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Item Open Access Quasicrystal structures from the crystallographic viewpoint(1988) Gähler, Franz; Fröhlich, J. (Prof. Dr.)Quasikristalle sind neuartige Phasen, die in schnell abgekühlten Metall-Legierungen vorkommen. Ihre wichtigste Eigenschaft ist, dass Ihre Fourier-Transformierte aus scharfen Bragg-Peaks besteht, deren Positionen und Intensitäten eine Punktsymmetrie haben, die mit einer dreidimensionalen periodischen Struktur nicht verträglich ist. Die Positionen der Bragg-Peaks eines Quasikristalls können jedoch alle als ganzzahlige Linearkombinationen von endlich vielen fundamentalen Wellenvektoren geschrieben werden; dies legt nahe, diese Strukturen als Schnitt durch eine höherdimensionale, periodische Struktur aufzufassen.Item Open Access Hochauflösende Beschleunigungssensoren mit Tunnelstrecke(2000) Strobelt, Tilo; Sandmaier, Hermann (Prof. Dr.-Ing.)Ziel der Arbeit ist es, alle zur Konstruktion von hochauflösenden Beschleunigungssensoren mit Tunnelstrecke erforderlichen theoretischen Grundlagen zur Verfügung zu stellen und deren Anwendbarkeit an Hand des Entwurfes und der Realisierung eines Sensors nachzuweisen. Wesentliche Themen des theoretischen Teiles der Arbeit sind hierbei: die Beschreibung der Funktion einer Tunnelstrecke sowie die Herleitung der Rauschanteile, welche die Auflösung von Beschleunigungssensoren beschränken. Es wird deutlich, dass die hohe Auflösung des Tunneleffektes den Einsatz einer aktiven Regelung und hochempfindlicher Stellelemente erforderlich macht. Vergleicht man alle in Beschleunigungssensoren auftretenden Rauschquellen, so stellt man fest, daß die Nachweisgrenze hochauflösender Beschleunigungssensoren vom thermische Rauschen des Feder-Masse-Systems bestimmt wird. Auf den hergeleiteten konstruktiven Grundlagen aufbauend, wird anschließend die Entwicklung eines Sensors mit einer Nachweisgrenze von 1E-6 g (g= Erdbeschleunigung) beschrieben. Hierbei wird die mikrotechnischen Konstruktion dargestellt und ausführlich auf die Entwicklung der erforderlichen Sensorelektronik eingegangen. Um die elektronische Regelung geeignet auslegen und optimieren zu können, wurde das dynamische Verhalten des Gesamtsystems analytisch beschrieben. Die geeignete Wahl der Parameter wurde mit Hilfe eines numerischen Modells des Gesamtsystems nochmals überprüft. Im Rahmen dieser Arbeit wurde ferner ein Messplatz entwickelt, auf dem die Beschleunigungssensoren nach ihrer Fertigstellung charakterisiert worden sind. Erstmalig wurde es so möglich, derartige Sensoren direkt an ihrer Auflösungsgrenze zu vermessen. Die dem Rauschen äquivalente Beschleunigung wird mit ca. 2.5E-6 g gemessen, stimmt also mit den theoretischen Vorhersagen gut überein.Item Open Access Mathematical modeling of signal transduction pathways in mammalian cells at the example of the EGF induced MAP kinase cascade and TNF receptor crosstalk(2004) Schoeberl, Birgit; Gilles, Ernst Dieter (Prof. Dr.-Ing. Dr. h.c. mult.)In this work we apply mathematical modeling to signal transduction networks in mammalian cells. In particular, we have developed models for a survival pathway, the EGF induced MAP kinase cascade and an apoptotic signal trans-duction network the TNF receptor crosstalk. The models presented here are based on and were validated with own experimental data. In the field of signal transduction the major proteins involved and their interactions are fairly well known and biochemically characterized . One characteristic of signal transduction networks is that they are highly interconected by positive and negative feedbacks. Therefore, the dynamics of these networks can not be understood by intuition alone. Mathematical modeling has proven to be a valuable tool in engineering that deals effectively with complexity. In both casese were able to verify hypotheses, which were obtained by the models, experimentally. This work shows that mathematical modeling in combination with quantitative experimental data can give new insights into the potential mechanisms of intracellular signal transduction and regulation.Item Open Access Whisker formation on Sn thin films(2010) Sobiech, Matthias Lukas; Mittemeijer, Eric Jan (Prof. Dr. Ir.)The system Sn on Cu will usually be applied for interconnection of modern electronic systems, i.e. for mechanical, thermal and electrical “integration” of electronic components (e.g. composed of Cu) on rigid substrates (i.e. printed circuit boards) by (e.g. Sn) solder-joint technology. Nowadays Sn is the material of choice for this purpose because the up to now commonly and successfully used SnPb alloys for soldering and coating applications are prohibited by law since 1st July 2006 due to environmental concerns (Pb-free and “green” legislation). However, it is well known since nearly 60 years that pure Sn thin films deposited on Cu substrates are very prone to spontaneous formation of needle-like Sn single-crystals, called whiskers, during ageing at room temperature. Such filamentary Sn whiskers exhibiting growth rates of about 1 Å/sec constitute an issue of great technological relevance for Sn coated leadframe legs of modern microelectronic devices because whisker-induced short-circuit failures of various electronic devices have resulted in enormous financial damage including breakdowns of satellites, computer centres and military and medical devices. Unfortunately, profound knowledge on this controversially discussed phenomenon of whisker-growth is still lacking. Therefore, particularly in recent years, the electronic industry promotes scientific activities to arrive at fundamental understanding of Sn whisker formation in order to implement industrially reliable (accelerated) whisker tests and/or mitigation strategies. Against the above background, the present thesis focuses in particular on revealing the driving force for Sn whiskering in the system Sn on Cu during room temperature ageing and thus to devise a coherent understanding of the processes leading to the formation and growth of Sn whiskers. The obtained fundamental interrelations of microstructural evolution, phase formation, residual stress development and the associated whiskering of Sn thin films electro- and sputter-deposited on Cu as well as of SnPb thin films electrodeposited on Cu during ageing at room temperature have lead to a qualitative understanding of whisker growth in terms of localized Coble-creep. On this basis, whisker mitigation strategies can be proposed.Item Open Access Die Rolle des p53-Status für die Sensitivität von Tumoren gegenüber unterschiedlichen p53 Aktivatoren(2014) Weilbacher, Andrea; Peter Scheurich (Prof. Dr.)Die tumorsuppressiven Eigenschaften von p53 gelten als zentral bei der Aufrechterhaltung der genomischen Stabilität. Auf Grund dessen spielt p53 eine Schlüsselrolle bei der Reaktion auf genomischen Stress welcher unter anderem durch klassische Chemotherapeutika, wie beispielsweise Cisplatin, induziert wird. In der vorliegenden Studie wurde die Rolle des p53-Status für die Sensitivität gegenüber p53-aktivierenden Substanzen untersucht. Hierfür wurde eine Auswahl von Zelllinien aus verschiedenen Entitäten und mit unterschiedlichen p53-Genotypen sowie Tumor-assoziierte Fibroblasten aus der humanen Lunge und PBMNCs von gesunden Donoren verwendet. Als p53-Aktivatoren wurden neben dem klassischen DNA-modifizierenden Molekül Cisplatin die direkten Aktivatoren von p53, Nutlin-3 und RITA eingesetzt. Die Behandlung mit Nutlin-3 führte selektiv in wtp53-exprimierenden Zellen zu einem G1-Arrest. Dieser trat auch in primären, nichtmalignen Zellen auf. Nutlin-3 agiert somit selektiv in wtp53-exprimierenden Zellen, nicht aber tumorselektiv. Die wtp53-selektive Wirkungsweise konnte weder nach Cisplatin- noch nach RITA-Behandlung nachgewiesen werden. Beide Substanzen induzierten Zelltod auch in mtp53-Systemen oder im Falle von RITA auch in der p53-null-Zelllinie OVCAR5. Der durch Cisplatin und RITA induzierte Zelltod in der wtp53-exprimierenden Zelllinie NTERA-2D1 konnte auf die Aktivierung von wtp53 zurückgeführt werden. Hingegen war der in den mtp53-exprimierenden Zelllinien OVCAR3 und OVCAR4 induzierte Zelltod im Falle einer Behandlung mit Cisplatin oder RITA unabhängig von mtp53. Zudem führte die siRNA-vermittelte Depletion von p63 und p73 zu keiner Verminderung des Zelltods. Cisplatin und RITA können somit unabhängig von der p53-Superfamilie Zelltod in p53-defekten Systemen induzieren. Dieser war für beide Substanzen auf die Aktivierung der mitochondrialen Effektoren BAX und BAK zurückzuführen. Die Induktion von Zelltod nach Cisplatin-Behandlung konnte weiterhin auf die Aktivierung der pro-apoptotischen Bcl-2-Proteine NOXA und PUMA zurückgeführt werden. Die Analyse der konstitutiven Expression der Bcl-2-Proteine in der gesamten Zelllinienauswahl zeigte eine signifikante Korrelation des Verhältnisses aus pro- und anti-apoptotischen Bcl-2-Proteinen gegenüber der Cisplatin-Sensitivität. NOXA und Bcl-w wurden in diesem Ansatz als prädiktive Marker der Cisplatin-Sensitivität innerhalb der Zellauswahl identifiziert. Die Kombinationsbehandlung von Cisplatin mit dem BH3-mimetic ABT-737 führte zu einer Sensitivierung von Cisplatin-insensitiven Zellen. Im Falle der Behandlung mit RITA konnte keine Korrelation zwischen dem Verhältnis aus pro- und anti-apoptotischen Bcl-2-Proteinen und dem durch RITA induzierten Zelltod festgestellt werden. Jedoch erwies sich die Herunterregulation von anti-apoptotischen Bcl-2-Proteinen nach RITA-Behandlung als wichtig für die Induktion von Apoptose. Infolgedessen führte die Kombinationsbehandlung von RITA mit ABT-737 zu einer Verstärkung des RITA-induzierten Zelltods in RITA-sensitiven Zellen. RITA-insensitive Zellen blieben dabei unbeeinflusst. Weiterhin konnte der durch RITA vermittelte Zelltod in p53-defekten Systemen auf die Aktivierung des JNK- und p38-Signaltransduktionsweges zurückgeführt werden. Insbesondere JNK1 erwies sich als entscheidend für die Induktion von Apoptose nach RITA-Behandlung. Im Vergleich der drei Substanzen zeigte sich überraschenderweise eine größere Ähnlichkeit von RITA zu Cisplatin als zu Nutlin-3. Cisplatin, als klassischer über DNA-Schädigung wirkender p53-Aktivator, führte zur Induktion von Zelltod in Zelllinien welche nahezu alle auch sensitiv gegenüber der Behandlung mit RITA waren. Potentiell könnte somit auch RITA über DNA-Schädigungen Zelltod induzieren. Intrazellulär führen jedoch beide Substanzen zu unterschiedlichen Effekten. Während Cisplatin zu einer Hochregulation von pro-apoptotischen BH3-only-Proteinen führt, induziert die Behandlung mit RITA eine Reduktion anti-apoptotischer Bcl-2-Proteine. Im Falle von Cisplatin konnte die Proteinkonzentration von NOXA und Bcl-w als Marker für die Sensitivität innerhalb der Zellauswahl identifiziert werden. RITA hingegen induzierte Zelltod nur in einer bestimmten Gruppe von Zellen, weshalb der Transport von RITA ein potentieller Marker für die RITA-Sensitivität darstellen könnte. Zusammenfassend konnte in der verwendeten Zellauswahl sowohl nach Nutlin-3-, als auch nach Cisplatin- oder RITA-Behandlung ein Einfluss von wtp53 für die Sensitivität nachgewiesen werden. Allerdings konnten durch Cisplatin und RITA auch Effekte unabhängig von p53 vermittelt werden. Interessanterweise führte die Behandlung mit RITA zu einer von der p53-Superfamilie und von der Aktivierung des JNK-Signaltransduktionsweges unabhängigen Regulation von p53-Zielgenen. Dementsprechend können im p53-defekten System p53-Zielgene sowie typische p53-Funktionen durch die Aktivierung p53-unabhängiger Signalwege vermittelt werden.Item Open Access Advances in the modelling of in-situ powder diffraction data(2013) Müller, Melanie; Dinnebier, Robert E. (Prof. Dr.)X-ray powder diffraction is a well-established technique to analyse structural and microstructural properties of materials. The possibility to record in-situ powder diffraction data allows studying changes within the structure and microstructure of a sample that occur in dependence on the applied external conditions (e.g. temperature, pressure). In the present thesis, in-situ X-ray powder diffraction was used to study structural and microstructural changes of different samples occurring at elevated temperature or upon UV illumination. Several structural phase transitions were studied using the approach of parametric Rietveld refinement. In parametric Rietveld refinement a set of powder diffraction pattern is refined simultaneously, constraining the evolution of some parameters using mathematical models, so that only the variables of the model need to be refined. In order to model and analyse the behaviour of structural parameters, Landau theory and its corresponding equations were used, owing to the fact that structural parameters (e.g. lattice strain, changes in atomic positions or occupancy) comprise an order parameter as defined in Landau theory. For description of the crystal structure of materials, several different approaches were tested, e.g. atomic coordinates, symmetry modes, rigid body rotations or rigid body symmetry modes. The dependence of preparation conditions on the properties of nanomaterials and their growth kinetics was studied using Whole Powder Pattern Modelling. This method allows modelling X-ray powder diffraction pattern using the microstructure of the sample without the use of arbitrary profile functions. The Fourier transforms of frequently observed effects as crystallite shape and size distribution or density of various defects, like dislocations and stacking faults, are utilised in order to get the resulting diffraction profile. Two different systems with industrial application, CeO2 and Cu2ZnSnS4, which were produced using a sol-gel approach, were investigated.Item Open Access Comparative analysis of the nuclear receptors CAR, PXR and PPARα in the regulation of hepatic energy homeostasis and xenobiotic metabolism(2014) Kandel, Benjamin; Zanger, Ulrich (Prof. Dr.)Nuclear receptors (NRs), most notably the constitutive androstane receptor (CAR) and the pregnane X receptor (PXR), regulate the transcription of several drug metabolizing enzymes and transporters (DMET) and thus represent important regulators of drug metabolism in the liver. Accordingly, the ligand dependent activation of these NRs by drugs and other xenobiotics contributes to the intra- and inter-individual variability of the drug detoxifying system. CAR and PXR were further shown to regulate the transcription of key enzymes involved in lipid and glucose metabolism. The NR peroxisome proliferator-activated receptor alpha (PPARa), a key regulator of fatty acid catabolism and target of lipid lowering fibrates, was recently identified as a direct regulator of cytochrome P450 3A4 (CYP3A4) and also potentially of other DMET genes. In this respect, CAR, PXR and PPARa are determinants of an overlapping number of liver functions including drug metabolism and energy homeostasis and are therefore associated with adverse drug reactions as well as liver disease like steatosis. Until now there have been no comparative studies investigating the transcriptomes of CAR, PXR and PPARa in humans. Therefore, a major focus of this study was to assess the genome-wide transcriptional changes provoked by these NRs in primary human hepatocytes (PHHs). To investigate human liver-specific gene expression and its regulation PHHs represent the most suitable available in vitro cell system. To identify the CAR-, PXR- and PPARa-specific genome-wide expression changes, hepatocyte cultures from six individual donors were treated with the prototypical ligands for CAR (CITCO), PXR (rifampicin) and PPARa (WY-14643) as well as DMSO (vehicle control). Afterwards, the mRNA expression in these samples was determined utilizing Affymetrix® microarrays. The obtained expression data were statistically evaluated to identify the genes that showed a differential expression in response to the agonist treatments and to investigate to which metabolic functions these genes contribute. The results of these experiments confirmed that CAR, PXR and PPARa regulated a highly overlapping but distinct set of genes coding for DMET. For example, according to KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analyses expression of 10 DMET genes were shown to be regulated by all three NRs, whereas other DMET genes responded exclusively to the activation of one of the NRs. In addition several DMET related genes previously not shown to be regulated by CAR [like CYP2E1, sulfotransferase 1B1 (SULT1B1), UDP-glucuronosyltransferase 2B4 (UGT2B4) and cytochrome P450 reductase (POR)], PXR [like CYP2E1, alcohol dehydrogenases (ADHs), flavin containing monooxygenase 5 (FMO5) and glutathione peroxidase 2 (GPX2)] or PPARa like UBT2B4, ADH1s and FMO5) were identified to respond to the respective agonists. For PXR and CAR, this extends the list of genes by which these NRs influence drug metabolism and potentially contribute to drug-drug interactions (DDIs). The results obtained further specify the role of PPARa as a regulator of drug metabolism in vitro by increasing expression of, e.g., CYP3A4, 2B6, 2C8 and UGT1A1, thus pointing to a potential role of PPARa in adverse drug reactions in vivo. Furthermore, several genes coding for proteins involved in energy homeostasis, were identified as differentially expressed in response to PXR activation [e.g., pyruvate dehydrogenase kinase 4 (PDK4), glycogen synthase 2 (GYS2), carnitine palmitoyltransferase 2 (CPT2)], where such a relation was not reported so far. These results further expanded the knowledge of how PXR potentially impact fatty acid catabolism, gluconeogenesis and lipid de novo synthesis and provide interesting starting points to investigate how PXR activation contributes to altered glucose and lipid levels or disease like hepatic steatosis. Besides ligand-dependent regulation of nuclear receptors, post-translational modification has also been shown to influence the activity of liver-enriched NRs and expression of their target genes. In this context, protein kinase A (PKA) had been shown to repress CYP3A4 expression via PXR in a species-dependent manner, whereas the influence of PKA on the expression of other DMET genes had not been investigated in detail so far. The second part of this work therefore investigated the impact of PKA activation on the expression and activity of important drug metabolizing enzymes in a PXR- as well as a CAR-dependent manner. In this work PKA activation in primary human hepatocytes was identified as a determinant of drug metabolism in vitro by repressing PXR- and CAR-mediated or reducing basal expression and activity of CYP1A1, CYP2B6, CYP2C8 and CYP3A4, but also expression of ATP-binding cassette B1 (ABCB1) and UGT1A1. Using reporter gene assays, these observed effects could be linked to PKA-mediated repression of PXR and CAR activity that may involve phosphorylation of these NRs. It could be further shown that expression of DMET genes was also repressed by the fasting hormone glucagon, a physiologically relevant activator of PKA signaling, which was not investigated in humans so far. Due to the promiscuous ligand-specificity of PXR, which includes numerous compounds, drug treatment often leads to PXR activation, even with so-called “natural” compounds like St. John’s wort (SJW). It would thus be highly desirable to develop strategies in drug development to assess or circumvent the activation of NRs without compromising the pharmacological effects. Therefore, the last part of this work consists of an in vitro study to investigate synthetic acylated phloroglucinols, designed as substitutes for hyperforin, regarding their potential to activate PXR. Hyperforin the major active constituent of the plant SJW used to treat depressions was shown to exert its antidepressant properties via indirect inhibition of serotonin reuptake by selectively activating the canonical transient receptor potential channel 6 (TRPC6). In addition, hyperforin is associated with clinically relevant drug-drug interactions in patients that had taken SJW concomitantly with other drugs due to potent activation of the nuclear receptor PXR by hyperforin. The phloroglucinol derivatives investigated in this thesis had previously been evaluated for their bioactivity. It had been reported that five of the nine synthetic acylated phloroglucinols activate TRPC6 with similar potency as hyperforin. In this work, all these nine synthetic phloroglucinol derivatives were investigated in comparison to hyperforin and rifampicin for their potential to activate PXR. Hyperforin and rifampicin treatment of HepG2 cells co-transfected with a human PXR expression vector and a CYP3A4 promoter reporter construct resulted in potent PXR-dependent induction, while all TRPC6-activating compounds failed to show any PXR activation or to antagonize rifampicin-mediated CYP3A4 promoter induction. Hyperforin and rifampicin treatment of primary human hepatocytes resulted in highly correlated induction of PXR target genes, whereas treatment with the phloroglucinol derivatives elicited moderate gene expression changes that only weakly correlated to those of rifampicin treatment. The herein observed lack of PXR activation by the TRPC6 activating phloroglucinols was further supported by in silico pharmacophore modeling that did not indicate potent agonist or antagonist interactions for the TRPC6 activating derivatives and docking studies that suggested interaction of only one of these compounds. These in silico studies performed by Prof. Sean Ekins are published together with the results presented in this work (Kandel et al., 2014). This approach shows that strategies avoiding PXR activation are conceivable in drug development in order to prevent DDIs and improve drug safety. Taken together, these results further increase the number of genes by which CAR, PXR, and PPARa contribute to the regulation of drug metabolism and energy homeostasis. Moreover it was demonstrated that the PKA, which is involved in the transduction of the effects of, e.g., the hormone glucagon, represents a determinant of the drug detoxifying system in humans. Furthermore, a strategy could be presented, taking the example of the hyperforin derivates, which can be used to investigate and avoid DDIs in drug development. Such information will become imperative in future personalized medicine and the ever-present polypharmacy in order to handle intra- and inter-individual variability and to minimize drug failure or drug-drug interactions.Item Open Access X-ray investigation of Nb/O interfaces(2008) Delheusy, Mélissa; Dosch, Helmut (Prof. Dr.)X-ray free electron lasers and the future International Linear Collider project are based on the performance of niobium superconducting rf cavities for efficient particle acceleration. A remarkable increase of the rf accelerating field is usually achieved by low-temperature annealing of the cavities (T<150°C, several hours). The microscopic origin of this effect has remained unclear; however, it has been argued that a redistribution of subsurface interstitial oxygen into niobium is involved. In this study, the near surface structure of oxidized niobium single crystals and its evolution upon vacuum annealing has been studied by means of non-destructive in-situ surface sensitive x-ray techniques: x-ray reflectivity (XRR), grazing incidence x-ray diffraction (GIXD), diffuse scattering (GIDXS), crystal truncation rods measurements (CTRs), and high-resolution core-level spectroscopy (HRCLS). A first insight into the interplay between the oxide formation/dissolution and the occurrence of subsurface interstitial oxygen has been given. The natural oxide on Nb(110) and Nb(100) surfaces is constituted of Nb2O5, NbO2 and NbO, from the surface to the interface. It reduces progressively upon heating from Nb2O5 to NbO2 at low temperatures, and to NbO at 300°C. The Nb(110)/NbO(111) interface presents a Nishiyma-Wassermann epitaxial orientation relationship. The depth-distribution of interstitial oxygen is established indicating that most of the oxygen is located in the direct vicinity of the oxide/niobium interface. No evidence of oxygen depletion below the oxide layer is observed for the low temperature thermal treatments and surface preparations investigated in this study.Item Open Access Eine neue Methode zur Berechnung der Energieskalen von Metallen mit Störstellen(1996) Fischer, Kurt; Fulde, Peter (Prof. Dr.)Das universelle Verhalten von Störstellen mit stark korrelierten Elektronen in einem Metall wird bewiesen. In solchen Systemen wechselwirken die lokalen, stark korrelierten Elektronen der Störstelle mit den niederenergetischen Teilchen-Loch-Anregungen des Metalls und bilden einen komplizierten Vielteilchen-Grundzustand. Alle Versuche, diesen Grundzustand, die elementaren Anregungen und die Energieskalen des universellen Verhaltens mit den sonst üblichen störungstheoretischen Methoden zu berechnen, sind gescheitert. In einigen Spezialfällen gelingt mit Hilfe des Bethe-Ansatzes oder der numerischen Renormierungsgruppe eine Bestimmung der Thermodynamik eines Modellsystems, das aber nicht direkt mit dem ursprünglichen System in Verbindung gebracht werden kann. Daher stellt sich die Frage, wie man das universelle Verhalten von solchen Systemen systematisch beschreiben kann. Die zentralen Aussagen dieser Dissertation lauten: 1. Die Energieskalen eines Systems mit Störstellen lassen sich mit Hilfe einer Diagrammtechnik exakt berechnen. 2. Der Störstellen-Anteil jeder Observablen skaliert im universellen Limes mit der gleichen Energie. 3. Allein die Diagrammstruktur liefert ein Kriterium dafür, wann eine Näherung das universelle Verhalten bewahrt. Die hier benutzte Diagrammtechnik für Störstellen in Metallen wird neu und vereinfacht hergeleitet. Die Zusammenfassung der Diagramme in Klassen von Skelettdiagrammen führt auf ein Variationsprinzip für ein erzeugendes Funktional. Der Störstellen-Anteil der jeweiligen Observablen ergibt sich am Sattelpunkt des Funktionals bezüglich der Variation nach den ionischen Propagatoren. Diese Methode stammt von Luttinger und Ward. In dieser Dissertation wird die Information benutzt, wie die Skelettdiagramme strukturiert sind. Der entscheidende Punkt ist, dass man die Parameter-Abhängigkeit der Propagatoren im erzeugenden Funktional am Sattelpunkt ignorieren kann: Sie wird nur durch die Struktur der Skelettdiagramme bestimmt. Die Skelettdiagramme können so allein aufgrund ihrer Struktur nach der inversen Bandbreite klassifiziert werden. Nur eine als relevant bezeichnete Mindestmenge der Skelettdiagramme bestimmt die exakten Energieskalen des Systems, ohne das Modell vorher gelöst zu haben. Dies ergibt die erste Aussage. Eine Observable ergibt sich stets als Antwort des Systems auf ein entsprechendes äusseres Feld. Es wurde gezeigt, dass es nicht von der Observablen abhängt, welche Skelettdiagramme relevant sind. So kann man zeigen, dass der Störstellen-Anteil jeder Observablen mit der gleichen Energieskala skaliert. Dies ergibt die zweite Aussage. Die relevanten Skelettdiagramme beschreiben damit das universelle Verhalten der Störstelle und garantieren eine systematische Näherung für die Thermodynamik und die Dynamik der Störstelle. Berücksichtigt man noch weitere Skelettdiagramme, so verhalten sich die damit berechneten Störstellen-Anteile von Observablen genau dann universell, wenn ganze Familien von Skelettdiagrammen mitgenommen werden. Diese Skelettdiagramme ändern nichts an der Energieskala. Dies ergibt die dritte Aussage. Für das Anderson-Modell Störstelle in einem Metall ergeben die relevanten Skelettdiagramme gerade die sogenannte non-crossing-approximation (NCA). Daher beschreibt die NCA das universelle Verhalten des Anderson-Modells qualitativ richtig. Skelettdiagramme höherer Ordnung ändern das Bild nicht qualitativ. Bisher war es unerklärlich, warum die NCA mit den Resultaten der numerischen Renormierungsgruppe so gut übereinstimmt. Die Erklärung findet sich im exakten Skalierungsverhalten der NCA, das hier gezeigt wurde. Um das Verfahren zu testen, haben wir für das Anderson-Modell im magnetischen Limes das crossover untersucht. Für hohe Temperaturen oder Magnetfelder verhält sich das magnetische Moment der Störstelle asymptotisch frei, während es für tiefe Temperaturen durch niederenergetische Teilchen-Loch-Anregungen des Fermi-Sees abgeschirmt wird. Dieses crossover vom perturbativen Hochtemperatur- zum nichtperturbativen Niedertemperaturbereich wird durch die Wilson-Zahl charakterisiert. Hier wird die Wilson-Zahl zum ersten Mal mit einer Diagrammtechnik (NCA) berechnet. Das Ergebnis stimmt im Rahmen der Näherung mit dem als exakt postulierten Resultat überein. Um das universelle Verhalten anderer Observablen zu demonstrieren, wurde sodann für das Anderson-Modell zum ersten Mal explizit gezeigt, dass das Einteilchen-Anregungsspektrum - und damit die Abrikosov-Suhl-Resonanz - richtig skaliert, wenn man es mit Hilfe der NCA berechnet. Die vorgestellte Methode ermöglicht es auch, für ein System mit Störstellen die Qualität von Näherungen zu beurteilen, ohne sie zuvor explizit lösen zu müssen.Item Open Access Role of inflammatory cytokine signaling in the regulation of detoxifying functions in human hepatocytes and liver(2014) Klein, Marcus; Zanger, Ulrich (Prof. Dr.)During inflammation, circulating pro-inflammatory cytokines such as TNFα, IL-1ß, and IL-6, which are produced by, e.g., Kupffer cells, macrophages, or tumor cells, play important roles in hepatocellular signalling pathways and in the regulation of cellular homeostasis. In particular, these cytokines are responsible for the acute phase response (APR) but also for a dramatic reduction of drug detoxification capacity due to impaired expression of numerous genes coding for drug metabolic enzymes and transporters (DMETs). Several pathways are known to be activated by IL-6 such as the JAK/STAT, MAPK/ERK, and PI3K/AKT pathways. Earlier work by others has shown that downregulation of CYP3A4 is independent of the JAK/STAT and MAPK/ERK pathways. However, there is evidence that MAPKs are able to phosphorylate nuclear receptors (NRs) such as RXR-α, which alters their function. Moreover, AKT, downstream of PI3K, may induce nuclear translocation of NF-κB which antagonizes RXR-α and other NRs. RXR-α, which heterodimerizes with subfamily 1 NRs (e.g., CAR and PXR), is an important regulator of detoxifying functions in liver. Inhibition of RXR-α or other NRs could therefore explain the simultaneous downregulation of large gene batteries including many DMET genes. The contributing signaling events and mechanisms remained, however, largely unexplained. Therefore, the major focus of this thesis was the investigation of the impact of the major inflammatory mediator IL-6 on the regulation of detoxifying functions in human liver. For this purpose, a large-scale investigation of DMET gene expression changes in IL-6-stimulated primary human hepatocytes (PHH) was carried out. Many important DMET genes were found to be downregulated in response to IL-6 stimulation of PHH. Most significantly suppressed were genes coding for cytochrome P450s (e.g., CYP1A2, 2C9, 2D6, and 3A4) and ATP-binding cassette (e.g., ABCB1 and ABCC2) and solute carrier (e.g., SLC10A1 and SLCO1B1) drug transporters. The average phase II metabolism gene expression appeared to be only moderately affected by IL-6, showing much stronger variability in gene expression, including genes with a trend towards upregulation (SULTs). Most notably, CYPs appeared to be highly downregulated in a coordinated fashion, demonstrating the broad suppressive potency of IL-6 towards this particular family of drug metabolizing enzymes (DMEs). Moreover, determination of metabolite formation rates in IL-6-treated PHH revealed impaired metabolic functionality of the major CYP isoenzymes CYP1A2, 2B6, 2C8, 2C9, 2C19, and 3A4. Therefore, it was shown that IL-6 signaling extensively interferes with drug detoxification capacity in human hepatocytes. Phosphoprotein analyses revealed activation of the JAK/STAT, MAPK, and PI3K cascades by IL-6. Whereas individual chemical inhibition of the two latter pathways attenuated many IL-6-mediated effects on DMET gene expression, co-inhibition almost completely abolished these effects. Inhibition of JAK/STAT signaling barely affected IL-6-mediated effects. Notably, activation of PI3K and knock-down (KD) of RXR-α demonstrated strikingly similar DMET gene expression patterns compared to IL-6 stimulation. Therefore, these data indicated a MAPK/ERK- and PI3K/AKT-dependent but JAK/STAT-independent downregulation of DMET genes in response to IL-6, possibly via interference with RXR-α. In conclusion, these data suggest that MAPKs and AKT-activated NF-κB antagonize NR signaling, causing a coordinated downregulation of DMET genes. The investigation of sensitive regulatory mechanisms is complicated by the interindividual variability of PHH. The human hepatocellular carcinoma derived HepaRG cell line has been shown to retain many functional characteristics of PHH, including the expression of functional DMETs, but the influence of inflammation has not been investigated so far. Thus, HepaRG cells were tested for their robustness and suitability in studying the inflammation-mediated impact on the drug detoxification capacity in human liver. Indeed, IL-6 stimulation of HepaRG cells led to highly induced expression of acute phase (AP) genes (e.g., CRP) and significantly repressed DMET gene expression in a coordinated fashion. The selectivity and magnitude of these effects were strikingly similar to those observed in IL-6-exposed PHH, with only few exceptions (e.g., CYP2E1 and SULTs). This was further supported by a strong positive correlation of IL-6-mediated expression changes of DMET and critical modifier genes in both cell models. Moreover, decreased protein expression and activity of major P450s could be determined in HepaRG cells, comparable to PHH. Exposure of HepaRG cells to different cytokines resulted in moderately different gene expression patterns, indicating specific responsiveness to particular pro-inflammatory cytokines. These data indicate that HepaRG cells retain the regulatory mechanisms that are responsible for the downregulation of the liver’s drug detoxification capacity during inflammation. This cell line may therefore provide a good alternative model for detailed mechanistic analyses during such conditions. The inflammation-mediated transcriptional changes that have major effects on drug detoxification in the liver have not been analyzed on a transcriptome-wide scale so far. Therefore, the last part of this work focused on the unbiased assessment of genome-wide transcriptional changes in response to inflammatory signaling in the human liver. For this purpose, microarray analysis was carried out in IL-6-stimulated PHH and compared to transcriptome data, previously acquired in samples from a liver cohort, including patients having undergone an APR (elevated CRP). Remarkably, major human-relevant CYPs, 2C8, 3A4, and 2A6 were the most strongly downregulated genes in IL-6-challenged PHH. Their transcription was at least 4-fold repressed. A total of 40 DMET genes were identified as significantly altered, of which 30 were downregulated, including almost all transcripts of major CYPs of importance in humans (e.g., 1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 3A4 and 3A5), phase II drug metabolizing enzymes (e.g., GSTAs, SULTs and UGTs), and drug transporters (ABCs and SLCs). In liver samples from patients with elevated CRP, 29 DMET genes were downregulated including important genes coding for phase I/II drug metabolizing enzymes (e.g., ADHs, ALDHs, CYPs, GSTs and UGTs) and drug transporters (e.g., ABCG2 and SLCs). In both studies, gene term enrichment analyses indicated a very strong influence on xenobiotic metabolic and related processes, containing mostly downregulated DMET genes. Moreover, pathway enrichment (KEGG) analyses revealed that drug and xenobiotic metabolic signaling pathways were the most strongly impacted reaction networks, clearly demonstrating that the drug detoxification system in the liver is largely affected during inflammation. Gene annotation analysis also identified enriched processes related to diverse lipid metabolic processes such as fatty-acid and steroid metabolism. Moreover, enriched biological processes and regulatory pathways related to amino acid metabolism were found, particularly in the retrospective study. The data indicated a conservation and allocation of specific amino acids, possibly in favor of acute phase protein (APP) synthesis. Taken together, these findings highlight the scale on which the human liver transcriptome is affected during inflammation. Extensive reorganization related to xenobiotic, lipid, and amino acid metabolism takes place. It appears that the liver devotes its transcriptional machinery to the immune response while other major liver functions are shut down. This may help to pave the way towards a better understanding of how the liver organizes its many responsibilities in different conditions.