@phdthesis{Bartl2012, author = {Bartl, Jasmin}, title = {Impairment of insulin signaling pathway in Alzheimer's disease}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-74197}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2012}, abstract = {The neurodegenerative disorder Alzheimer's disease (AD) is the cause of approximately 60\% of the world's 35 million patients suffering from dementia. Current research focuses here are on association with other diseases such as diabetes type 2 (T2DM), possible genetic markers, specific signal transduction pathways within the brain and potential protein modification, because the pathogenesis and etiology of AD are still not fully understood. Specifically association of T2DM with AD came to the focus with the so-called "Rotterdam study" in 1999, indicating that T2DM doubles the risk of developing AD. In the meantime, it is known that the prevalence rate in patients with T2DM is 30\%. Drugs commonly used in the treatment of T2DM such as peroxisome proliferator-activated receptors gamma (PPARγ) agonists show improvement of the cognitive abilities in patients with early stage of dementia, with potential therapeutically relevance. Therefore it is important not only to investigate a link between these diseases, but also to investigate the insulin signaling pathway in the brain of AD patients. In order to investigate this complex issue in more details and demonstrate additional links between T2DM and AD, the present study used several basic biological methods to clarify the question: "Is impaired insulin signaling pathway within the brain crucial for the development of AD?" from several points of view. The methods used in this work have been i) an analysis of single nucleotide (SNP) polymorphism of the insulin-degrading enzyme gene (IDE) in relation to risk of AD and / or of T2DM, ii) post-mortem histochemical studies of brain tissue of patients with only AD, with AD combined with T2DM and with only T2DM compared with an age-matched control group, and iii.) investigations of neurochemical pathways and gene/protein expression changes of a human cell culture as a consequences of amyloid β (Aβ) treatment. After analysis of the IDE SNP polymorphism in the selected VITA (Vienna Trans Danube Aging) cohort disease-specific effects were discovered. The upstream polymorphism (IDE2) was found to influence AD risk in a protective manner, while the downstream polymorphism (IDE7) modified the T2DM risk. Based on the SNP results, the presented study delineate the model that IDE promoter and 3‟ untranslated region/downstream variation can have different effects on IDE expression, maybe a relevant endophenotype with disorder-specific effects on AD and T2DM susceptibility. Furthermore, the human post-mortem studies could show that both AD as well as T2DM patients had a significantly lower density of the insulin receptor (IR) in the hippocampus, whereas a significantly increased density of inactive phosphorylated PPARγ has been found and this persisted even in patients with both diseases. Summarizing the histological study, it was possible to reveal common histological features of AD and T2DM, but no direct connection between the two diseases. Although AD is nowadays not only characterized by amyloid-containing plaque deposits and by the hyperphosphorylation of tau protein, the excessive Aβ42 presence in the brains of AD patients is still playing a key role. Up to date it is still not entirely clear which physical form of Aβ42 is responsible for the development of AD. The present work investigated, what impact has the state of aggregation of Aβ42 on genes and proteins of the insulin signaling pathway and the amyloid cascade. It could be shown that the oligomeric variant enhanced specifically the gene and protein expression of glycogen synthase kinase (GSK) 3β and also the enzyme activity was significantly increased, but has in turn strongly inhibited the IR gene and protein expression. Additionally, the effect of Aβ42 on monoamine oxidase B (MAO-B) was examined. An effect of both aggregated forms of Aβ42 had on enzyme activity was discovered. However, the fibrillar variants led to significantly increased activity of MAO-B while the oligomeric variants inhibited the enzyme activity. Several previous studies have demonstrated the involvement of increased MAO-B activity in AD, but the present work provides for the first time a direct link between the states of aggregation of Aβ42 to enzyme activity. Finally the results of the presented thesis can be summarized to following conclusion: Although AD and T2DM sharing some degrees of common features, still there is a lack of direct association, and therefore the diseases must be considered more independent rather than linked. But the impaired cerebral insulin signaling pathway seems to be another manifested hallmark of AD.}, subject = {Alzheimer-Krankheit}, language = {en} } @phdthesis{Bernhardt2012, author = {Bernhardt, Marcel}, title = {Diagnostik der invasiven Aspergillose bei immunsupprimierten Patienten}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-97386}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2012}, abstract = {Die invasive Aspergillose ist eine schwerwiegende opportunistische systemische Infektion bei immunsupprimierten Patienten. In dieser Arbeit wurden verschiedene PCR-Verfahren und ein ELISA-Verfahren zum Antigennachweis in Hinblick auf Sensitivit{\"a}t, Sensibilit{\"a}t und positiver bzw. negativer Vorhersagewahrscheinlichkeit verglichen. Die konventionelle 18S-PCR ist ein panfungales Assay und wegen zahlreicher falsch-positiver Ergebnisse nicht geeignet zur Fr{\"u}hdiagnose. Die ITS-PCR hat sich aufgrund guter Spezifit{\"a}t als vielversprechend erwiesen, muss aber noch mit gr{\"o}ßeren Fallzahlen evaluiert werden. Der Antigennachweis (Platelia, Fa. Bio-Rad) weist eine hohe Sensitivit{\"a}t und negativen pr{\"a}diktiven Wert auf. Als vielversprechend d{\"u}rfte zuk{\"u}nftig eine Kombination aus PCR und Antigennachweisverfahren gelten.}, subject = {Aspergillose}, language = {de} } @phdthesis{Heffels2012, author = {Heffels, Karl-Heinz}, title = {Functional nanofibres for regenerative medicine}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-75684}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2012}, abstract = {This thesis concerned the design and examination of a scaffold for tissue engineering applications. The template for the presented scaffold came from nature itself: the intercellular space in tissues that provides structure and support to the cells of the respective tissue, known as extracellular matrix (ECM). Fibres are a predominant characteristic feature of ECM, providing adhesion sites for cell-matrix interactions. In this dissertation a fibrous mesh was generated using the electrospinning technique to mimic the fibrous structure of the ECM. Two base polymers were explored: a biodegradable polyester, poly(D,L-lactide-co-glycolide); and a functional PEG-based star polymer, NCO-sP(EO-stat-PO). This topic was described in three major parts: the first part was materials based, concerning the chemical design and characterisation of the polymer scaffolds; the focus was then shifted to the cellular response to this fibrous scaffold; and finally the in vivo performance of the material was preliminarily assessed. The first steps towards an electrospun mesh started with adjusting the spinning parameters for the generation of homogeneous fibres. As reported in Chapter 3 a suitable setup configuration was on the one hand comprised of a spinning solution that consisted of 28.5 w/v\% PLGA RG 504 and 6 w/v\% NCO-sP(EO-stat-PO) in 450 µL acetone, 50 µL DMSO and 10 µL of an aqueous trifluoroacetic acid solution. On the other hand an ideal spinning behaviour was achieved at process parameters such as a flow rate of 0.5 mL/h, spinneret to collector distance of 12-16 cm and a voltage of 13 kV. The NCO-sP(EO-stat-PO) containing fibres proved to be highly hydrophilic as the functional additive was present on the fibre surface. Furthermore, the fibres featured a bulk degradation pattern as a consequence of the proportion of PLGA. Besides the morphologic similarity to ECM fibres, the functionality of the electrospun fibres is also decisive for a successful ECM mimicry. In Chapter 4, the passive as well as active functionality of the fibres was investigated. The fibres were required to be protein repellent to prevent an unspecific cell adhesion. This was proven as even 6.5 \% sP(EO-stat-PO) in the PLGA fibres reduced any unspecific protein adsorption of bovine serum albumin and foetal calf serum to less than 1 \%. However, avidin based proteins attached to the fibres. This adhesion process was avoided by an additional fibre surface treatment with glycidol. The active functionalisation of NCO-sP(EO-stat-PO)/PLGA fibres was investigated with two fluorescent dyes and biocytin. A threefold, chemically orthogonal, fibre modification was achieved with these dyes. The chapters about the chemical and mechanical properties laid the basis for the in vitro chapters where a specific fibre functionalisation with peptides was conducted to analyse the cell adhesion and biochemical expressions. Beginning with fibroblasts in Chapter 5 the focus was on the specific cell adhesion on the electrospun fibres. While NCO-sP(EO-stat-PO)/PLGA fibres without peptides did not allow any adhesion of fibroblasts, a fibre modification with GRGDS (an adhesion mediating peptide sequence) induced the adhesion and spreading of human dermal fibroblasts on the fibrous scaffolds. The control sequence GRGES that has no adhesion mediating qualities did not lead to any cell adhesion as observed on fibres without modifications. While the experiments of Chapter 5 were a proof-of-concept, in Chapter 6 a possible application in cartilage tissue engineering was examined. Therefore, primary human chondrocytes were seeded on fibrous scaffolds with various peptide sequences. Though the chondrocytes exhibited high viability on all scaffolds, an active interaction of cells and fibres was only found for the decorin derived sequence CGKLER. Live-cell-imaging revealed both cell attachment and migration within CGKLER-modified meshes. As chondrocytes undergo a de-differentiation towards a fibroblast-like phenotype, the chondrogenic re-differentiation on these scaffolds was investigated in a long term cell culture experiment of 28 days. Therefore, the glycosaminoglycan production was analysed as well as the mRNA expression of genes coding for collagen I and II, aggrecan and proteoglycan 4. In general only low amounts of the chondrogenic markers were measured, suggesting no chondrogenic differentiation. For conclusive evidence follow-up experiments are required that support or reject the findings. The success of an implant for tissue engineering relies not only on the response of the targeted cell type but also on the immune reaction caused by leukocytes. Hence, Chapter 7 dealt with primary human macrophages and their behaviour and phenotype on two-dimensional (2D) surfaces compared to three-dimensional (3D) fibrous substrates. It was found that the general non-adhesiveness of NCO-sP(EO-stat-PO) surfaces and fibres does not apply to macrophages. The cells aligned along the fibres on surfaces or resided in the pores of the meshes. On flat surfaces without 3D structure the macrophages showed a retarded adhesion kinetic accompanied with a high migratory activity indicating their search for a topographical feature to adhere to. Moreover, a detailed investigation of cell surface markers and chemokine signalling revealed that macrophages on 2D surfaces exhibited surface markers indicating a healing phenotype while the chemokine release suggested a pro-inflammatory phenotype. Interestingly, the opposite situation was found on 3D fibrous substrates with pro-inflammatory surface markers and pro-angiogenic cytokine release. As the immune response largely depends on cellular communication, it was concluded that the NCO-sP(EO-stat-PO)/PLGA fibres induce an adequate immune response with promising prospects to be used in a scaffold for tissue engineering. The final chapter of this thesis reports on a first in vivo study conducted with the presented electrospun fibres. Here, the fibres were combined with a polypropylene mesh for the treatment of diaphragmatic hernias in a rabbit model. Two scaffold series were described that differed in the overall surface morphology: while the fibres of Series A were incorporated into a thick gel of NCO-sP(EO-stat-PO), the scaffolds of Series B featured only a thin hydrogel layer so that the overall fibrous structure could be retained. After four months in vivo the treated defects of the diaphragm were significantly smaller and filled mainly with scar tissue. Thick granulomas occurred on scaffolds of Series A while the implants of Series B did not induce any granuloma formation. As a consequence of the generally positive outcome of this study, the constructs were enhanced with a drug release system in a follow-up project. The incorporated drug was the MMP-inhibitor Ilomastat which is intended to reduce the formation of scar tissue. In conclusion, the simple and straight forward fabrication, the threefold functionalisation possibility and general versatile applicability makes the meshes of NCO-sP(EO-stat-PO)/PLGA fibres a promising candidate to be applied in tissue engineering scaffolds in the future.}, subject = {Nanofaser}, language = {en} } @phdthesis{Meusel2012, author = {Meusel, Marcus}, title = {Der positive Effekt von Indometacin im isch{\"a}misch bedingten akuten Nierenversagen}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-74219}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2012}, abstract = {Aus Voruntersuchungen geht hervor, dass die Expression der organischen Anionentransporter OAT-1 und OAT-3 durch Prostaglandin E2 herabgesetzt wird und dass beide Transporter im isch{\"a}misch bedingten akuten Nierenversagen herunter reguliert werden. Da zudem Prostaglandin E2 im iANV vermehrt vorliegt und von Cyclooxygenasen gebildet wird, wurde in dieser Arbeit der Effekt von Indometacin als nicht-selektiver COX-Inhibitor auf die Expression und Funktion von OAT-1 und OAT-3, sowie auf die gesamte Nierenfunktion untersucht. Das isch{\"a}misch bedingte akute Nierenversagen wurde bei Ratten durch bilaterales Abklemmen der Aa. renales {\"u}ber 45 Minuten induziert. Indometacin (1mg/kg) wurde hierbei intraperitoneal gegen Ende der Isch{\"a}miephase appliziert. Die Gruppeneinteilung erfolgte in Gruppen mit Gef{\"a}ßabklemmung jeweils mit bzw. ohne Indometacingabe und in Gruppen mit Scheinoperationen. Die Expression von OAT-1 und OAT-3 wurde mithilfe von rt-PCR und Western Blot Verfahren bestimmt, deren Funktion anhand der PAH Nettosekretion ermittelt und die Nierenfunktion mithilfe von PAH- und Inulin Clearance analysiert. Alle Parameter wurde 24h nach renaler Isch{\"a}mie betrachtet. In Isch{\"a}mie-Tieren konnte Indometacin die Expression von OAT-1 und OAT-3 und die PAH Nettosekretion wiederherstellen. Zus{\"a}tzlich vermochte Indometacin auch die Nierenfunktion signifikant gegen{\"u}ber den Isch{\"a}mie-Tieren ohne Therapie zu verbessern. So l{\"a}sst sich zusammenfassend sagen, dass niedrig dosiertes Indometacin eine Herunterregulation von OAT-1 und OAT-3 nach isch{\"a}misch bedingtem akuten Nierenversagen verhindert und einen relevanten protektiven Effekt auf die Nierenfunktion zeigt.}, subject = {Indometacin}, language = {de} } @phdthesis{Schuster2012, author = {Schuster, Thomas Friedrich}, title = {Akute exogene Intoxikationen im Patientengut der Intensivstation der Medizinischen Universit{\"a}tsklinik W{\"u}rzburg : 1996 - 2000}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-90523}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2012}, abstract = {Akute exogene Intoxikationen, Arten, H{\"a}ufigkeiten, Therapien und Komplikationen im 5 Jahresr{\"u}ckblick 1996 bis 2000 und im Vergleich zu 1986 bis 1990 im Patientengut der Intensivstation der Medizinischen Universit{\"a}tsklinik W{\"u}rzburg.}, subject = {Vergiftung}, language = {de} }