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Der Wilms Tumor (WT), auch Nephroblastom genannt, zählt zu den im Kindesalter am häufigsten auftretenden malignen Tumoren und entsteht meist unilateral (90 – 95 %) und sporadisch (98 – 99 %). Leider sind bis heute die molekularen Ursachen, die zur Entwicklung dieser Tumoren führen nur unzureichend aufgeklärt. So werden bisher nur drei Gene mit dem Auftreten von WT in Verbindung gebracht: WT1, CTNNB1 und WTx. Während WT1 und CTNNB1 jeweils Mutationsraten von etwa 10 – 15 % aufweisen, die zudem häufig gemeinsam vorliegen, werden für WTx Mutationsraten von etwa 30 % beobachtet. Die genetischen Alterationen der anderen Tumoren sind noch immer komplett unbekannt. Ziel dieser Arbeit war aus diesem Grund die Identifikation von relevanten Regionen und Genen, die an der Entstehung bzw. dem klinischen Fortschreiten von Wilms Tumoren beteiligt sind. Zusätzlich sollten weitere Untersuchungen zur Einschätzung ihres prognostischen Potenzials dienen. In einem ersten Ansatz wurden die Chromosomenbereiche 11q und 16q in einer großen Anzahl von Wilms Tumoren auf LOH (=loss of heterozygosity), d.h. den (partiellen) Verlust von genetischem Material, untersucht. In beiden Fällen wurden erhöhte LOH-Raten von etwa 20 % beobachtet, jedoch war keine Eingrenzung der relevanten Regionen möglich, da Allelverluste nicht stets ab einem bestimmten Marker beobachtet wurden. Ein Vergleich mit der Histologie ergab signifikante Assoziationen der Allelverluste mit anaplastischen und Mischtyp-Tumoren (nur für LOH 11q), wohingegen kaum LOHs in epithelialen und stromareichen Tumoren festgestellt wurden. Somit scheinen auf 11q und 16q Gene vorzuliegen, die einerseits die Differenzierung in Epithel und Stroma begünstigen oder andererseits ein blastemreiches und anaplastisches Erscheinungsbild verhindern. Jedoch könnte auch die Assoziation von bestimmten Subtypen mit LOH 11q und 16q auf eine Entstehung aus unterschiedlichen Zellen hindeuten. Weiterhin war das Auftreten von LOH, v.a. wenn jeweils der komplette Chromosomenarm betroffen war, mit einem erhöhten Rezidiv- und Sterberisiko (nur LOH 11q) verbunden. Somit konnte gezeigt werden, dass LOH-Untersuchungen auf 11q und 16q zur Identifikation von Hochrisikopatienten für die Entwicklung von Rezidiven bzw. erhöhter Mortalität eingesetzt werden können, wodurch eine individuelle Anpassung der Therapiemaßnahmen ermöglicht wird. In einem zweiten Ansatz wurden eine Reihe von bereits publizierten potenziellen Markergenen in einer großen Anzahl von Wilms Tumoren mit Hilfe der Realtime RT-PCR auf ihre Relevanz überprüft. Allen diesen Genen wurde zuvor eine Funktion bei der histologischen Klassifikation der Tumoren bzw. bei der Vorhersage bestimmter klinischer Verläufe zugeschrieben. Die univariate Analyse diente der Beurteilung der Relevanz einzelner Gene, wohingegen die multivariate Analyse zur Bestimmung von prognostischen Genkombinationen eingesetzt wurde. Anschließend erfolgte die Validierung mittels eines zweiten und unabhängigen Tumorsatzes. Auch wenn viele der bereits publizierten Marker und in der ersten Analyse erhaltenen Assoziationen in einem weiteren und unabhängigen Tumorsatz nicht verifizierbar waren, konnten dennoch einige frühere Ergebnisse repliziert und die Relevanz der entsprechenden Gene nachgewiesen werden. Neben der Verbindung der Repression von HEY2 und TRIM22 mit Hochrisikotumoren bzw. einer höheren Sterbewahrscheinlichkeit fanden sich schwach signifikante Assoziationen auch für die verminderte Expression von TRIM22 und VEGF mit der Histologie. Ebenso waren erhöhte Level von TERT und die Repression von TRIM22 mit der Entwicklung eines Rezidivs verbunden. Vor allem aber die Korrelation der Repression von HEY2 und VEGF sowie einer Überexpression von CA9 mit Rezidiven, Tumoren hoher Malignität oder primären Metastasen verweisen auf die Notwendigkeit, besonders die Hypoxie- und Angiogenese-Signalkaskaden in Wilms Tumoren zu untersuchen, um deren Einfluss v.a. auf das Fortschreiten und die Ausbreitung der Tumoren zu evaluieren. Auch wenn die multivariate Analyse nicht zu relevanten Genkombinationen führte, konnte hier dennoch eine schwache Assoziation der verminderten Expression von TOP2A und TRIM22 mit primären Metastasen oder einer erhöhten Mortalität, sowie der Überexpression von TERT mit der Rezidivbildung bestätigt werden. Interessanterweise stellte sich die Histologie, die derzeit das Hauptkriterium für die Risikoklassifikation darstellt, weder als geeigneter prognostischer Marker für die Beurteilung des Rezidiv- noch des Sterberisikos heraus. Somit sollten Realtime RT-PCR Analysen in Zukunft als weiterer Faktor zur Beurteilung des Rezidiv- und Sterberisikos eingesetzt werden, um eine individuelle Anpassung der Therapie zu ermöglichen. Basierend auf den Ergebnissen der Realtime RT-PCR Analyse wurde der Einfluss der Expression ausgewählter Gene auf Primärkulturen, die aus nativem Wilms Tumormaterial gewonnen wurden, untersucht. Nach der Überexpression von HEY2, EGR1, MYCN und TRIM22 wurden bei allen Zellen hohe Sterberaten beobachtet, v.a. bei HEY2 und EGR1. Leider konnte weder für HEY2 noch für EGR1 der Grund hierfür aufgeklärt werden, allerdings war bei EGR1 weder die Apoptose noch die Seneszenz beteiligt. Im Gegensatz hierzu wurde die Apoptose als entscheidender Mechanismus bei MYCN und v.a. TRIM22 ermittelt. Außerdem scheint bei MYCN ein großer Anteil an Zellen in die Seneszenz einzutreten. Auch wenn diese ersten Untersuchungen an Primärkulturen von Wilms Tumoren eindeutig die Relevanz dieser Gene für die Entwicklung bzw. das Fortschreiten der Tumoren bestätigten, so sind trotz alledem weitere Experimente v.a. in einer größeren Anzahl genetisch unterschiedlicher Primärkulturen nötig, um das endgültige Potenzial dieser Gene aufzuklären. / Wilms tumor (WT), also called Nephroblastoma, belongs to the most common malignant tumors occurring in childhood. Most of these Wilms tumors develop unilaterally (90 – 95 %) and sporadically (98 – 99 %). Unfortunately, only little is known about the molecular background underlying their development with only three genes known so far: WT1, CTNNB1 and WTx. Mutations in WT1 and CTNNB1 occur only in a minor fraction of Wilms tumors of about 10 - 15 % and are often associated with each other, while mutations in WTx can be found in about 30 % of tumors. The genetic alterations of the other tumors are completely unknown. Hence, the aim of this thesis was to identify important regions and genes that are involved in Wilms tumor formation and/or progression and to further characterize their potential as markers for the prediction of certain clinical outcomes. First, chromosome arms 11q and 16q were screened for LOH (= loss of heterozygosity), which means the (partial) loss of the genome in a cell, in a large cohort of Wilms tumors. In both regions LOH rates of about 20 % were detected, but since allele losses did not always start at the same marker in the different tumors it was not possible to delimit any relevant subregions. Since there were significantly higher rates of allele loss in anaplastic and mixed-type (only 11q) tumors and almost no allele loss in epithelial and stromal tumors, 11q and 16q must contain genes that either facilitate the epithelial and stromal differentiation of cells or hamper the appearance of blastemal and anaplastic phenotypes. Otherwise, the obvious possibility to discriminate these histological subtypes by allele loss on 11q and 16q might be explained by a development from different precursor cells. Higher rates of LOH could also be linked to higher risks for relapse and death (11q only), especially when the whole chromosome arms were involved. Therefore, investigation of LOH on 11q and 16q may help to adjust the therapeutic regimens by identifying high-risk patients for relapse and death. A second approach was to reinvestigate the expression of a number of published marker genes in a larger set of Wilms tumors with a uniform method, the realtime RT-PCR. All of the genes were suggested to facilitate classification and/or prediction of certain clinical outcomes. Univariate analysis was performed to screen for relevant genes, followed by multivariate analysis to search for predictive gene combinations. Finally, validation of associations found in the first cohort was performed in a second and independent tumor set. Unfortunately, many of the previously published markers as well as associations of the first tumor set could not be verified in a new and independent tumor set. Nevertheless, it was possible to replicate the results of a number of genes and evidence their prognostic relevance. These included the repression of HEY2 and TRIM22 for high-risk tumors or mortality. Weaker correlations were verified for the repression of TRIM22 and VEGF with the histological risk and for overexpression of TERT and repression of TRIM22 with later relapse. Since the weaker expression of HEY2 and VEGF as well as the overexpression of CA9 was significantly linked to relapse, high malignant tumors or metastasis the hypoxia / angiogenesis pathways should be investigated in Wilms tumors especially with regard to the progression and spreading of tumors. Finally, multivariate analysis substantiated a weak association of repression of TOP2A and TRIM22 with metastasis or death and of overexpression of TERT with subsequent relapse. Most interestingly, histology, the current gold standard used for prediction of risks for relapse and death, could not be verified as potent prognostic factor for neither of them. Hence, realtime RT-PCR analyses can aid in stratification of tumors and prediction of relapse and death risks to intensify therapy for high-risk patients on one hand and to reduce therapy and side-effects in low-risk patients on the other hand. Based on the results of the realtime RT-PCR analyses the expression of several genes was ascertained in primary cell cultures cultivated from native Wilms tumor material. Overexpression of MYCN, TRIM22 and especially HEY2 and EGR1 by viral transduction resulted in high rates of cell death. Unfortunately, the underlying mechanism of death could be determined neither for HEY2 nor for EGR1, though for EGR1 the involvement of apoptosis and senescence could be excluded. In contrast, death in MYCN and especially in TRIM22 overexpressing cells could be attributed to high rates of apoptosis. Furthermore, a large fraction of MYCN cells seem to enter cell senescence and stop to proliferate. These results clearly corroborate the proposed relevance of the investigated genes in the development and/or progression of Wilms tumors. Nevertheless, further experiments in different primary cell cultures of Wilms tumors are necessary to clarify the real potential of these genes.

The work presented in this thesis details a novel engine evaluation approach utilising real-time simulation and advanced engine testing systems for general applicability to new generations of air charging systems. A novel engine air charging system including a charge air handling unit (CAHU) has been developed which is connected to an engine to emulate advanced boosting system conditions. Significant analytical and development work has focused on generating a real-time turbocharger model such that the CAHU can be effectively controlled to emulate the turbocharger performance under both steady and pulsating conditions. Experimental work was carried out to evaluate this new engine air charge testing system against a production turbocharged baseline engine. The accuracies with respect to the boost pressure, turbocharger speed, mass air flow, and fuel consumption in the steady state tests are above 95%, and the level of confidence for the engine back pressure is approximately 90%. The difference of turbocharger speed between the steady turbocharger model and the pulsation model is also identified. In engine transient tests, the boost pressure and engine torque have shown fast response characteristics similar to that of the baseline engine. While general transient trends were achieved, some issues were identified with the high speed control of the CAHU interacting with the dSPACE real time turbocharger model. It is proposed that future improvement can be achieved via applying new control algorithms to improve the accuracy and tracking the CAHU control without increasing the system instabilities.

The individual soft tissues in the human body, such as liver, prostate, thyroid and breast, can each be characterized by a set of mechanical properties. Among these properties, the stiffness, or Young’s modulus, is of particular interest, as disease processes or abnormal growths introduce changes in the tissue stiffness. For example, cirrhosis is associated with an increase in stiffness in the affected region(s) of the liver, and the severity has a strong positive correlation with the measured liver tissue stiffness. Although the conventional ultrasound image is produced by changes in acoustic properties, most notably acoustic impedance (equal to density times sound speed), it is in fact possible to measure tissue strain ultrasonically, by performing ultrasound imaging while the tissue region of interest is mechanically perturbed. Although in principle incorrect, such strain imaging methods are commonly referred to as ultrasound elastography imaging. While tissue strain can reveal the presence of stiffness changes, its diagnostic value is limited due to the inability to reveal the magnitude of the stiffness change. Still, strain imaging is a feature on several commercial scanners. There does in fact exist an elegant, but complex and quite expensive, quantitative ultrasound method of imaging the elasticity of soft tissues, called Supersonic Shear imaging (SSI). However, a much lower cost method of quantitatively imaging tissue elasticity would be useful, especially if the method can be implemented with only minor modifications to existing ultrasound scanner design. This dissertation research deals with an attempt of designing and testing such a method. Ultrasound elastography encompasses a number of diverse techniques, roughly categorized by the mechanical perturbation method into two main groups: quasi-static and dynamic methods. Dynamic elastography requires a vibrating source, either separate or integrated with a transducer, making the imaging system cumbersome, especially for the portable systems. Quasi-static elastography only requires conventional ultrasound hardware, however current techniques remain qualitative with unknown stress distribution. This dissertation focuses on the investigation of free hand quantitative quasi-static elastography, aiming to real time assessment. Our proposed low cost real-time ultrasound elastography system is based on determining an axial strain and an axial stress over a region of interest, i.e., an axial strain image and an axial stress image are required. By taking the axial stress/axial strain ratio for each pixel in the image, an actual elasticity image is established. To achieve this goal, our system needs to ultrasonically measure the mechanical strain fast and accurately over a specified image plane; likewise, the system needs to be able to calculate the mechanical stress over the same image plane in real time. Now, the stress imaging will require us to apply a quasi-static force function and also to be able to quantify this force function. There are two major research efforts we have made to implement a low cost real-time ultrasound elastography system. The first important topic of this dissertation involves the development of a novel displacement and strain estimator based on analytical phase tracking (APT), which has been demonstrated to give better performance in terms of accuracy, resolution and computational efficiency (approximately 40 times faster than the standard time domain cross correlation method). The second important topic is the stress field reconstruction, with efforts in: 1) integrate force sensors into a single linear array transducer probe, with the goal of quantifying the applied force function; 2) propose a superposition method based on Love’s analytical equation to calculate the stress distribution, where this solution is computationally fast enough to allow real time stress field estimation; 3) analyze the accuracy of the proposed stress method using finite element analysis as a reference on different simulated phantoms. The final objective is to combine the strain and stress information together for quantitative elastography. Correspondingly, we have implemented experiments to evaluate the method on homogeneous and inhomogeneous phantoms of various types. Results show that this method is able to distinguish medium with different stiffness. We have conducted experiments to study the feasibility and improve the accuracy of this estimation technique based on phantoms with known elasticity. In principle, such a technique could be used to image the distribution of Young’s modulus under quasi-static compression, with specific applications to medical imaging.

Software used in medical settings operate in complex and variable environments. Programs need to integrate well not only with their electrical and mechanical components, but also within the socio-technological setting they participate in. In this Master's Thesis, a modular software architecture for controlling surgical robot systems within magnetic resonance scanners is designed and implemented. The C++ program runs on a sbRIO 9651 real-time operating system and an object oriented design is taken. Robot kinematics and controls are put into effect in software and validated. Communication with up to ten daughter cards occurs via SPI and external information is exchanged via OpenIGTLink. A web-based engineering console made with ReactJS is also constructed to provide a visual interface for actuating motor axes and executing robot functionality. Documentation of the code is provided and the program was validated quantitatively with software tests and qualitatively through experimentation in MRI suites.

The Real-time Specification of Java (RTSJ) is an intention to introduce Java as a language for developing real-time system. However, the complexity of their development and a non-trivial programming model of RTSJ with its manual memory management often lead to programming errors. To mitigate the development of RTSJ systems it would be beneficial to provide an internal domain-specific language (DSL) extending the Java language which would allow to develop the systems in more intuitive and safer way. However, it is needed to find compromise between solution's power and level of usability, because this two attributes go often against each other. One possible way of DSLs creation concerns the Meta-Programming System (MPS). It allows to develop new domain-specific languages and corresponding projectional editors enabling different views on code. This thesis proposes a design and implementation of the DSL on the top of the MPS platform and corresponding code generator enabling development of RTSJ systems. Furthermore, the thesis provides a simple case-study to evaluate a proposed DSL. Additionally, the thesis assesses the suitability of MPS as a DSL-development platform.

Doctor of Philosophy(PhD) / Recent advances in miniaturization and low power design have led to a flurry of activity in wireless sensor networks. Sensor networks have different constraints than traditional wired networks. A wireless sensor network is a special network with large numbers of nodes equipped with embedded processors, sensors, and radios. These nodes collaborate to accomplish a common task such as environment monitoring or asset tracking. In many applications, sensor nodes will be deployed in an ad-hoc fashion without careful planning. They must organize themselves to form a multihop, wireless communication network. In sensor network environments, much research has been conducted in areas such as power consumption, self-organisation techniques, routing between the sensors, and the communication between the sensor and the sink. On the other hand, real-time communication with the Quality of Service (QoS) concept in wireless sensor networks is still an open research field. Most protocols either ignore real time or simply attempt to process as fast as possible and hope that this speed is sufficient to meet the deadline. However, the introduction of real-time communication has created additional challenges in this area. The sensor node spends most of its life routing packets from one node to another until the packet reaches the sink; therefore, the node functions as a small router most of the time. Since sensor networks deal with time-critical applications, it is often necessary for communication to meet real time constraints. However, research that deals with providing QoS guarantees for real-time traffic in sensor networks is still in its infancy.This thesis presents a real-time communication framework to provide quality of service in sensor networks environments. The proposed framework consists of four components: First, present an analytical model for implementing Priority Queuing (PQ) in a sensor node to calculate the queuing delay. The exact packet delay for corresponding classes is calculated. Further, the analytical results are validated through an extensive simulation study. Second, report on a novel analytical model based on a limited service polling discipline. The model is based on an M/D/1 queuing system (a special class of M/G/1 queuing systems), which takes into account two different classes of traffic in a sensor node. The proposed model implements two queues in a sensor node that are served in a round robin fashion. The exact queuing delay in a sensor node for corresponding classes is calculated. Then, the analytical results are validated through an extensive simulation study. Third, exhibit a novel packet delivery mechanism, namely the Multiple Level Stateless Protocol (MLSP), as a real-time protocol for sensor networks to guarantee the traffic in wireless sensor networks. MLSP improves the packet loss rate and the handling of holes in sensor network much better than its counterpart, MMSPEED. It also introduces the k-limited polling model for the first time. In addition, the whole sending packets dropped significantly compared to MMSPEED, which it leads to decrease the consumption power. Fourth, explain a new framework for moving data from the sink to the user, at a low cost and low power, using the Universal Mobile Telecommunication System (UMTS), which is standard for the Third Generation Mobile System (3G). The integration of sensor networks with the 3G mobile network infrastructure will reduce the cost of building new infrastructures and enable the large-scale deployment of sensor networks

Neurturin (NTN) belongs to the glial cell-line derived neurotrophic factor (GDNF) family of growth factors. Both NTN and GDNF have been shown to potently prevent the degeneration of dopaminergic neuron in vitro and in vivo. The GDNF family receptor alpha 2 (GFRα-2) is the preferred receptor for NTN. In addition to the known full-length isoform (GFRα-2a), we have previously reported the isolation of two novel alternatively spliced isoforms (GFRα-2b and GFRα-2c). The expression levels of these isoforms have yet to be quantified and the functional properties determined. In this report, we have developed a real time polymerase chain reaction (PCR) using SYBR Green I to detect the expression levels of the three splice variants (GFRα-2a, GFRα-2b and GFRα-2c) in murine tissues. Both GFRα-2a and GFRα-2c were expressed at similar levels in all tissues examined. GFRα-2b was found to be 10 fold lower in expression. All three isoforms activated MAPK (ERK1/2) and Akt. Transcriptional profiling with DNA microarrays demonstrated that the spliced isoforms do not share similar profiles. In conclusion, we have now shown the expression levels of the spliced variants. All three isoforms are functional. However, each isoform appeared to have unique transcriptional profiles when activated. / Singapore-MIT Alliance (SMA)

Thesis (Ph. D.)--University of Texas at Austin, 2006. / Vita. Includes bibliographical references.

Steroid hormones, such as estrogen, are produced in one tissue and carried through the blood stream to target tissues in which they bind to highly specific nuclear receptors and trigger changes in gene expression and metabolism. Industrial chemicals, such as bisphenol A and many agricultural chemicals, including permethrin and fervalerate, are known to have estrogenic potential and therefore are estrogen mimics. Widely used agricultural chemicals, Enable (fungicide) and Diazinon (insecticide), were evaluated to examine their toxicity and estrogenicity. MCF-7 cells, an estrogen-dependent human breast cancer line, were utilized for this purpose. MCF-7 cells were treated with 0.033-3.3 ppb (ng/ml) of Enable and 0.3-67 ppm of Diazinon and gene expression was compared to that in untreated cells. Microarray analysis showed down-regulation of eight genes and up-regulation of thirty four genes in cells treated with 3.3 ppb of Enable, compared to untreated cells. Similarly, in cells treated with 67 ppm of Diazinon, there were three genes down-regulated and twenty seven genes up-regulated. For both chemicals, specific genes were selected for special consideration. RT-PCR confirmed results obtained from analysis of the microarray. These studies were designed to provide base-line data on gene expression-altering capacity of specific chemicals and will allow assessment of the deleterious effects caused by exposure to the aforementioned chemicals.