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A brief analysis of the nonresonant-coupled parallel resonant converter is performed. The converter is modeled and a reference classical analog controller is designed and simulated. Infrastructure required for digital control of the converter (including anti-aliasing filters and a modulator) is designed and a classical digital controller is designed and simulated, yielding a ~30% degradation in control bandwidth at the worst-case operating point as compared with the analog controller. Based on the strong relationship observed between low-frequency converter gain and operating point, a gain-scheduled digital controller is proposed, designed, and simulated, showing 4:1 improved worst-case control bandwidth as compared with the analog controller. A complete prototype is designed and built which experimentally validates the results of the gain-scheduled controller simulation with good correlation. The three approaches that were investigated are compared and conclusions are drawn. Suggestions for further research are presented. / Master of Science

Chemokines are chemotactic cytokines which play an important role in the migration of immune cells to distant tissues or compartments within tissues. These proteins have also been demonstrated to play a major role in cancer metastasis. The C-C chemokine receptor type 7 (CCR7) is a member of the chemokine receptor family. CCR7 along with its ligands CCL19 and CCL21 plays an important role in innate immune response by trafficking of lymphocytes. In cancer, tumour cells expressing CCR7 migrate to lymphoid organs and thus disseminate to other organs. Neutralizing the interactions between CCL21/CCR7 would therefore be expected to inhibit the progression and metastasis of many different types of cancer to regional lymph nodes or distant organs. Our objective was to identify a potent small molecule antagonist of CCR7 as a prelude to the investigation of the role of this axis in cancer metastasis. In this study, we provided a brief description of chemokines and their role in health and disease with an emphasis on the CCR7/CCL19/CCL21 axis, as well as identification of a CCR7 antagonist “hit”. The potency of the CCR7 antagonist “hit” was optimised by synthesizing different CCR7 antagonist analogues. The “hit” optimization process has led to discover the most active compound amongst a series of different analogues which have the ability to bind and block CCR7 receptor. The efficacy of the most active compound and other analogues were evaluated in vitro using a calcium flux assay which is based on detecting fluorescent light emitted upon release of calcium ions. To identify a suitable cell line, which expresses CCR7 and capably respond to it, amongst a panel of cell lines for in vitro assessment of potency of synthesised compounds, we used Western blot assay and later by flow cytometry assay. The activity and selectivity of the most effective compound against CCR7 receptor was evaluated in vitro by other functional assays such as “configured agarose spot assay” and scratch assay. We first configured the existing under agarose assay to fulfil our requirements and then used it to assess activity and selectivity of compounds. The configured agarose spot assay also describes the application of the agarose spot for evaluation of cells chemotactic response to multiple chemokines under identical experiment conditions.

Directional couplers are passive components in radio frequency (RF) engineering and have a broad set of applications. With the scope of how a directional coupler can be implemented in a design, it is important to have specified goals in mind when designing a coupler, for the component to be precise and behave in a desired way. Different design variations also have benefits and disadvantages, and in this project several variations of couplers were investigated. The limiting factors in this project were the rather small area to work with, combined with the design requirements. This is ultimately what made the design approach unique and the reason for using an iterative process in a simulation software, where conventional methods of designing could not be used. Out of the different designs that were tested, two directional couplers on the RO4350B substrate with a gap of 0.34 mm between the transmission lines fulfilled every design parameter except the one regarding the coupling factor. There are three notable conclusions that were drawn. The first is that all directivity compensation methods that were evaluated are valid after optimizing relevant parameters. Some methods could be combined to have an even greater effect, such as the soldermask and wiggly line methods. The second is that ground vias can affect the directivity of a directional coupler substantially. Designing a coplanar waveguide coupler can also have benefits in respect to the given design requirements in this project. Thirdly, a lossy material such as FR4 could be used and still achieve the design parameters at these frequencies.

Introduction Adhesion GPCRs (aGPCRs) constitute the second largest family of the GPCR superfamily, and yet their properties are also the least understood. Growing research on the biological functions of aGPCRs suggest their implications in various (patho)physiological processes, such as cell migration, organ development and cancers. Moreover, due to the unique architecture of a large extracellular region (ECR) containing a plethora of adhesion motifs, aGPCRs are vital as a mechanosensor which transduces extracellular mechanical stimuli into intracellular signal transduction. One distinct feature of aGPCRs among the GPCR superfamily is the possession of a conserved extracellular fold termed GPCR autoproteolysis-inducing (GAIN) domain in perhaps all members within the class. The cleavage at the last loop of the GAIN domain leads to the formation of two non-covalently associated N- and C-terminal fragments (NTF/CTF). A peptide stretch in the start of the CTF acts as a tethered agonist (TA) which is responsible for at least part of the signaling volumes of an activated receptor. Despite the strict conservation of the GAIN domain and its importance in the activation mechanism of aGPCRs, some other fundamental properties of the receptors, with reference to GAIN domain cleavage, have not been rigorously analysed in a biological context. Thus, this study aims to: 1. Explore the structural and molecular determinants that affects GAIN domain cleavage; 2. Investigate the consequences of GAIN domain cleavage towards (i) surface trafficking, and (ii) phosphorylation of receptors. Results Abolishment of GAIN domain cleavage in Polycystin-1, the only other protein family possessing the GAIN domain, was found to eliminate its surface expression, which is a cause of polycystic kidney/liver disease. However, whether such relationship is also true for aGPCRs has not been systematically analysed. Therefore, the study started with profiling the kinetics of surface delivery of several members of aGPCRs. Mutations on the -2 or +1 residues of the GPCR proteolytic site (GPS) (thereby abolishing GAIN domain cleavage) affected the steady-state surface and total expressions of the receptors differently, and had variable effect towards different receptor members. However, the observations from steady-state kinetics are also a resultant output from numerous processes involved in proteostasis. To further dissect whether GPS mutations affect the surface trafficking of the receptors, a pulse-chase assay called the ‘Retention Upon Selective Hook’ (RUSH) assay was employed, wherein the synthesised receptor molecules conjugated to a streptavidin-binding peptide are withheld in the ER by the co-expressed, ER-resident streptavidin, and are only released upon the addition of biotin that outcompete the receptor-streptavidin binding, creating a synchronised transport. By adapting the RUSH assay on some aGPCR members, the attenuation of surface trafficking by GPS mutations has become more apparent. The tested receptors were found to have a deficit in the quantity of surface population, rather than a change in rate of trafficking, upon the introduction of GPS mutations. This implies that the cells may utilise GAIN domain cleavage as a quality checkpoint for ER exit of aGPCRs. As the GAIN domains of at least some aGPCRs were found to be cleaved before ER exit, and as the rate of surface delivery was generally not affected by GAIN domain cleavage, the influence of GAIN domain cleavage may arise earlier during the receptor maturation in the ER. However, while the mechanisms of GAIN domain cleavage have been elucidated previously, they rely heavily on purified domains. The fundamental questions of when exactly the GAIN domain is cleaved and what additional determinants apart from the GPS sequence contribute to GAIN domain cleavage during receptor biogenesis have still not been answered. In combination with molecular dynamics (MD) simulation studies on the GAIN domain of rat isoform of ADGRL1, F803 was found to be crucial in the proteolysis by forming an edge-π interaction with H836 (-2 position of the cleavage site), such that H836 is in close proximity to the hydroxyl group of T838 for the initiation of the nucleophilic attack. Reconstruction of the edge-π interaction into ADGRB3, a naturally uncleavable receptor, partially reinstates its GAIN domain cleavage; but similar reintroduction on ADGRB2 has no effect on restoring the proteolysis. Nonetheless, this observation highlights the vitality of a proper folding of the GAIN domain, specifically the microenvironment of the cleavage site, in assisting in cleavage. The study continued with a systematic series of experiments that ultimately discover the roles of the CTF towards GAIN domain cleavage of aGPCRs. Firstly, to mimic the biogenesis of the receptor, the seven transmembrane (7TM) region of ADGRE2 (E2) was stepwisely truncated and then analysed for GAIN domain cleavage. It was observed that the extent of GAIN domain cleavage increases when the ECR of E2 precedes with more number of TMs. The proteolysis occurs, although less efficiently, as early as the first TM is synthesised. Interestingly, GAIN domain cleavage is unaffected when the TM region of the E2-1TM mutant was replaced by other single-pass TM, and whether it is trafficked to the surface or held in the ER, while the proteolysis of TM-less ECR mutants is largely impeded. Based on this observation, the ECR and the TM region was spaced either by a fluorophore moiety or a variable number of helical turns. Remarkably, the extent of GAIN domain cleavage of all tested receptors declined upon the increase in displacement with the lumenal side of the ER membrane, defining the importance of membrane proximity in the completion of proteolysis during the maturation of GAIN domain. In that, a new model of GAIN domain cleavage during biogenesis has been proposed, with appreciation of the GAIN domain as part of a higher-order stuctural organisation rather than an independent domain. A physiological extent of GAIN domain cleavage does not only require the folding of the GAIN domain, but also the membrane tethering property of the CTF, allowing a partial cleavage as little as one TM is generated, and a dynamic stability provided by the full CTF. In some aGPCRs, the contributions from CTF are more significant than the autonomous GAIN domain folding. The findings implicate more complex requirements for GAIN domain cleavage in a biological context, and hence supporting a possibility that GAIN domain cleavage is the rate-determining step for ER exit of the receptor, leading to the observations obtained in the kinetic study. Phosphorylation of L3 by PKC activated by distant signaling cascade(s) The last part of the study focused on characterising the mechanism of phosphorylation of ADGRL3 (L3) at Thr1140 (pT1140), which is a poorly explored field of aGPCRs. It was made possible by exploiting a phosphospecific antibody developed in collaboration. Coincidently, pT1140 was not dependent on the examined GPCR properties of the receptor, such as G protein coupling, dependence of the TA, and GRK-mediated phosphorylation. Instead, by series of pharmacological inhibitions, it was discovered that pT1140 originates from the action of novel PKCs (nPKCs). Co-expression of L3 and dominant-negative mutants or the catalytic domains of individual members of nPKCs reveals that PKC acts as a master regulator of the phosphorylation event, by directly phosphorylating the receptor and priming other members of the nPKCs for pT1140. Finally, possible origins of the PKC activation were explored. It was found that the stimulation of PKC occurs via actin disassembly, which can act downstreams of VEGF-A/VEGFR2 signaling, although the physiological relevance is still yet to be deciphered. Nonetheless, the observations opened up new directions of research in the aspect of crosstalks between different signaling cascades and the possible modulations of the signaling fidelity of aGPCRs. Additionally, the complexity of aGPCR signaling has been clearly demonstrated. Conclusion This study has further defined the importance of GAIN domain cleavage for surface trafficking of aGPCRs, a process crucial for extracellular interactions. Moreover, a novel mechanistic model of GAIN domain cleavage in relevance to biogenesis and maturation of the receptors has been postulated. Characterisation of a site-specific phosphorylation mechanism of L3 has illustrated the potential of complex interactions of aGPCRs with other signaling pathways in cells. The results collectively shed light on the structure-function relationship of aGPCRs, and pave ways for numerous potential areas for explorations in the future.

Metal-Organic Frameworks (MOFs) are porous materials consisting of organic ligands connected by inorganic nodes. Their structural uniformity, high surface area, and synthetic tunability, position these frameworks as suitable active materials to achieve efficient and clean electrochemical energy storage. In spite of recent demonstrations of MOFs undergoing diverse electrochemical processes, a fundamental understanding of the mechanism of electron, proton, and ion transport in these porous structures is needed for their application in electronic devices. The current work focuses on contributing to such understanding by investigating proton-coupled electron transfer, capacitance performance, and the relative contribution of electron and ionic transport in the voltammetry of zirconium-based MOFs. First, we investigated the effects that the quinone ligand orientation inside two new UiO-type metal-organic frameworks (2,6-Zr-AQ-MOF and 1,4-Zr-AQ-MOF) have on the ability of the MOFs to achieve proton and electron conduction. The number of electrons and protons transferred by the frameworks was tailored in a Nernstian manner by the pH of the media, revealing different electrochemical processes separated by distinct pKa values. In particular, the position of the quinone moiety with respect to the zirconium node, the effect of hydrogen bonding, and the amount of defects in the MOFs, lead to different PCET processes. The ability of the MOFs to transport discrete numbers of protons and electrons, suggested their application as charge carriers in electronic devices. With that purpose in mind, we assembled 2,6-Zr-AQ-MOF and 1,4-Zr-AQ-MOF into two different types of working electrodes: a slurry-modified glassy carbon electrode, and as solvothermally-grown MOF thin films. The specific capacitance and the percentage of quinone accessed in the two frameworks were calculated for the two types of electrodes using cyclic voltammetry in aqueous buffered media as a function of pH. Both frameworks showed an enhanced capacitance and quinone accessibility in the thin films as compared to the powder-based electrodes, while revealing that the structural differences between 2,6-Zr-AQ-MOF and 1,4-Zr-AQ-MOF in terms of defectivity and the number of electrons and protons transferred were directly influencing the percentage of active quinones and the ability of the materials to store charge. Additionally, we investigated in detail the redox-hopping electron transport mechanism previously proposed for MOFs, by utilizing the chronoamperometric response (I vs. t) of three metallocene-doped metal-organic frameworks (MOFs) thin films (M-NU-1000, M= Fe, Ru, Os) in two different electrolytes (TBAPF6 and TBATFAB). We were able to elucidate, for the first time, the diffusion coefficients of electrons and ions (De and Di, respectively) through the structure in response to an oxidizing applied bias. The application of a theoretical model for solid state-voltammetry to the experimental data revealed that the diffusion of ions is the rate-determining step at the three different time stages of the electrochemical transformation. Remarkably, the trends observed in the diffusion coefficients (De and Di) of these systems obtained in PF61- and TFAB1- based electrolytes at the different stages of the electrochemical reaction, demonstrated that the redox hopping rates inside frameworks can be controlled through the modifications of the self-exchange rates of redox centers, the use of large MOF channels, and the utilization of smaller counter anions. These structure-function relationships provide a foundation for the future design, control, and optimization of electronic and ionic transport properties in MOF thin films. / PHD / The necessity of implementing new energy storage systems that enable the utilization of clean energy in diverse technologies such as electric vehicles and smart power grids, has generated great research efforts in the field of materials science. In particular, the development of nanoscale-based materials that can be utilized in batteries and supercapacitors is essential for achieving effective and clean electrochemical energy storage. Two of the main desired properties for such materials to be employed as electrodes in energy storage devices are high surface area and the possibility of incorporating redox-active moieties that are able to store electricity. Metal-Organic Frameworks (MOFs) are a relatively new kind of porous materials with high surface area and structural uniformity, consisting of organic ligands connected by inorganic nodes. The application of these materials in charge transport and storage is still in its early stages. Therefore, fundamental understanding of the mechanism of electron, proton, and ion transport in MOFs is necessary for a rational design of these porous structures. In order to contribute to such understanding, the present work is focus on two main concepts: (1) elucidating the effect that the tridimensional orientation of redox moieties inside the MOF could have on the charge storage performance and the ability of the material to achieve proton and electron conduction; and (2) quantifying for the first time the individual relative contribution of electron and ionic transport in MOF materials.

Motivated by the recent commercialization of gallium-nitride (GaN) switches, an effort was initiated to determine whether it was feasible to switch the flyback converter at 5 MHz in order to improve the power density of this versatile isolated topology. Soft switching techniques have to be utilized to eliminate the switching loss to maintain high efficiency at multi-megahertz. Compared to the traditional modeling of zero-voltage-switching quasi-square-wave converters, a numerical methodology of parameters design is proposed based on the steady-state model of zero-voltage switching quasi-square-wave flyback converter. The magnetizing inductance is selected to guarantee zero-voltage switching for the entire input and load range with the trade-off design for conduction loss and turn-off loss. A design methodology is introduced to select a minimum core volume for an inductor or coupled inductors experiencing appreciable core loss. The geometric constant Kgac = MLT/(Ac2WA) is shown to be a power function of the core volume Ve, where Ac is the effective core area, WA is the area of the winding window, and MLT is the mean length per turn for commercial toroidal, ER, and PQ cores, permitting the total loss to be expressed as a direct function of the core volume. The inductor is designed to meet specific loss or thermal constraints. An iterative procedure is described in which two- or three-dimensional proximity effects are first neglected and then subsequently incorporated via finite-element simulation. Interleaved and non-interleaved planar PCB winding structures were also evaluated to minimize leakage inductance, self-capacitance and winding loss. The analysis on the trade-off between magnetic size, frequency, loss and temperature indicated the potential for a higher density flyback converter. A small-signal equivalent circuit of QSW converter was proposed to design the control loop and to understand the small-signal behavior. By adding a simple damping resistor on the traditional small-signal CCM model, it can predict the pole splitting phenomenon observed in QSW converter. With the analytical expressions of the transfer functions of QSW converters, the impact of key parameters including magnetizing inductance, dead time, input voltage and output power on the small-signal behavior can be analyzed. The closed-loop bandwidth can be pushed much higher with this modified model, and the transient performance is significantly improved. With the traditional fix dead-time control, a large amount of loss during dead time occurred, especially for the eGaN FETs with high reverse voltage drop. An adaptive dead time control scheme was implemented with simple combinational logic circuitries to adjust the turn on time of the power switches. A variable deadtime control was proposed to further improve the performance of adaptive dead-time control with simplified sensing circuit, and the extra conduction loss caused by propagation delay in adaptive dead-time control can be minimized at multi-megahertz frequency. / Ph. D. / With the fast development of telecom, computer and network systems, high efficient and small volume power supplies are highly desired. A typical method for achieving high power density involves increasing the frequency and implement soft-switching techniques to minimize loss. Thanks to the recent commercialization of the advanced semiconductor gallium-nitride (GaN) switches, it is feasible to design high density power supplies and cost effective power system. Several challenges including optimization of power converter, high frequency magnetics and implementation of control architecture have been addressed in this dissertation which helps to realize this compact power system. With the implementation of proposed circuit model and seminumerical design procedures for magnetics, a 30W high-frequency isolated DC/DC converter with planar inductor is fabricated to verify the theoretical analysis, which also demonstrates much improved performances.

G protein-coupled receptors (GPCRs) play a major role in physiological functions by transducing extracellular information into intracellular responses, thereby, allowing auto-, para , and endocrine communication. In combination with their ubiquitous expression, GPCRs are of special interest for developing therapeutic approaches. Due to their high targetability, GPCRs are also interesting for obesity research. Even though the prevalence for obesity and its comorbidities like type 2 diabetes mellitus is rapidly rising worldwide, the physiological function of many GPCRs is not well resolved in adipose tissue (AT). Here, the high percentage of orphan GPCRs (65 %), meaning GPCRs characterized by lacking information about an endogenous ligand, signal transduction and/or physiological functions, is an enormous restricting factor to understand AT (patho)physiology. In this study the overall goal was the identification and characterization of GPCRs hitherto unrecognized in AT and adipocyte functionality. Therefore, the first part of this study focused on incorporating a web application for the analysis of publicly available RNA-seq data of human and mouse AT and adipocytes, called FATTLAS. With this application, the GPCRome was analysed to identify GPCRs connected to AT function. Furthermore, those GPCRs were characterized in 3T3-L1, a murine preadipocyte cell line with adipogenic potential. Following, the main results are described: 1. Analysing the GPCR expression patterns in AT of lean individuals and individuals with obesity lead to the identification of a multitude of highly and differentially expressed genes, many of them being already characterized in AT or adipocyte function. Nevertheless, four GPCRs were identified being either highly expressed in both conditions (Gpr146) or differentially expressed in individuals with obesity (Fzd5, Mrgprf, and Ptger2) having an unknown function in AT. For all four receptors an agonist has been previously described. 2. Besides expression in AT, all four receptors are also present in primary and 3T3-L1 (pre)adipocytes. Thus, GPR146, FZD5, MRGPRF, and PTGER2 are suitable for characterizing their role in adipocyte function. 3. Investigating receptor overexpression in the heterologous expression system HEK293T, FZD5, MRGPRF and PTGER2 were found to be expressed at the cell surface, whereas GPR146 was mainly located in endosomes. 4. While analysing receptor signalling in HEK293T cells high basal receptor activity was detected for PTGER2 (Gɑs protein) and GPR146 (Gɑi protein). As the published agonists could not induce receptor activation in 3T3-L1 (pre)adipocytes, siRNA-mediated knockdown was the method of choice for the characterization of endogenously expressed receptors in 3T3-L1 preadipocytes. 5. Receptor knockdown analysis for these four receptors revealed a reduced adipogenesis acting via the PPARγ axis. Here, MRGPRF and PTGER2 act via a cAMP-dependent mechanism. Furthermore, the receptors are involved in preadipocyte viability, which also contribute to the regulation of adipogenesis. As the receptor knockdown in preadipocytes leads to a reduced viability and adipogenesis, adipocyte function was analysed after knockdown in adipocytes. 6. Before the investigation of adipocyte function, the effect of receptor knockdown in mature adipocytes on adipocyte viability was analysed with only Mrgprf knockdown showing an effect. 7. In adipocyte function, MRGPRF was involved in adiponectin secretion most likely by changing intracellular cAMP accumulation. Moreover, GPR146 regulates lipolysis via basal Gɑi-protein signalling. PTGER2 and FZD5 did not show an involvement in the analysed adipocyte functionalities. Taken together, in this study an interactive public database (FATTLAS) was implemented, incorporating publicly available RNA-seq data for AT and adipocytes for improved access and analysis of these complex datasets. Using this database, four hitherto unrecognized GPCRs were identified and their involvement in adipogenesis and AT function was proved. In the second part of this thesis, the subgroup adhesion GPCRs (aGPCRs) of the GPCRome, were analysed. Genome-wide association studies have linked this orphan GPCR class to AT function and metabolic dysfunction (e.g. obesity). However, they have not been thoroughly investigated yet. By performing a comprehensive study, the following main findings about aGPCR expression in AT and adipocytes and about their role in adipogenesis and adipocyte function were revealed: 1. In mice 30 aGPCRs are genomically encoded. 25 receptors were identified to be expressed in subcutaneous AT (scAT), 28 in visceral AT (vAT). Only Emr4, Gpr133 and Gpr116 showed a differential expression between scAT and vAT. 2. Under high-fat diet seven receptors (Lphn1-3, Eltd1, Emr1, Gpr124, Gpr116, and Gpr56) were significantly higher expressed in scAT, whereas in vAT three receptors were upregulated (Emr4, Gpr124, and Celsr3) and four downregulated (Gpr113, Gpr116, Gpr64, and Gpr97). 3. As AT consists of diverse cell types, cell-specific expression of aGPCRs in adipocytes and the stroma vascular fraction (SVF) was determined. Here, four receptors were significantly higher expressed in adipocytes (Lphn2, Gpr125, Gpr111, and Gpr64), six receptors were lower expressed (Emr1, Emr4, Gpr133, Gpr113, and Gpr97, Gpr126). However, most aGPCRs are expressed in adipocytes making them interesting for investigating their role in adipocyte function. 4. In human scAT CELSR1, CELSR2, EMR2, and GPR126 were upregulated comparing lean and obese conditions, GPR64 and GPR97 were downregulated. The expression of aGPCRs in human scAT was comparable to mouse scAT indicating a transferability of aGPCR function in mouse and human samples. 5. Analysing aGPCR expression during adipogenesis in 3T3-L1, three distinct expression patterns were found: no changes (Lphn1-3, Cd97, Gpr124, Gpr125, Gpr116, Gpr56, Gpr64, Gpr97, and Gpr126), steady upregulation (Gpr124 and Gpr126), and downregulation (Cd97, Gpr116, Gpr56, and Gpr64). 6. Knockdown of Lphn2, Gpr124, Gpr125, Gpr116, Gpr64, and Gpr126 in 3T3-L1 lead to a reduced lipid accumulation and droplet size indicating an impaired differentiation into mature adipocytes. 7. Exemplarily, GPR64 was selected to study the role of aGPCRs in adipocyte function due to its significantly reduced expression in obesity and its role in adipogenesis. Knockdown and stimulation by a tethered agonist-derived peptide uncovered Gαs protein-mediated signalling. Furthermore, peptide stimulation resulted in a reduced adiponectin secretion and glucose uptake in 3T3-L1 adipocytes. Lipolysis was induced in 3T3-L1 and primary adipocytes after peptide stimulation indicating a transferability of cell culture experiments to ex vivo analysis. In summary, this comprehensive study describes aGPCR expression in human and mouse AT, adipocytes and SVF in depth. Furthermore, their involvement in adipogenesis under physiological and knockdown conditions was studied in the model cell line 3T3-L1. The functional role of aGPCRs was exemplarily analysed for GPR64. As the model cell line 3T3-L1 is difficult to transfect and knockdown only reduces gene expression, the suitability of CRISPR/Cas technology for generating a receptor knockout (KO) in 3T3-L1 cell line was tested. For validation, the aGPCRs, Gpr64 and Gpr126, were chosen. The main results of the third part of this study are the following: 1. Gpr64- and Gpr126 KO cells were generated using a commercially avaible 3T3-L1 cell line constitutively overexpressing Cas9 (3T3-L1 Cas9). 2. An impaired adipogenesis and adipocyte function was found already in 3T3-L1 Cas9 compared to 3T3-L1 wildtype. Creating a self-made Cas9-overexpressing 3T3-L1 was not feasible. Thus, Cas9-overexpressing 3T3-L1 cells are not suitable for analysing adipocyte function after KO. 3. Evaluating other approaches for CRISPR/Cas9-mediated KO strategies, transient overexpression of Cas9 using plasmid-based methods were not feasible, too. However, introducing Cas9 protein was successful and did not interfere with adipogenesis making this approach the method of choice for CRISPR/Cas9-mediated KO in 3T3-L1. In brief, KO studies in 3T3-L1 cell line using CRISPR/Cas technology should not be carried out using plasmid-based approaches. However, transfecting the Cas9 protein or its ribonucleoprotein complex is feasible to create 3T3-L1 KO cell lines without interfering with adipogenesis and adipocyte function. In conclusion, this thesis provides a comprehensive study about GPCR expression in AT of lean individuals and individuals with obesity. The identification and characterization of GPCRs hitherto unrecognized in AT or even orphan GPCRs supports the understanding of AT (patho)physiology. In particular, the expression of all GPCRs in AT of lean individuals and individuals with obesity was uncovered for human scAT as well as mouse scAT and vAT. Furthermore, three rhodopsin-like GPCRs (GPR146, MRGPRF, and PTGER2), one Fzd-like receptor (FZD5) and the adhesion GPCR class were basally characterized regarding adipogenesis and (pre)adipocyte functionality using the model cell line 3T3-L1. These data are fundamental for understanding the importance of the whole repertoire of GPCR in AT (patho)physiology and can be used as a starting point for their characterization in AT and adipocyte function in depth, possibly even leading to novel therapeutic approaches in the future.

Dans l’industrie pharmaceutique, la granulation sèche par compactage à rouleaux est un procédé d’agglomération de poudres en granulés pour améliorer les propriétés d’écoulement nécessaire pour le procédé de compression en matrice. Comprendre le procédé de compactage à rouleaux et optimiser l’efficacité de production est limitée par l’utilisation de l’approche expérimentale à cause du coût élevé des poudres, le temps des essais et la complexité du procédé. Dans ce travail, une méthode d’éléments finis en 3D, est développée dans le but d’identifier les paramètres critiques du matériau et du procédé pour le contrôle de la qualité de la production. Le modèle de comportement de Drucker-Prager Cap est utilisé pour décrire le comportement en compression de poudres et sa calibration est déterminée à partir des essais standard. Pour surmonter la complexité liée à l’existence de deux mécanismes différents, l’alimentation en poudre par une vis sans fin et le compactage entre les rouleaux, une nouvelle méthode d’interfaçage entre la méthode des éléments discrets (DEM) employée pour décrire l’écoulement dans l’alimentation et la méthode des éléments finis (FEM) utilisée pour le compactage entre les rouleaux est développée. Enfin, pour une modélisation de compactage de rouleaux plus réaliste, prenant en compte la variation de l’entrefer entre les rouleaux, une nouvelle approche de couplage Euler-Lagrange est proposée. Les résultats de simulations par éléments finis montrent clairement l’effet des différents paramètres du procédé sur les distributions de pression et de densité dans la zone de compactage. En outre, les résultats montrent que l'utilisation de plaques de confinement de la poudre entre les rouleaux, développe une distribution de pression et de densité non homogène dans le compact, avec une densité plus élevée au centre et plus faible aux bords. D'autre part, l’utilisation de rouleaux dont l’un est surmonté d’une jante de confinement, a montré une distribution de propriétés globalement plus uniforme sur la largeur du compact avec des valeurs légèrement plus élevées aux bords qu’au centre. La méthodologie combinant les méthodes DEM & FEM montre clairement une corrélation directe entre la vitesse des particules entraînées par la vis dans la zone d’alimentation et la pression du rouleau. Tous les deux oscillent avec la même période. Cela se traduit par un compact anisotrope avec un profile de densité variant de manière sinusoïdale le long de sa largeur. Afin d'étudier la capacité du modèle à prédire les propriétés des compacts produits par compactage à rouleaux, les prédictions par simulations numériques sont comparées aux données de la littérature et validées par des mesures spécifiques. / In the pharmaceutical industry, dry granulation by roll compaction is a process of size enlargement of powder into granules with good flowability for subsequent die compaction process. Understanding the roll compaction process and optimizing manufacturing efficiency is limited using the experimental approach due to the high cost of powder, time-consuming and the complexity of the process. In this work, a 3D Finite Element Method (FEM) model was developed to identify the critical material properties, roll press designs and process parameters controlling the quality of the product. The Drucker-Prager Cap (DPC) model was used to describe the powder compaction behavior and was determined based on standard calibration method. To overcome the complexity involving two different mechanisms of powder feeding by the screw and powder compaction between rolls, a novel combined approach of Discrete Element Method (DEM), used to predict the granular material flow in the feed zone and the Finite Elements Method (FEM) employed for roll compaction, was developed. Lastly, for a more realistic roll compaction modelling, allowing the fluctuation of the gap between rolls, a Coupled-Eulerian Lagrangian (CEL) approach was developed. FEM simulation results clearly show the effect of different process parameters on roll pressure and density distribution in the compaction zone of powder between the rolls. Moreover, results show that using a cheek-plates sealing system causes a nonuniform roll pressure and density distribution with the highest values in the middle and the lowest at the edges. On the other hand, the resultant pressure and density distributions with the rimmed-roll obtained higher values in the edges than in the middle and overall a more uniform distribution. The combined DEM-FEM methodology clearly shows a direct correlation between the particle velocity driven by the screw conveyor to the feed zone and the roll pressure, both oscillating in the same period. This translates into an anisotropic ribbon with a density profile varying sinusoidally along its length. To validate the results, the simulations are compared with literature and experimentally measured values in order to assess the ability of the model to predict the properties of the produced ribbons.

In this thesis the application of novel tensor decomposition and tensor representation techniques in highly accurate post Hartree-Fock methods is evaluated. These representation techniques can help to overcome the steep scaling behaviour of high level ab-initio calculations with increasing system size and therefore break the "curse of dimensionality". After a comparison of various tensor formats the application of the "canonical polyadic" format (CP) is described in detail. There, especially the casting of a normal, index based tensor into the CP format (tensor decomposition) and a method for a low rank approximation (rank reduction) of the two-electron integrals in the AO basis are investigated. The decisive quantity for the applicability of the CP format is the scaling of the rank with increasing system and basis set size. The memory requirements and the computational effort for tensor manipulations in the CP format are only linear in the number of dimensions but still depend on the expansion length (rank) of the approximation. Furthermore, the AO-MO transformation and a MP2 algorithm with decomposed tensors in the CP format is evaluated and the scaling with increasing system and basis set size is investigated. Finally, a Coupled-Cluster algorithm based only on low-rank CP representation of the MO integrals is developed. There, especially the successive tensor contraction during the iterative solution of the amplitude equations and the error propagation upon multiple application of the reduction procedure are discussed. In conclusion the overall complexity of a Coupled-Cluster procedure with tensors in CP format is evaluated and some possibilities for improvements of the rank reduction procedure tailored to the needs in electronic structure calculations are shown. / Die vorliegende Arbeit beschäftigt sich mit der Anwendung neuartiger Tensorzerlegungs- und Tensorrepesentationstechniken in hochgenauen post Hartree-Fock Methoden um das hohe Skalierungsverhalten dieser Verfahren mit steigender Systemgröße zu verringern und somit den "Fluch der Dimensionen" zu brechen. Nach einer vergleichenden Betrachtung verschiedener Representationsformate wird auf die Anwendung des "canonical polyadic" Formates (CP) detailliert eingegangen. Dabei stehen zunächst die Umwandlung eines normalen, indexbasierten Tensors in das CP Format (Tensorzerlegung) und eine Methode der Niedrigrang Approximation (Rangreduktion) für Zweielektronenintegrale in der AO Basis im Vordergrund. Die entscheidende Größe für die Anwendbarkeit ist dabei das Skalierungsverhalten das Ranges mit steigender System- und Basissatzgröße, da der Speicheraufwand und die Berechnungskosten für Tensormanipulationen im CP Format zwar nur noch linear von der Anzahl der Dimensionen des Tensors abhängen, allerdings auch mit der Expansionslänge (Rang) skalieren. Im Anschluss wird die AO-MO Transformation und der MP2 Algorithmus mit zerlegten Tensoren im CP Format diskutiert und erneut das Skalierungsverhalten mit steigender System- und Basissatzgröße untersucht. Abschließend wird ein Coupled-Cluster Algorithmus vorgestellt, welcher ausschließlich mit Tensoren in einer Niedrigrang CP Darstellung arbeitet. Dabei wird vor allem auf die sukzessive Tensorkontraktion während der iterativen Bestimmung der Amplituden eingegangen und die Fehlerfortpanzung durch Anwendung des Rangreduktions-Algorithmus analysiert. Abschließend wird die Komplexität des gesamten Verfahrens bewertet und Verbesserungsmöglichkeiten der Reduktionsprozedur aufgezeigt.