesem Projekt wird untersucht, ob sich humane GBM-Gewebe als sogenannte Slice-Kultur halten lassen und als Testsysteme zur Untersuchung der Wirkung von Chemotherapeutika sowie Bestrahlung geeignet sind. Bei dieser Kultivierungsmethode wird das Gewebe in Scheiben (Slices) geschnitten, wobei alle Zellen im Verband sowie die 3D-Struktur erhalten bleiben. Wegen des humanen Ursprungs entfällt das Problem des Speziesunterschiedes. Das Gewebe wird direkt aus dem Operationssaal ins Labor transferiert und weiterverarbeitet. Wir konnten bislang zeigen, dass Slice-Kulturen von humanem GBM über mindestens zwei Wochen in Kultur vital bleiben und ihre ursprüngliche charakteristische Morphologie beibehalten. Etablierte Behandlungsmethoden wie die Gabe von Temozolomid oder Röntgenbestrahlung zeigen auch in kultivierten Slices bekannte Effekte wie Induktion von DNA-Doppelstrangbrüchen, Reduktion von Proliferation und Aktivierung des Apoptose-Enzyms Caspase 3. Eine neue Therapieoption besteht seit einigen Jahren in der Bestrahlung mit Kohlenstoffionen (12C), die an der GSI Helmholtzzentrum für Schwerionenforschung in Darmstadt entwickelt und getestet wurde. Derzeit wird diese Therapie sehr erfolgreich an soliden Tumoren im Kopf- und Halsbereich angewendet und soll nun auf weitere Tumorarten ausgedehnt werden. Eine Kooperation mit der dortigen Biophysik-Gruppe wurde initiiert, um humane GBM-Slices mit 12C zu bestrahlen. Bislang wurde das entsprechende Setup etabliert und erste Experimente durchgeführt. Die ersten Ergebnisse wurden kürzlich publiziert. Weiterhin soll nun geprüft werden, ob das Ansprechen der GBM Slice-Kulturen mit dem Überleben der Patienten korreliert bzw. ob resistente Kulturen aus Patienten stammten, die schlecht auf die Therapie reagierten. Außerdem sollen überlebende Zellen in den Slices nach Behandlung auf ihre molekularen Eigenschaften geprüft werden, um Hinweise auf die Mechanismen der Tumorresistenz zu erhalten. Langfristig könnten diese Slice-Kulturen genutzt werden, um neuartige Wirkstoffe in der Vorklinik zu prüfen oder eine optimierte, personalisierte Therapie für Patienten zu ermitteln.