The development of new therapeutical approaches, devised to foster the regeneration of neuronal circuits after injury and/or in neurodegenerative diseases, is of great importance. The impairment of dopaminergic projections is especially severe, because these projections are involved in crucial brain functions such as motor control, reward and cognition. In the work presented here, organotypic brain slice co-cultures of (a) the mesostriatal and (b) the mesocortical dopaminergic projection systems consisting of tissue sections of the ventral tegmental area/substantia nigra (VTA/SN), in combination with the target regions of (a) the striatum (STR) or (b) the prefrontal cortex (PFC), respectively, were used to evaluate different approaches to stimulate neurite outgrowth: (i) inhibition of cAMP/cGMP turnover with 3’,5’ cyclic nucleotide phosphodiesterase inhibitors (PDE-Is), (ii) blockade of calcium currents with nimodipine, and (iii) the co-cultivation with bone marrow-derived mesenchymal stromal/stem cells (BM-MSCs). The neurite growth-promoting properties of the tested substances and cell populations were analyzed by neurite density quantification in the border region between the two brain slices, using biocytin tracing or tyrosine hydroxylase labeling and automated image processing procedures. In addition, toxicological tests and gene expression analyses were conducted. (i) PDE-Is were applied to VTA/SN+STR rat co-cultures. The quantification of neurite density after both biocytin tracing and tyrosine hydroxylase labeling revealed a growth promoting effect of the PDE2A-Is BAY60-7550 and ND7001. The application of the PDE10-I MP-10 did not alter neurite density in comparison to the vehicle control. (ii) The effects of nimodipine were evaluated in VTA/SN+PFC rat co-cultures. A neurite growth-promoting effect of 0.1 µM and 1 µM nimodipine was demonstrated in a projection system of the CNS. In contrast, the application of 10 µM nimodipine did not alter neurite density, compared to the vehicle control, but induced the activation of the apoptosis marker caspase 3. The expression levels of the investigated genes, including Ca2+ binding proteins (Pvalb, S100b), immediate early genes (Arc, Egr1, Egr2, Egr4, Fos and JunB), glial fibrillary acidic protein, and myelin components (Mal, Mog, Plp1) were not significantly changed (with the exception of Egr4) by the treatment with 0.1 µM and 1 µM nimodipine. (iii) Bulk BM-MSCs that were classically isolated by plastic adhesion were compared to the subpopulation Sca-1+Lin-CD45--derived MSCs (SL45-MSCs). The neurite growth-promoting properties of both MSC populations were quantified in VTA/SN+PFC mouse co-cultures. For this purpose, the MSCs were seeded on glass slides that were placed underneath the co-cultures. A significantly enhanced neurite density within the co-cultures was induced by both bulk BM-MSCs and SL45-MSCs. SL45-MSCs increased neurite density to a higher degree. The characterization of both MSC populations revealed that the frequency of fibroblast colony forming units (CFU-f ) is 105-fold higher in SL45-MSCs. SL45-MSCs were morphologically more homogeneous and expressed higher levels of nestin, BDNF and FGF2 compared to bulk BM-MSCs. Thus, this work emphasizes the vast potential for molecular targeting with respect to the development of therapeutic strategies in the enhancement of neurite regrowth.

The entorhinal cortex (EC) in the medial temporal lobe plays a critical role in memory formation and is implicated in several neurological diseases including temporal lobe epilepsy and Alzheimer’s disease. Despite the known importance of this brain region, little is known about the normal bioelectrical activity patterns of the EC in awake, behaving primates. In order to develop effective therapies for diseases affecting the EC, we must first understand its normal properties. To contribute to our understanding of the EC, I monitored the activity of individual neurons and populations of neurons in the EC of rhesus macaque monkeys during free-viewing of photographs using electrophysiological techniques. The results of these experiments help to explain how primates can form memories of, and navigate through, the visual world. These experiments revealed neurons in the EC that represent visual space with triangular grid receptive fields and other neurons that prefer to fire near image borders. These properties are similar to those previously described in the rodent EC, but here the neuronal responses relate to viewing of remote space as opposed to representing the physical location of the animal. The representation of visual space may be aided by another EC neuron type that was discovered, free-viewing saccade direction cells, neurons that signaled the direction of upcoming saccades. Such a signal could be used by other cells to prepare to fire according to the future gaze location. Many of these spatially-responsive neurons also represented memory for images, suggesting that they may be useful for associating items with their locations. I also examined the neuronal circuitry of recognition memory for visual stimuli in the EC, and I found that population synchronization within the gamma-band (30-140 Hz) in superficial layers of the EC was modulated by stimulus novelty, while the strength of memory formation modulated gamma-band synchronization in the deep layers and in layer III. Furthermore, the strength of connectivity in the gamma-band between different layers was correlated with the strength of memory formation, with deep to superficial power transfer being correlated with stronger memory formation and superficial to deep transfer correlated with weaker memory formation. These findings support several previous investigations of hippocampal-entorhinal connectivity in the rodent and advance our understanding of the functional circuitry of the medial temporal lobe memory system. Finally, I explored the design of a device that could be used to investigate properties of brain tissue in vitro, potentially aiding in the development of treatments for disorders of the EC and other brain structures. We designed, fabricated, and validated a novel device for long-term maintenance of thick brain slices and 3-dimensional dissociated cell cultures on a perforated multi-electrode array. To date, most electrical recordings of thick tissue preparations have been performed by manually inserting electrode arrays. This work demonstrates a simple and effective solution to this problem by building a culture perfusion chamber around a planar perforated multi-electrode array. By making use of interstitial perfusion, the device maintained the thickness of tissue constructs and improved cellular survival as demonstrated by increased firing rates of perfused slices and 3-D cultures, compared to unperfused controls. To the best of our knowledge, this is the first thick tissue culture device to combine forced interstitial perfusion for long-term tissue maintenance and an integrated multi-electrode array for electrical recording and stimulation.

Die koronare Herzkrankheit (KHK) gehört zu den häufigsten Todesursachen in den westlichen Industrienationen. Die Diagnostik der Erkrankung hat somit großen Stellenwert in der Medizin. Akzeptierter Goldstandard zur Diagnostik einer KHK ist die Herzkatheteruntersuchung (HKU). Als nicht-invasive Alternative zur HKU hat sich in den letzten Jahren die Mehrzeilen-Computertomographie als zuverlässiges Verfahren für den KHK-Ausschluss bei mittlerer Vortestwahrscheinlichkeit etabliert. Ziel dieser Arbeit ist es, die diagnostischen Eigenschaften der prospektiv getriggerten mit der retrospektiv getriggerten CT-Koronarangiographie (CTCA) an einem 64-Zeilen Gerät in einem heterogenen Patientenkollektiv mit unterschiedlichen kardiovaskulären Erkrankungen (Verdacht auf Koronare Herzkrankheit, Aortenaneurysma, präoperativ zum Aortenklappenersatz oder zur Pulmonalvenenablation, zum Ausschluss eines Tumors oder Perikarditiden) in Genauigkeit, Bildqualität und ihrer Anwendbarkeit gegenüberzustellen und sie mit dem Referenzstandard, der HKU, zu vergleichen. In diese Studie wurden retrospektiv 77 Patienten eingeschlossen, die ein EKG-getriggertes kardiales CT erhielten. Wenn es möglich war, d.h. die Herzfrequenz <75/min, BMI <35 und ein Sinusrhythmus vorlag, wurde die prospektive EKG-getriggerte CTCA durchgeführt, alternativ kam die retrospektive EKG-getriggerte Technik zur Anwendung. Alle Segmente der Koronararterien, deren Lumendiameter ≥1.5mm betrug, wurden hinsichtlich Stenosen und Bildqualität analysiert und beurteilt. Die retrospektive EKG-getriggerte CTCA wurde bei 39 Patienten und die prospektive EKG-getriggerte CTCA bei 38 Patienten durchgeführt. Die mittlere Herzfrequenz (HF) betrug jeweils 69.5±9.1/min und 62.8±5.9/min. Bei der Detektion von Stenosen ≥50% zeigt die segment-(patienten-) basierte Betrachtung bei der retrospektiven EKG-getriggerten CTCA eine Sensitivität, Spezifität, positiven (PPV) und negativen prädiktiven Wert (NPV) von 97%, 98%, 71%, 100% (91%, 82%, 67%, 96%) und die prospektiv EKG-getriggerte CTCA 94%, 97%, 75%, 99% (93%, 96%, 93%, 96%). In der prospektiv EKG-getriggerten CTCA-Gruppe steigt die Sensitivität und der NPV bei Patienten mit einer HF ≤65/min. Gefäßspezifische Untersuchungen weisen bei der prospektiven Technik eine geringere diagnostische Aussagekraft bezüglich der rechten Koronararterie (RCA) auf, welche jedoch bei einer HF ≤65/min ansteigt. Die Bildqualität unterscheidet sich nicht signifikant in beiden Gruppen. Die Arbeit hat gezeigt, dass die prospektive EKG-getriggerte CTCA in einer heterogenen Patientenpopulation eine hohe diagnostische Genauigkeit und Bildqualität bei HF ≤65/min aufweist. Eine niedrige HF ist für die Beurteilung der RCA von besonderer Bedeutung.

This dissertation presents an investigation of micromachined three-dimensional microelectrode arrays (3-D MEAs) targeted toward in-vitro and in-vivo biomedical applications. Current 3-D MEAs are predominantly silicon-based, fabricated in a planar fashion, and are assembled to achieve a true 3-D form: a technique that cannot be extended to micro-manufacturing. The integrated 3-D MEAs developed in this work are polymer-based and thus offer potential for large-scale, high volume manufacturing. Two different techniques are developed for microfabrication of these MEAs - laser micromachining of a conformally deposited polymer on a non-planar surface to create 3-D molds for metal electrodeposition; and metal transfer micromolding, where functional metal layers are transferred from one polymer to another during the process of micromolding thus eliminating the need for complex and non-repeatable 3-D lithography processes. In-vitro and in-vivo 3-D MEAs are microfabricated using these techniques and are packaged utilizing Printed Circuit Boards (PCB) or other low-cost manufacturing techniques. To demonstrate in-vitro applications, growth of 3-D co-cultures of neurons/astrocytes and tissue-slice electrophysiology with brain tissue of rat pups were implemented. To demonstrate in-vivo application, measurements of nerve conduction were implemented. Microelectrode impedance models, noise models and various process models were evaluated. The results confirmed biocompatibility of the polymers involved, acceptable impedance range and noise of the microelectrodes, and potential to improve upon an archaic clinical diagnostic application utilizing these 3-D MEAs.

The thesis entitled "Pushing the Limits of NMR Sensitivity and Chiral Analysis: Design of New NMR Methods and Bio-molecular Tools" consists of six chapters. The research work reported in this thesis is focused on the development of novel chemical and NMR methodological approaches for enantiomeric analysis and mea- surement of residual dipolar couplings (RDCs), and the development of sensitivity enhanced slice selective NMR experiments for obtaining pure shift 1H spectra and the measurement of scalar couplings. The thesis is divided into two parts. The Part I comprises chapters 2-4, where the enantiomeric analysis is discussed, which includes newly developed chiral reagents, two new weak chiral aligning media and design of novel NMR techniques. Part II comprises chapters 5 and 6, which discusses new sensitivity enhanced slice selective NMR techniques. Chapter 1 gives a general introduction to NMR and the problems investigated in the remaining chapters of the thesis. The chapter starts with a brief discussion on the introduction, advancements and general applications of NMR, discussion is also given on the NMR approaches for enantiomeric analysis both in isotropic and anisotropic phases and the measurement of RDCs, including the benefits and limitations associated with each approach. The chapter sets the tone by discussing limitations of the existed NMR enantiomeric approaches and slice-selective techniques, and builds the bridge for the rest of the chapters by addressing these limitations. The chapter also introduces slice selective experiments, their benefits over other conventional methods and limitations. Additional introductory notes are also given on some related concepts. Part I : NMR Chiral analysis and RDCs measurements Chapter 2 discusses chiral sensing properties of RNA nucleosides and their utility as chiral derivatizing agents for the enantio-discrimination of 1o-amines using one dimensional 1H NMR. A three component protocol has been proposed for the complexation of nucleosides with amines, which is rapid, economical and provides maximum diastereomeric conversion. The chiral differentiating ability of nucleosides are examined for different amines based on the 1H NMR chemical shift differences between the diastereomers (∆δ R, S ). Enantiomeric differentiation has been observed at multiple chemically distinct proton sites. It is observed that adenosine and guanosine exhibit large chiral differentiation (∆δ R, S ) due to the presence of a purine ring. The comparison of the diastereomeric excess (de) measured by NMR with those of the gravimetrically prepared ratios are in excellent agreement with each other confirming the robustness of these RNA nucleosides in discriminating primary amines. Chapter 3 establishes the smooth connectivity with the chapter 2 by discussing the limitations of the enantiomeric discrimination using NMR in isotropic solutions. This chapter discusses two new water compatible aligning media that were developed based on self-assembling strategy of small bio-molecules. The self-assembled folic acid, and the binary mixture of 50-GMP and guanosine are introduced as two novel weak aligning media. The properties of these low ordered media have been systematically studied for their easy preparation, physical parameter dependent tunability of their degree of alignment, mesosphere sustainability over a broad range of temperature and the concentration of the ingredients, and the phase reproducibility. The applications of both these new media are demonstrated for chiral and pro-chiral discrimination and also for the measurement of RDCs. Both these liquid crystalline media could be tuned to very low degree of alignment (order parameter of the order of 10−4), which provides simple first order spectra of molecules aligned in them, the analysis provide order dependent NMR spectral parameters. The 50-GMP:guanosine orienting medium can be prepared in less than 1 hour, and has been demonstrated to be an ideal medium for the determination of RDCs that are used as restraints in the structure calculations of small molecules. Chapter 4 describes 1H NMR spectral complexity in isotropic and anisotropic phases and its consequences on enantiomeric analysis. In circumventing such problems, new NMR techniques have been developed and the spin dynamics involved in the designed sequences are discussed. The newly developed 2D 1H NMR experimental method termed as RES-TOCSY, and its applicability for resolving R and S enantiomeric or diastereomeric peaks of all the coupled proton spins in isotropic phase is discussed. The utility of the developed method is demonstrated in diverse situations, such as, for suppressing impurities peaks, resolving the severely overlapped peaks and unraveling the peaks masked due to severe line broadening when metal complexes are used as chiral auxiliaries. The advantages and limitations of the method over other methods available in the literature are discussed and the significant advantage of the present method is illustrated by spectral comparison with J-resolved experiment. The appli- cation of the method for the accurate measurement of enantiomeric excess has also been demonstrated. The chapter also introduces another NMR experimental technique developed for resolving enantiomeric peaks and complete unraveling of R and S spectra in anisotropic phase. The developed 2D NMR method is cited in the literature as CH-RES-TOCSY. In addition to spectroscopic visualization of R and S spectra, the method also yields C-H RDCs. The applicability of the new experiment has been demonstrated on a chosen example. The wide utility of the method has also been demonstrated for the assignment of symmetric cis- and trans- isomers. Part II : Sensitivity Enhancement of Slice selective NMR Experiments Chapter 5 describes applications of slice selective NMR experiments over conven tional NMR methods and their limitations as far as the sensitivity of signal detection is concerned, especially in low concentrated samples. The chapter introduces the implementation of Acceleration by Sharing Adjacent Polarization (ASAP) technique in slice selective experiments. It is convincingly demonstrated that ASAP helps in reducing inter scan relaxation delay and consequently permits acquisition of more number of scans in a given time, resulting in the gain in signal enhancement by a factor of two. The pulse sequences have been suitably designed for obtaining the pure shift 1H spectra and in G-SERF experiment for the measurement of 1H-1H couplings, both with significantly enhanced signal intensities. Chapter 6 describes new sensitivity enhanced slice selective NMR methods for mea- surement of scalar couplings. A new experiment has been developed which is named as Quick G-SERF (QG-SERF). It is a 1D NMR slice selective method developed based on real time spin manipulation technique. The method gives multiple scalar couplings of a selected spin with simplified multiplets, which is analogous to the 2D G-SERF but with considerable saving in instrument time by 1-2 orders of magnitude. The rapidness of the experiment arises due to reduced dimensionality. The spin dynamics involved in the pulse sequence and its working principle have been described. The application of the method is illustrated for the measurement of 1H-1H couplings. The sequence has been further improved to obtain the heteronuclear couplings between two abundant spins in an orchestrated manner and has been demonstrated for measurement of 1H-19F couplings. This sequence cited as HF-QG-SERF has been implemented on the molecules containing number of chemically non-equivalent fluorine atoms.

The visualization of a 3D image obtained from computerized tomography examinations has shown itself to be an important factor for increasing the quality of medical diagnoses and, consequently, treatment efficacy. There already exist on the market, several visualization softwares, which use different techniques to show the 3D tomography image. However, to show a high quality 3D image, sophisticated devices must be used to obtain slices, close to one another, thus increasing the incidence of X-ray given to the patient. An interpolation slice method which resulted from the TC examination produces good results, and is able to reduce the X-ray incidence upon the patient. This method must reconstruct the curvature from the patient s internal structures without using slices in close proximity. This work proposes a method of 3D image interpolation, composed of a juxtaposition of the slices from CT examination results. The goal of this method is to increase the quality of 3D visualization through the production of sharp and precise structure contours. This thesis proposes the division of the interpolation method into two steps. In the first step, the goal is to obtain an initial representation of the image in 3D, which is composed of real slices as well as virtual slices which are referred to in this work as initial virtual slices. In the second step, the empty spaces of the structure are recovered by the 3D image inpainting process. This work also proposes a method to obtain the initial virtual slice and two different methods for inpainting the 3D image. These inpainting methods are the transversal slice line prolongation method and the transportation and diffusion of information. Both methods use the differential equation theory. The transportation and diffusion of information method shows better results than other methods proposed in this work, besides this, this method presents better results than the linear interpolation and Goshtasby et al. [1] methods also implemented in this work. Visual and numerical comparisons are used to obtain this conclusion. The numerical measures used are statistical correlation, the PSNR and the Hausdorff distance [2]. The transportation and diffusion of information method shows itself able to produce better results than all the other tested methods. Besides this principal contribution, this work also developed a KIT to implement 2D and 3D CT visualize applications. / A visualização de imagens resultantes de exame de tomografia computadorizada (TC) em 3D ´e um fator importante para o aumento da precisão nos diagnósticos médicos e, consequentemente, na eficácia dos tratamentos. Atualmente existem diversos produtos no mercado, que fazem uso de várias técnicas existentes para apresentação de imagens tomográficas em 3D. Contudo, para se obter maior suavidade e precisão nos contornos das estruturas visualizadas em 3D, utiliza-se equipamentos capazes de produzir fatias paralelas do corpo humano muito próximas uma das outras, aumentando a exposição dos pacientes aos raios X. Um método de interpolação de fatias resultantes de exame de TC que forneça bons resultados, pode reduzir a incidência de raios X no paciente, pois esse método pode recuperar a curvatura das estruturas sem a necessidade de uma grande proximidade entre as fatias. Este trabalho propõe um método para a interpolação de imagem em 3D, formada pela justaposição de fatias de resultados de exames de tomografia computadorizada. O objetivo desse método ´e obter contornos suaves e precisos, melhorando os processos de visualização em 3D. Para isso, esta tese propõe a divisão do processo de interpolação em duas etapas. Na primeira etapa obtém-se uma representação inicial da imagem em 3D composta por fatias reais e por fatias denominadas de fatias virtuais iniciais e, na segunda etapa, restaura-se essas estruturas geradas com um processo de retoque de imagem em 3D. Este trabalho propõe também um método para obtenção da fatia virtual inicial e dois métodos diferentes para a realização do passo de retoque da imagem em 3D resultante da justaposição das fatias reais e virtuais iniciais. Esses métodos são o prolongamento de linhas nas fatias transversais e transporte e difusão de informações. Ambos os métodos utilizam a teoria de equações diferenciais. O método de transporte e difusão de informações demonstrou melhores resultados do que outro método proposto neste trabalho, além de obter melhores resultados do que os métodos de interpolação linear e Goshtasby e outros [1] implementados neste trabalho. Comparações visuais e comparações numéricas utilizando a correlação estatística, a PSNR e a distância de Haussdorff [2] foram realizadas para se obter essas conclusões. O método de transporte e difusão de informações é capaz de gerar contornos mais suaves e precisos que esses outros métodos testados. Além dessa contribuição principal, este trabalho também desenvolveu um KIT para a construção de aplicações visualizadoras de tomografias computadorizadas em 2D e em 3D. / Mestre em Ciências

The development of new therapeutical approaches, devised to foster the regeneration of neuronal circuits after injury and/or in neurodegenerative diseases, is of great importance. The impairment of dopaminergic projections is especially severe, because these projections are involved in crucial brain functions such as motor control, reward and cognition. In the work presented here, organotypic brain slice co-cultures of (a) the mesostriatal and (b) the mesocortical dopaminergic projection systems consisting of tissue sections of the ventral tegmental area/substantia nigra (VTA/SN), in combination with the target regions of (a) the striatum (STR) or (b) the prefrontal cortex (PFC), respectively, were used to evaluate different approaches to stimulate neurite outgrowth: (i) inhibition of cAMP/cGMP turnover with 3’,5’ cyclic nucleotide phosphodiesterase inhibitors (PDE-Is), (ii) blockade of calcium currents with nimodipine, and (iii) the co-cultivation with bone marrow-derived mesenchymal stromal/stem cells (BM-MSCs). The neurite growth-promoting properties of the tested substances and cell populations were analyzed by neurite density quantification in the border region between the two brain slices, using biocytin tracing or tyrosine hydroxylase labeling and automated image processing procedures. In addition, toxicological tests and gene expression analyses were conducted. (i) PDE-Is were applied to VTA/SN+STR rat co-cultures. The quantification of neurite density after both biocytin tracing and tyrosine hydroxylase labeling revealed a growth promoting effect of the PDE2A-Is BAY60-7550 and ND7001. The application of the PDE10-I MP-10 did not alter neurite density in comparison to the vehicle control. (ii) The effects of nimodipine were evaluated in VTA/SN+PFC rat co-cultures. A neurite growth-promoting effect of 0.1 µM and 1 µM nimodipine was demonstrated in a projection system of the CNS. In contrast, the application of 10 µM nimodipine did not alter neurite density, compared to the vehicle control, but induced the activation of the apoptosis marker caspase 3. The expression levels of the investigated genes, including Ca2+ binding proteins (Pvalb, S100b), immediate early genes (Arc, Egr1, Egr2, Egr4, Fos and JunB), glial fibrillary acidic protein, and myelin components (Mal, Mog, Plp1) were not significantly changed (with the exception of Egr4) by the treatment with 0.1 µM and 1 µM nimodipine. (iii) Bulk BM-MSCs that were classically isolated by plastic adhesion were compared to the subpopulation Sca-1+Lin-CD45--derived MSCs (SL45-MSCs). The neurite growth-promoting properties of both MSC populations were quantified in VTA/SN+PFC mouse co-cultures. For this purpose, the MSCs were seeded on glass slides that were placed underneath the co-cultures. A significantly enhanced neurite density within the co-cultures was induced by both bulk BM-MSCs and SL45-MSCs. SL45-MSCs increased neurite density to a higher degree. The characterization of both MSC populations revealed that the frequency of fibroblast colony forming units (CFU-f ) is 105-fold higher in SL45-MSCs. SL45-MSCs were morphologically more homogeneous and expressed higher levels of nestin, BDNF and FGF2 compared to bulk BM-MSCs. Thus, this work emphasizes the vast potential for molecular targeting with respect to the development of therapeutic strategies in the enhancement of neurite regrowth.:Table of contents Abbreviations 1 1. Introduction 2 1.1 The dopaminergic system 2 1.2 Neurite regeneration following mechanical lesions of the CNS 7 1.3 Organotypic brain slice co-cultures 8 1.4 Promising substances and cells to enhance neuroregeneration 10 1.5 The aim of the thesis 14 2. The original research articles 16 2.1 Phosphodiesterase 2 inhibitors promote axonal outgrowth in organotypic slice co-cultures 17 2.2 Nimodipine enhances neurite outgrowth in dopaminergic brain slice co-cultures 35 2.3 Mesenchymal stem cells support neuronal fiber growth in an organotypic brain slice co-culture model 50 3. References 66 Appendices 73 Summary 73 Zusammenfassung 78 Curriculum Vitae 84 Track Record 85 Selbständigkeitserklärung 87 Acknowledgments 88

Maschinelle Beatmung kann neben den gewünschten Effekten eine vorbestehende Lungenerkrankung weiter aggravieren und sogar das Lungenparenchym zuvor lungengesunder Patienten schädigen. Mit Hilfe der quantitativen Computertomographie (qCT) können pathologische Belüftungszustände und gegebenenfalls durch maschinelle Beatmung verursachte Schäden analysiert werden. Solche auf der qCT basierende Analysen der Lungenbelüftung werden jedoch potentiell durch CT-Akquisitions- und Bildrekonstruktionsparameter beeinflusst. Um die Ergebnisse vor allem von Analysen des überbelüfteten Lungenvolumens richtig bewerten zu können, müssen solche Einflüsse untersucht werden. Bei 10 Versuchstieren (Schweine) wurden bei einem konstanten Atemwegsdruck von 25 cm H2O zuerst bei gesunder Lunge und dann erneut nach experimenteller Lungenschädigung CT-Bildserien mit zwei unterschiedlichen Strahlendosen angefertigt. Von diesen Rohdaten wurden Bildserien mit unterschiedlichen Rekonstruktionsparametern angefertigt und in jeder dieser Bildserien das überbelüftete Lungenvolumen bestimmt. Sowohl die Schichtdicke, der Filter als auch die Stromstärke hatten einen signifikanten Einfluss auf das eigentlich konstante überbelüftete Lungenvolumen, der jedoch nur teilweise klinisch relevant war. Bei der Interpretation von Messungen des überbelüfteten Lungenvolumens sollten dennoch die Einflüsse der genannten Parameter beachtet und für Vergleichsuntersuchungen gleiche Parametereinstellungen verwendet werden. Eine Dosisreduktion scheint dabei für Messungen des überbelüfteten Lungenvolumens praktikabel.:Inhaltsverzeichnis 0 Abkürzungsverzeichnis 1 1 Einleitung 3 1.1 Ventilator-associated Lung Injury (VALI) 3 1.2 Computertomographie und Diagnostik von Lungenerkrankungen 5 1.3 Spiral-CT 9 1.4 Datenerfassung und Bildrekonstruktion 10 1.5 Grundlagen zur Dichtemessung 12 1.6 Einfluss von Filter und Schichtdicke auf das Bild 13 1.7 Einfluss von Filter und Schichtdicke auf die Analyse der pulmonalen Überbelüftung 15 1.8 Zielstellung 17 2 Materialien und Methodik 18 2.1 Versuchstiere 18 2.2 Überblick über den Versuchsablauf 18 2.2.1 Prämedikation und Narkoseführung 18 2.2.2 Induktion des Lungenschadens 20 2.2.3 CT-Scans und Bildrekonstruktionen 21 2.3 Segmentierung und volumetrische Analyse 22 2.4 Statistische Analyse 24 3 Ergebnisse 26 3.1 Einfluss von Schichtdicke, Filter und Stromstärke auf normale Lungen 26 3.2 Einfluss von Schichtdicke, Filter und Stromstärke auf geschädigte Lungen 34 3.3 Vergleich der automatischen und manuellen Segmentierung 38 4 Diskussion 40 4.1 Einfluss von Schichtdicke und Filter 42 4.2 Einfluss der Stromstärke 49 4.3 Einfluss der experimentell induzierten Lungenschädigung 53 4.4 Vergleich der Segmentierungssoftware 55 4.5 Diskussion der Methodik 55 4.6 Schlussfolgerung 58 5 Zusammenfassung der Arbeit 60 6 Literaturverzeichnis 63 7 Danksagung 77 8 Erklärung über die eigenständige Abfassung der Arbeit 78

Die koronare Herzkrankheit (KHK) gehört zu den häufigsten Todesursachen in den westlichen Industrienationen. Die Diagnostik der Erkrankung hat somit großen Stellenwert in der Medizin. Akzeptierter Goldstandard zur Diagnostik einer KHK ist die Herzkatheteruntersuchung (HKU). Als nicht-invasive Alternative zur HKU hat sich in den letzten Jahren die Mehrzeilen-Computertomographie als zuverlässiges Verfahren für den KHK-Ausschluss bei mittlerer Vortestwahrscheinlichkeit etabliert. Ziel dieser Arbeit ist es, die diagnostischen Eigenschaften der prospektiv getriggerten mit der retrospektiv getriggerten CT-Koronarangiographie (CTCA) an einem 64-Zeilen Gerät in einem heterogenen Patientenkollektiv mit unterschiedlichen kardiovaskulären Erkrankungen (Verdacht auf Koronare Herzkrankheit, Aortenaneurysma, präoperativ zum Aortenklappenersatz oder zur Pulmonalvenenablation, zum Ausschluss eines Tumors oder Perikarditiden) in Genauigkeit, Bildqualität und ihrer Anwendbarkeit gegenüberzustellen und sie mit dem Referenzstandard, der HKU, zu vergleichen. In diese Studie wurden retrospektiv 77 Patienten eingeschlossen, die ein EKG-getriggertes kardiales CT erhielten. Wenn es möglich war, d.h. die Herzfrequenz <75/min, BMI <35 und ein Sinusrhythmus vorlag, wurde die prospektive EKG-getriggerte CTCA durchgeführt, alternativ kam die retrospektive EKG-getriggerte Technik zur Anwendung. Alle Segmente der Koronararterien, deren Lumendiameter ≥1.5mm betrug, wurden hinsichtlich Stenosen und Bildqualität analysiert und beurteilt. Die retrospektive EKG-getriggerte CTCA wurde bei 39 Patienten und die prospektive EKG-getriggerte CTCA bei 38 Patienten durchgeführt. Die mittlere Herzfrequenz (HF) betrug jeweils 69.5±9.1/min und 62.8±5.9/min. Bei der Detektion von Stenosen ≥50% zeigt die segment-(patienten-) basierte Betrachtung bei der retrospektiven EKG-getriggerten CTCA eine Sensitivität, Spezifität, positiven (PPV) und negativen prädiktiven Wert (NPV) von 97%, 98%, 71%, 100% (91%, 82%, 67%, 96%) und die prospektiv EKG-getriggerte CTCA 94%, 97%, 75%, 99% (93%, 96%, 93%, 96%). In der prospektiv EKG-getriggerten CTCA-Gruppe steigt die Sensitivität und der NPV bei Patienten mit einer HF ≤65/min. Gefäßspezifische Untersuchungen weisen bei der prospektiven Technik eine geringere diagnostische Aussagekraft bezüglich der rechten Koronararterie (RCA) auf, welche jedoch bei einer HF ≤65/min ansteigt. Die Bildqualität unterscheidet sich nicht signifikant in beiden Gruppen. Die Arbeit hat gezeigt, dass die prospektive EKG-getriggerte CTCA in einer heterogenen Patientenpopulation eine hohe diagnostische Genauigkeit und Bildqualität bei HF ≤65/min aufweist. Eine niedrige HF ist für die Beurteilung der RCA von besonderer Bedeutung.:1 Bibliographische Beschreibung 2 Einleitung 2.1 Die koronare Herzerkrankung (KHK) 2.1.1 Definition und Epidemiologie 2.1.2 Ätiologie 2.1.3 Anatomie und Pathophysiologie 2.1.4 Symptomatik 2.2 Diagnostik der KHK 2.2.1 Basisdiagnostik 2.2.2 Bildgebende Diagnostik zur direkten Beurteilung der Koronargefäße 2.3 CT-Verfahren 2.3.1 Retrospektives EKG-Gating 2.3.2 Prospektives EKG-Gating 2.3.3 Diagnostische Genauigkeit der CT-Koronarangiographie (CTCA) 2.4 Aspekte zur Strahlendosis 2.5 Indikationen zur HKU und kardialen CT 2.6 Zielsetzung der Studie 3 Publikation 4 Zusammenfassung 5 Literaturverzeichnis 6 Anlagen 6.1 Selbständigkeitserklärung 6.2 Lebenslauf 6.2.1 Persönliche Daten 6.2.2 Beruflicher Werdegang 6.3 Danksagung

PlanetLab as a distributed systems testbed offers a unique opportunity for developing and testing new applications useful for future Internet. This work brings up a scheme and a solution of the problem with accessing PlanetLab by a larger group of students e.g. for the purpose of solving their courseworks. A designed system empowers its administrator to create and control virtual user accounts which provide possibility for all its users to connect to selected nodes in the PlanetLab.