O objetivo principal deste trabalho foi analisar o controverso conceito de informação em estatística. Para tal, primeiramente foi estudado o conceito de informação dado por Basu. A seguir, a análise foi dividida em três partes: informação nos dados, informação no experimento e diagramas de influência. Nas duas primeiras etapas, sempre se tentou definir propriedades que uma função de informação deveria satisfazer para se enquadrar ao conceito. Na primeira etapa, foi estudado como o princípio da verossimilhança é uma classe de equivalência decorrente de acreditar que experimentos triviais não trazem informação. Também foram apresentadas métricas que satisfazem o princípio da verossimilhança e estas foram usadas para avaliar um exemplo intuitivo. Na segunda etapa, passamos para o problema da informação de um experimento. Foi apresentada a relação da suficiência de Blackwell com experimentos triviais e o conceito usual de suficiência. Também foi analisada a equivalência de Blackwell e a sua relação com o Princípio da Verossimilhança anteriormente estudado. Além disso, as métricas apresentadas para medir a informação de conjuntos de dados foram adaptadas para também medir a informação de um experimento. Finalmente, observou-se que nas etapas anteriores uma série de simetrias mostraram-se como elementos essenciais do conceito de informação. Para ganhar intuição sobre elas, estas foram reescritas através da ferramenta gráfica dos diagramas de influência. Assim, definições como suficiência, suficiência de Blackwell, suficiência mínima e completude foram reapresentadas apenas usando essa ferramenta. / The main objective of this work is to analyze the controversial concept of information in Statistics. To do so, firstly the concept of information according to Basu is presented. Next, the analysis is divided in three parts: information in a data set, information in an experiment and influence diagrams. In the first two parts, we always tried to define properties an information function should satisfy in order to be in accordance to the concept of Basu. In the first part, it was studied how the likelihood principle is an equivalence class which follows from believing that trivial experiments do not bring information. Metrics which satisfy the likelihood principle were also presented and used to analyze an intuitive example. In the second part, the problem became that of determining information of a particular experiment. The relation between Blackwell\'s suciency, trivial experiments and classical suciency was presented. Blackwell\'s equivalence was also analyzed and its relationship with the Likelihood Principle was exposed. The metrics presented to evaluate the information in a data set were also adapted to do so with experiments. Finally, in the first parts a number of symmetries were shown as essencial elements of the concept of information. To gain more intuition about these elements, we tried to rewrite them using the graphic tool of influence diagrams. Therefore, definitions as sufficiency, Blackwell\'s sufficiency, minimal sufficiency and completeness were shown again, only using influence diagrams.

<div>The thesis includes two application studies of computational fluid dynamics. The first is new and efficient drug delivery to the posterior part of the eye, a growing health necessity worldwide. Current treatment of eye diseases, such as age related macular degeneration (AMD), relies on repeated intravitreal injections of drug-containing solutions. Such a drug delivery has significant drawbacks, including short drug life, vital medical service, and high medical costs. In this study, we explore a new approach of controlled drug delivery by introducing unique porous implants. Computational</div><div>modeling contains physiological and anatomical traits. We simulate the IgG1 Fab drug delivery to the posterior eye to evaluate the effectiveness of the porous implants to control the drug delivery. The computational model was validated by established computation results from independent studies and experimental data. Overall, the results indicate that therapeutic drug levels in the posterior eye are sustained for</div><div>eight weeks, similar to those performed with intravitreal injection of the same drug. We evaluate the effects of the porous implant on the time evaluation of the drug concentrations in the sclera, choroid, and retina layers of the eye. Subsequent simulations were carried out with varying porosity values of a porous episcleral implant.</div><div>Our computational results reveal that the time evolution of drug concentration is distinctively correlated to drug source location and pore size. The response of this porous implant for controlled drug delivery applications was examined. A correlation between porosity and fluid properties for the porous implants was revealed in this study. The second application lays in the computational modeling of the oscillating flow in rectangular ducts. This computational study has further applications in investigating the fluid flow motion in bodily organs. It can be useful in studying the</div><div>response of bone cells to the wall shear stress in the human body. </div>

Purpose: The aim of this work was to characterize the healthy human cornea and the cornea of patients suffering from posterior polymorphous corneal dystrophy (PPCD) using different antibodies. Despite the fact that PPCD is a very rare disorder, one of the largest groups of PPCD patients in the world comes from the Czech Republic. This offers us the opportunity to investigate the changes on the clinical, cellular and molecular levels. Material and Methods: A collection of 25 control corneas as well as 16 pathological corneas from PPCD patients were used. Epithelial (cytokeratins) and mesothelial markers (mesothelin, calbindin 2, HBME-1 protein) were detected in all layers of the healthy corneas using immunocyto- and immunohistochemistry. The expression of all markers was confirmed using molecular methods as well (RT-PCR and Western blot). Changes in the expression of cytokeratins and changes in the extracellular matrix structure (collagen IV and VIII) were studied in the PPCD corneas. Combined fluorescent immunohistochemistry with fluorescence in situ hybridization were used in order to characterize the origin of abnormal cells on the posterior graft surface, which cause the recurrence of the PPCD after penetrating keratoplasty surgery. Results: Changes in the cytokeratin expression (strong...

BACKGROUND: Posterior inferior cerebellar artery (PICA) aneurysms are an uncommon, heterogeneous group of aneurysms with poorer neurological outcomes compared to other intracranial aneurysms. At first, as part A, we conducted a systematic review of the literature to evaluate the safety and efficacy of treatment strategies for PICA-aneurysms. Subsequently, as part B, we performed a multicenter retrospective study to analyze the outcome in a large series of patients treated with contemporary microsurgical and endovascular techniques. METHODS: For the meta-analysis, a systematic search of Medline, EMBASE, Scopus and Web of Science was done for studies published through November 2015. We included studies that described treatment of PICA-aneurysms with ≥10 patients. Random-effects meta-analysis was used to pool the following outcomes: complete occlusion, technical success, periprocedural morbidity/mortality, stroke rates, aneurysm recurrence/rebleed, CN-palsies rates, and long-term neurological morbidity/mortality. As the second part, aiming to report the current trends and results in treatment strategies for PICA-aneurysms, records of 94 patients treated for PICA-aneurysms between 2000 and 2015 at 3 large referral neurovascular centers were retrospectively reviewed. RESULTS: In the meta-analysis, we...

Joint models for the association of a longitudinal binary and a longitudinal continuous process are proposed for situations in which their association is of direct interest. The models are parameterized such that the dependence between the two processes is characterized by unconstrained regression coefficients. Bayesian variable selection techniques are used to parsimoniously model these coefficients. A Markov chain Monte Carlo (MCMC) sampling algorithm is developed for sampling from the posterior distribution, using data augmentation steps to handle missing data. Several technical issues are addressed to implement the MCMC algorithm efficiently. The models are motivated by, and are used for, the analysis of a smoking cessation clinical trial in which an important question of interest was the effect of the (exercise) treatment on the relationship between smoking cessation and weight gain.

The common practice for testing measurement invariance is to constrain parameters to be equal over groups, and then evaluate the model-data fit to reject or fail to reject the restrictive model. Posterior predictive checking (PPC) provides an alternative approach to evaluating model-data discrepancy. This paper explores the utility of PPC in estimating measurement invariance. The simulation results show that the posterior predictive p (PP p) values of item parameter estimates respond to various invariance violations, whereas the PP p values of item-fit index may fail to detect such violations. The current paper suggests comparing group estimates and restrictive model estimates with posterior predictive distributions in order to demonstrate the pattern of misfit graphically.

No description available.

The vertebrate tail is unique for each species and fulfils a broad spectrum of functions. In the axolotl (Ambystoma mexicanum), a tailed amphibian, the tail constitutes one-third of the full body length and is necessary for swimming. Despite its size, most of the tail's tissues are derived from the posterior neural plate of the neurula. Although giving rise to neuronal structures of the central nervous system along most of its length, the most posterior part of the neural plate develops preponderantly into presomitic mesoderm (PSM) which forms muscle, bone and cartilage of the tail and posterior trunk. During development, the posterior neural plate reverses its orientation during an anterior turn movement (Taniguchi et al., 2017). Cells of the most posterior plate region become now localised in an anterior position while previously more anterior neural plate cells land at a more posterior site. Simultaneously, the axial neural tube and notochord extend themselves posteriorly. The PSM, developing bilaterally to the central axis, is integrated into posterior tail expansion while forming new somites at its anterior end. It is still elusive which morphological changes the PSM undergoes to facilitate tail formation and posterior elongation of the embryo. Furthermore, it remains enigmatic in what way PSM cells change their shape, orientation, migration behaviour and distribution to meet the requirements needed for adjusting PSM and somite morphology. With homotopic tissue transplantations of posterior neural plate cells from a gfp-expressing donor to a white (d/d) recipient, enabled specific labelling of all mesodermal cells of the tail. Otherwise, mesodermal cells of the trunk and tail can not be distinguished, neither genetically nor morphologically. With this cell labelling approach, the entire tail mesoderm could be imaged in toto. Thus, measurements of the morphological changes of the PSM and cell tracking in 3D was possible during development. With this technique, posterior neural plate cells could be shown to form parts of the posterior neural tube, the entire posterior PSM and the somites of the tail. During this course of development, the PSM becomes longer but does not increase its volume. Only when forming the somites, an increase in volume could be measured in the mesoderm. Single-cell labelling showed an anterior shift of cell movement led by medial PSM cells and followed by more laterally located cells. The anterior displacement happens simultaneously to the posterior elongation of the embryo. A hypothetical push by newly generated cells at the tail tip could be ruled out. Mitotic cells were evenly distributed in all tissues of the tail with a low proliferation rate. The morphological changes and anterior relocations of the tail mesoderm could, therefore, mainly be explained by cell migration. Therefore, further analyses focussed on cell migration, particularly on cellular characteristics displayed during migration such as shape, orientation, volume, distribution and filopodia organisation to obtain more profound information about how PSM cells migrate and contribute to somite formation. The net movement of tail elongation is directed posteriorly regardless of anteriorly relocating PSM cells. That is only feasible if a lateral expansion of the PSM by laterally migrating PSM cells is counteracted. There have been no studies on the lateral boundary so far. In the axolotl, the PSM is covered laterally by a two-layered epidermis and a fibronectin-rich extracellular matrix. After removing the tail epidermis, operated embryos showed missing or malformed tails, especially with lateral and dorsal curvatures and shortenings. Tail mesoderm examined in these cases showed an increased PSM volume and a lateral expansion of the tissue. A nearly normal tail developed when, after removing the epidermis, the embryos developed in 1% agarose supplemented with fibronectin. In contrast, a simple covering of the PSM with a nitrocellulose membrane, incubation in the softer methylcellulose or in agarose without fibronectin did not rescue tail formation. The lateral pressure on the PSM and a fibronectin-rich extracellular matrix seem necessary to preserve the tissue architecture of the PSM during tail formation. This study unravels the behaviour of individual PSM cells during their morphogenesis from single cells in the posterior plate of the neurula until somite formation in the tail bud. Overall, with specific labelling of tail mesodermal cells, their contribution to PSM morphology could be elucidated, and a more detailed model of tail elongation could be proposed: The posterior expansion of the neural tube and notochord pushes the posterior neural plate tissue posteriorly and squeezes the cells into an elongated mediolaterally oriented form. Labelling experiments of small individual cell groups showed that the ventral posteriormost cells are the first to escape this pressure by relocating anteriorly. Then, more anteriorly located cells follow, as well as dorsally located cells. These movements explain the anterior turn. Thereby, mesodermal cells start to migrate randomly, become elongated and change their orientation from mediolateral to anterior-posterior. Random cell migration leads to homogeneous cell mixing, which results in an aligned uniform tissue of trunk and tail PSM. The lateral constriction by the epidermis channels the undirected migration movements in an anterior direction. In this way, cells are directed towards the site of somite formation, the PSM narrows, and the embryo elongates posteriorly. This extension model includes the individual cell behaviour, which on the whole shapes PSM morphology. The analysed dynamic morphological changes of the PSM can be linked to the developmental processes of the tail and the posterior elongation of the axis.:1 Introduction 1.1 Embryonic tail formation . . . . . . . . . . . . . . . . . . . . . . . . . . 1 1.1.1 Mechanism of tail formation . . . . . . . . . . . . . . . . . . . . . 1 1.1.2 Molecular determination of cell populations in the tail bud . . . . . 5 1.2 Axial elongation of the vertebrate body plan . . . . . . . . . . . . . . . . . 8 1.2.1 Anterior body elongation (elongation of the trunk) . . . . . . . . . 8 1.2.2 Posterior body elongation (tail elongation) . . . . . . . . . . . . . . 9 1.3 Studying tissue morphology during development . . . . . . . . . . . . 11 1.4 Aim of the project . . . . . . . . . . . . . . . . . . . . . . .. . . . . . 12 2 Materials 2.1 Chemicals and solutions . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14 2.2 Antibodies and dyes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16 2.3 Techniqual equipment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16 2.4 Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18 3 Methods 3.1 Animals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19 3.1.1 Breeding of axolotls and embryo collection . . . . . . . . . . . . 19 3.1.2 Injections with the vital dye DiI . . . . . . . . . . . . . . . . . . . 19 3.1.3 Tissue transplantation techniques . . . . . . . . . . . . . . . . . . . 19 3.2 Immunohistochemical staining . . . . . . . . . . . . . . . . . . . . . . . . 20 3.2.1 Vibratome sections . . . . . . . . . . . . . . . . . . . . . . . . . . . 20 3.2.2 Whole-mount staining . . . . . . . . . . . . . . . . . . . . . . . . . 21 3.3 Optical tissue clearing protocols . . . . . . . . . . . . . . . . . . 21 3.3.1 Ethyl cinnamate based optical tissue clearing protocol . . . . . . . 21 3.3.2 SeeDB optical clearing protocol . . . . . . . . . . . . . . . . . . . . 22 3.4 Image analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22 3.4.1 3D image generation and processing . . . . . . . . . . .. . . . . . 22 3.4.2 Length measurements . . . . . . . . . . . . . . . . . . . . . . . . . . 22 3.4.3 Manual segmentation . . . . . . . . . . . . . . . . . . . . . . . . . 24 3.4.4 Automatic segmentation . . . . . . . . . . . . . . . . . . . . . . . . 25 3.5 Determination of cellular parameters . . . . . . . . . . . . . . .. . . . . 25 3.5.1 Cell shape . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25 3.5.2 Cell and tissue volume . . . . . . . . . . . . . . . . . . . . . . . . . 25 3.5.3 Cellular distribution . . . . . . . . . . . . . . . . . . . . . . . . . . 25 3.5.4 Closest neighbour analysis . . . . . . . . . . . . . . . . . . . . . . . 26 3.5.5 Cell orientation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27 3.5.6 Length and orientation of filopodia . . . . . . . . . . . . . . . . . . 31 3.5.7 Distance of cells to a plane . . . . . . . . . . . . . . . . . . . . . . . 31 3.5.8 Mitotic rate and spindle orientation . . . . . . . . . . . . . . . . . 32 4 Results 4.1 The presomitic mesoderm is associated with axial elongation. . . . . . 33 4.1.1 Elongation of the body axis . . . . . . . . . . . . . . . . . . . . . . 33 4.1.2 Contribution of different tissues . . . . . . . . . . . . . . . . . . . . 34 4.1.3 Differential contribution of mesoderm and epidermis . . . . . . . . . 40 4.1.4 Dual potential of mesodermal progenitors . . . . . . . . . . . . . . . 42 4.1.5 Mesodermal tissue expansion . . . . . . . . . . . . . . . . . . . . . 46 4.2 Cellular behaviour influences mesodermal morphology . . . . . . . . . 50 4.2.1 Cell division . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50 4.2.2 Positional changes . . . . . . . . . . . . . . . . . . . . . . . . . . . 53 4.2.3 Cellular characteristics . . . . . . . . . . . . . . . . . . . . . . . . . 59 Cell shape changes . . . . . . . . . . . . . . . . . . . . . . . . . . . 59 Change of cell orientation . . . . . . . . . . . . . . . . . . . . . . . 61 Orientation of filopodia . . . . . . . . . . . . . . . . . . . . . . . . . 63 Cell distribution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66 4.3 The epidermis fascilitates mesodermal tissue integrity . . . . . . .. . . . 67 4.3.1 Mesodermal tissue integrity . . . . . . . . . . . . . . . . . . . . . . 68 4.3.2 Malformed tails after epidermis removal . . . . . . . . . . . . . . . 70 4.3.3 Alteration in mesodermal tissue dimensions . . . . . . . . . . . . . 73 4.3.4 Alteration of cell density after epidermis removal . . . . . . . . . . 77 4.3.5 Rescue of tail formation . . . . . . . . . . . . . . . . . . . . . . . . 80 5 Discussion 5.1 Cell migration of the presomitic mesodermal cells . . . . . . . . .. . . . 85 5.1.1 Continuity of gastrulation movements . . . . . . . . . . . . . . . . . 85 5.1.2 Directed migration . . . . . . . . . . . . . . . . . . . . . . . . . . . 86 5.1.3 Random cell migration . . . . . . . . . . . . . . . . . . . . . . . . . 88 5.1.4 Lateral mechanical constriction . . . . . . . . . . . . . . . . . . . . 90 5.2 Non-volumetric growth of the presomitic mesoderm . . . . . . . . . . . . . 91 5.3 Models of tail presomitic mesoderm formation . . . . . . . . . . . . . . . . 93 / Der Schwanz der Wirbeltiere ist bei jeder Art einzigartig und erfüllt ein breites Spektrum an Funktionen. Beim Salamander Axolotl (Ambystoma mexicanum), macht der Schwanz ein Drittel der gesamten Körperlänge aus und ist zum Schwimmen notwendig. Trotz seiner Größe stammen die meisten Gewebe des Schwanzes von der posterioren Neuralplatte der Neurula ab. Obwohl der größte Teil der Neuralplatte neuronale Strukturen des Zentralnervensystems hervorbringt, entwickelt sich der posteriore Teil der Neuralplatte überwiegend zu präsomitischem Mesoderm (PSM), das Muskeln, Knochen und Knorpel des Schwanzes und des hinteren Rumpfes bildet. Während der Entwicklung kehrt die posteriore Neuralplatte ihre Orientierung in einer anterioren Drehbewegung um (Taniguchi et al., 2017). Zellen der hintersten Plattenregion werden in eine anteriore Position verschoben, während zuvor anteriorere Neuralplattenzellen an einer posterioren Stelle landen. Gleichzeitig verlängert sich das axiale Neuralrohr und das Notochord nach posterior. Das PSM, das sich bilateral zur Zentralachse entwickelt, ist im Prozess der Schwanzverlängerung involviert, während es gleichzeitig an seinem vorderen Ende neue Somiten bildet. Es ist immer noch unklar, welche morphologischen Veränderungen das PSM durchläuft, um die Schwanzbildung und die posteriore Ausdehnung des Embryos zu ermöglichen. Darüber hinaus ist unbekannt, auf welche Weise PSM-Zellen ihre Form, Orientierung, ihr Migrationsverhalten und ihre Verteilung ändern, die für eine Veränderung der PSM- und Somitenmorphologie erforderlich sind. Mit homotopen Gewebetransplantationen von posterioren Neuralplattenzellen von einem gfp-exprimierenden Spender auf einen weißen (d/d) Empfänger, konnte eine spezifische Markierung aller mesodermalen Zellen des Schwanzes erreicht werden. Andernfalls können mesodermale Zellen des Rumpfes und des Schwanzes weder genetisch noch morphologisch unterschieden werden. Mit diesem Zellmarkierungsansatz konnte das gesamte Schwanzmesoderm in toto abgebildet werden. So waren Messungen der morphologischen Veränderungen des PSM und Zellverfolgung in 3D während der Entwicklung möglich. Mit dieser Technik konnte gezeigt werden, dass die Zellen der posterioren Neuralplatte Teile des posterioren Neuralrohrs, das gesamte posteriore PSM und die Somiten des Schwanzes bilden. Dabei wird das PSM länger, ohne sein Volumen zu vergrößern. Erst während der Bildung von Somiten wurde eine Volumenzunahme gemessen Einzelzellmarkierungen zeigten eine anteriore Verschiebung der Zellen, angeführt von medialen PSM-Zellen und gefolgt von lateral gelegenen Zellen. Diese anteriore Verschiebung geschieht gleichzeitig mit der posterioren Streckung des Embryos. Ein hypothetischer Schub durch neugebildete Zellen an der Schwanzspitze konnte ausgeschlossen werden. Mitotischen Zellen waren gleichmäßig in allen Geweben des Schwanzes verteilt und wiesen eine geringe Proliferationsrate auf. Die morphologischen Veränderungen und anterioren Verlagerungen des Schwanzmesoderms können daher hauptsächlich durch Zellmigration erklärt werden. Die Analysen konzentrierten sich daher auf die Zellmigration, insbesondere auf die zellulären Charakteristika, die sich während der Migration zeigen, wie z.B. Form, Orientierung, Volumen, Verteilung und Filopodienorganisation. So konnten neue Informationen darüber gewonnen werden, wie PSM-Zellen wandern und zur Somitenbildung beitragen. Die Nettobewegung der Schwanzverlängerung ist, unabhängig von nach anterior wandernden PSM-Zellen, nach posterior gerichtet. Das ist nur möglich, wenn einer lateralen Ausdehnung des PSM durch ungerichtet migrierenden Zellen entgegengewirkt wird. Über die Rolle einer laterale Begrenzung bei diesem Prozess gibt es bisher keine Untersuchungen. Beim Axolotl ist das PSM seitlich von einer zweischichtigen Epidermis und einer Fibronektin-reichen extrazellulären Matrix bedeckt. Nach Entfernung der Schwanzepidermis zeigten operierte Embryonen fehlende oder missgebildete Schwänze, insbesondere mit einer lateralen und dorsalen Krümmung und einer Verkürzung. Untersuchungen des Schwanzmesoderms zeigten ein erhöhtes PSM-Volumen und eine laterale Ausdehnung des Gewebes. Ein nahezu normaler Schwanz entwickelte sich, wenn die Embryonen nach Entfernung der Epidermis mit 1% Agarose, ergänzt mit Fibronektin, bedeckt wurden. Im Gegensatz dazu konnte eine einfache Abdeckung des PSM mit einer Nitrozellulosemembran, die Inkubation in der weicheren Methylzellulose oder in Agarose ohne Fibronektin die Schwanzbildung nicht normalisieren. Der seitliche Druck auf das PSM und eine Fibronektin-reiche extrazelluläre Matrix scheinen notwendig zu sein, um die Gewebearchitektur des PSM während der Schwanzbildung zu erhalten. Diese Studie zeigt das Verhalten einzelner PSM-Zellen während der Morphogenese der hinteren Neuralplatte bis zur Somitenbildung. Insgesamt konnte durch die spezifische Markierung von mesodermalen Zellen des Schwanzes deren Beitrag zur PSM-Morphologie aufgeklärt und ein detaillierteres Modell der Schwanzverlängerung vorgeschlagen werden: Die posteriore Ausdehnung des Neuralrohrs und des Notochords schiebt das posteriore Neuralplattengewebe nach hinten und quetscht die Zellen in eine verlängerte, mediolateral orientierte Form. Markierungsexperimente einzelner Zellgruppen zeigten, dass die ventralen, posterior gelegenen Zellen diesem Druck als erste entkommen, indem sie sich nach anterior verschieben. Ihnen folgen weiter anterior gelegene Zellen sowie dorsal gelegene Zellen. Diese Bewegungen erklären die anteriore Drehung. Dabei beginnen mesodermale Zellen ungerichtet zu wandern, verlängern sich und ändern ihre Orientierung von mediolateral nach anterior-posterior. Die ungerichtete Zellwanderung führt zu einer homogenen Zelldurchmischung, so dass zusammen mit dem PSM des Rumpfes ein einheitliches Gewebe gebildet wird. Die laterale Begrenzung durch die Epidermis kanalisiert die ungerichteten Migrationsbewegungen in anteriore Richtung. Auf diese Weise werden die Zellen in Richtung der Somitenbildungsstelle gelenkt, das PSM verengt sich, und der Embryo streckt sich nach hinten. Dieses Ausdehnungsmodell beinhaltet das individuelle Zellverhalten, das insgesamt die Morphologie des PSM prägt. Die analysierten dynamischen morphologischen Veränderungen des PSM können mit Schwanzentwicklungsprozessen und der posterioren Elongation der Achse in Verbindung gebracht werden.:1 Introduction 1.1 Embryonic tail formation . . . . . . . . . . . . . . . . . . . . . . . . . . 1 1.1.1 Mechanism of tail formation . . . . . . . . . . . . . . . . . . . . . 1 1.1.2 Molecular determination of cell populations in the tail bud . . . . . 5 1.2 Axial elongation of the vertebrate body plan . . . . . . . . . . . . . . . . . 8 1.2.1 Anterior body elongation (elongation of the trunk) . . . . . . . . . 8 1.2.2 Posterior body elongation (tail elongation) . . . . . . . . . . . . . . 9 1.3 Studying tissue morphology during development . . . . . . . . . . . . 11 1.4 Aim of the project . . . . . . . . . . . . . . . . . . . . . . .. . . . . . 12 2 Materials 2.1 Chemicals and solutions . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14 2.2 Antibodies and dyes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16 2.3 Techniqual equipment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16 2.4 Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18 3 Methods 3.1 Animals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19 3.1.1 Breeding of axolotls and embryo collection . . . . . . . . . . . . 19 3.1.2 Injections with the vital dye DiI . . . . . . . . . . . . . . . . . . . 19 3.1.3 Tissue transplantation techniques . . . . . . . . . . . . . . . . . . . 19 3.2 Immunohistochemical staining . . . . . . . . . . . . . . . . . . . . . . . . 20 3.2.1 Vibratome sections . . . . . . . . . . . . . . . . . . . . . . . . . . . 20 3.2.2 Whole-mount staining . . . . . . . . . . . . . . . . . . . . . . . . . 21 3.3 Optical tissue clearing protocols . . . . . . . . . . . . . . . . . . 21 3.3.1 Ethyl cinnamate based optical tissue clearing protocol . . . . . . . 21 3.3.2 SeeDB optical clearing protocol . . . . . . . . . . . . . . . . . . . . 22 3.4 Image analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22 3.4.1 3D image generation and processing . . . . . . . . . . .. . . . . . 22 3.4.2 Length measurements . . . . . . . . . . . . . . . . . . . . . . . . . . 22 3.4.3 Manual segmentation . . . . . . . . . . . . . . . . . . . . . . . . . 24 3.4.4 Automatic segmentation . . . . . . . . . . . . . . . . . . . . . . . . 25 3.5 Determination of cellular parameters . . . . . . . . . . . . . . .. . . . . 25 3.5.1 Cell shape . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25 3.5.2 Cell and tissue volume . . . . . . . . . . . . . . . . . . . . . . . . . 25 3.5.3 Cellular distribution . . . . . . . . . . . . . . . . . . . . . . . . . . 25 3.5.4 Closest neighbour analysis . . . . . . . . . . . . . . . . . . . . . . . 26 3.5.5 Cell orientation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27 3.5.6 Length and orientation of filopodia . . . . . . . . . . . . . . . . . . 31 3.5.7 Distance of cells to a plane . . . . . . . . . . . . . . . . . . . . . . . 31 3.5.8 Mitotic rate and spindle orientation . . . . . . . . . . . . . . . . . 32 4 Results 4.1 The presomitic mesoderm is associated with axial elongation. . . . . . 33 4.1.1 Elongation of the body axis . . . . . . . . . . . . . . . . . . . . . . 33 4.1.2 Contribution of different tissues . . . . . . . . . . . . . . . . . . . . 34 4.1.3 Differential contribution of mesoderm and epidermis . . . . . . . . . 40 4.1.4 Dual potential of mesodermal progenitors . . . . . . . . . . . . . . . 42 4.1.5 Mesodermal tissue expansion . . . . . . . . . . . . . . . . . . . . . 46 4.2 Cellular behaviour influences mesodermal morphology . . . . . . . . . 50 4.2.1 Cell division . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50 4.2.2 Positional changes . . . . . . . . . . . . . . . . . . . . . . . . . . . 53 4.2.3 Cellular characteristics . . . . . . . . . . . . . . . . . . . . . . . . . 59 Cell shape changes . . . . . . . . . . . . . . . . . . . . . . . . . . . 59 Change of cell orientation . . . . . . . . . . . . . . . . . . . . . . . 61 Orientation of filopodia . . . . . . . . . . . . . . . . . . . . . . . . . 63 Cell distribution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66 4.3 The epidermis fascilitates mesodermal tissue integrity . . . . . . .. . . . 67 4.3.1 Mesodermal tissue integrity . . . . . . . . . . . . . . . . . . . . . . 68 4.3.2 Malformed tails after epidermis removal . . . . . . . . . . . . . . . 70 4.3.3 Alteration in mesodermal tissue dimensions . . . . . . . . . . . . . 73 4.3.4 Alteration of cell density after epidermis removal . . . . . . . . . . 77 4.3.5 Rescue of tail formation . . . . . . . . . . . . . . . . . . . . . . . . 80 5 Discussion 5.1 Cell migration of the presomitic mesodermal cells . . . . . . . . .. . . . 85 5.1.1 Continuity of gastrulation movements . . . . . . . . . . . . . . . . . 85 5.1.2 Directed migration . . . . . . . . . . . . . . . . . . . . . . . . . . . 86 5.1.3 Random cell migration . . . . . . . . . . . . . . . . . . . . . . . . . 88 5.1.4 Lateral mechanical constriction . . . . . . . . . . . . . . . . . . . . 90 5.2 Non-volumetric growth of the presomitic mesoderm . . . . . . . . . . . . . 91 5.3 Models of tail presomitic mesoderm formation . . . . . . . . . . . . . . . . 93

Bayesian reference analysis is a method of determining the prior under the Bayesian paradigm. It incorporates as little information as possible from the experiment. Estimation of the ratio of two independent Poisson rates is a common practical problem. In this thesis, the method of reference analysis is applied to derive the posterior distribution of the ratio of two independent Poisson rates, and then to construct point and interval estimates based on the reference posterior. In addition, the Frequentist coverage property of HPD intervals is verified through simulation.

Drug development is time consuming, expensive with high failure rates. It takes 10-15 years for a drug to go from discovery to approval, while the mean cost of developing a drug is $1.5 billions dollars. Pharmacometric models (PM) play a pivotal role in knowledge driven drug development and these models require validation prior to application. The purpose of the current study was to evaluate the posterior predictive check (PPC) and the bootstrap as population model validation tools. PPC was evaluated to determine, if it was able to distinguish between population pharmacokinetic (PPK) models that were developed/estimated from influence data versus models that were not derived/estimated from influence data. Bootstrap was examined to see if there was a correspondence between the root mean squared prediction errors (RMSPE) for serum concentrations when estimated by external prediction methods versus when estimated by the standard bootstrap. In the case of PPC, C last , C first -C last and C mid values from initial data sets were compared to corresponding posterior distributions. In the case of no influence data for C last , C first -C last and C mid on average 76%, 30% and 52% of the values from the posterior distributions were below the initial C last , C first -C last and C mid on average 93%, 13% and 67% of the values from the posterior distributions were below the initial C last , C first -C last and C mid respectively. PPC was able to classify models from influence versus no influence data. In the case of bootstrap when the original model was used to predict into the external data the WRMSPE for drug 1, drug 2, drug 3, drug 4 and simulated data set was 10.40 mg/L, 20.36 mg/L, 0.72 mg/L, 15.27 mg/L and 14.24 mg/L respectively. From the bootstrap the improved WRMSPE for drug 1 drug 2, drug 3, drug 4 and simulated data set was 9.35 mg/L, 19.85 mg/L, 0.50 mg/L, 14.44 mg/L and 13.98mg/L respectively. The bootstrap provided estimates of WRMSPE that corresponded to the external validation methods. From the results obtained, it was concluded that both the PPC and the Bootstrap were demonstrated to have value as validation tools.