Hepatocellular carcinoma (HCC) is one of the most lethal cancers in Hong Kong and Southeast Asian countries. Cancer progression is often symptomless, making the early diagnosis difficult, thus leading to a high mortality rate. Treatments against HCC were often found to be less effective than other cancers. Systemic chemotherapy, which is widely used in cancer treatments, has a low response rate in HCC. New treatment regimes, such as targeted therapy, have shown partial responses in clinical trials and therefore continuous effort in searching new drug targets is warranted. Granulin-epithelin Precursor (GEP) is a pluripotent growth factor, and has been shown to be overexpressed in HCC and various cancers. Our group has demonstrated that GEP promotes tumor growth, and regulates chemoresistance in HCC. It shares a highly similar expression pattern with one of the members of ATP-binding cassette (ABC) transporter family, ABCF1. Blocking GEP, both in vitro and in vivo, showed inhibition on HCC growth. These suggest that GEP is a potential target for HCC treatment. However, there is still little information on how GEP and ABCF1 is overexpressed in HCC. This project aims to investigate the mechanisms involved. GEP and ABCF1 genes are located on chromosomes 17q and 6p, respectively, which both are frequently amplified in HCC. We used quantitative microsatellite analysis (QuMA) to detect GEP and ABCF1 amplification in HCC samples. Both GEP and ABCF1 showed about 20% of HCC cases having amplification, and their copy numbers correlated to the mRNA expression levels. The copy numbers of GEP were also found to correlate to those of ABCF1 significantly. Clinico-pathological analysis showed that GEP copy numbers correlated with gender, serum AFP levels and HBV status, while ABCF1 did not associate with any of the clinico-pathological features. Fluorescence in situ hybridization (FISH) was performed to validate the results on DNA copy number by QuMA. The cases with highest DNA copy number on GEP and ABCF1, were examined. The average difference between FISH and QuMA results ranged ± 0.3 copies, indicating QuMA and FISH results were corroborated on DNA copy number. Furthermore, the FISH results indicated that there are different degrees of aneuploidy involved in chromosome 6p and 17q in 5 out of 6 cases investigated. These suggest that the copy number variations in GEP and ABCF1 were partly caused by the abnormal number of chromosomes. In summary, we observed that GEP and ABCF1 gene copy numbers were increased in subsets of HCC cases, and the increase correlated to their respective transcript expression levels. Furthermore, these copy number variations partly could be explained by aneuploidy as demonstrated by FISH analysis. The current study may help to understand the complex genomic aberrations in HCC and allow better treatment designs in the future. / published_or_final_version / Surgery / Master / Master of Philosophy

Σκοπός της παρούσας διπλωματικής είναι η μελέτη των αναπτυσσόμενων υπερτάσεων στις γραμμές μεταφοράς υψηλής τάσης (132kV) του δικτύου της Αρχής Ηλεκτρισμού Κύπρου. Γίνεται μια παρουσίαση του κάθε στοιχείου του δικτύου ξεχωριστά, με ποιο τρόπο μπορεί να μοντελοποιηθεί και πως συμπεριφέρεται σε κεραυνικό πλήγμα εύρους 100kA με τρία διαφορετικά σήματα. Τέλος παρουσιάζονται τα αποτελέσματα της προσομοίωσης στο λογισμικό ATP-EMTP και εξάγονται παρατηρήσεις και συμπεράσματα. Αρχικά στο πρώτο κεφάλαιο γίνεται μια εισαγωγική παρουσίαση της διπλωματικής εργασίας, μια ιστορική αναδρομή για τα συστήματα ηλεκτρικής ενέργειας και την επανάσταση που έφερε η ανάπτυξη τους. Στη συνέχεια παρουσιάζεται η πορεία του ηλεκτρισμού στη Κύπρο κάτω από το φορέα διαχείρισης του, την Αρχή Ηλεκτρισμού Κύπρου. Στο δεύτερο κεφάλαιο παρατίθενται λεπτομερώς τα στοιχεία που συνθέτουν σήμερα το δίκτυο της Α.Η.Κ από την παραγωγή, τη μεταφορά και τη διανομή της ηλεκτρικής ενέργειας στο νησί. Ακόμη γίνεται μία σύντομη αναφορά στο κλίμα της Κύπρου και πως αυτό ευνοεί τη δημιουργία κεραυνών. Στο τρίτο κεφάλαιο παρουσιάζεται και περιγράφεται αναλυτικά η αιτία της δημιουργίας των εξωτερικών υπερτάσεων, δηλαδή ο κεραυνός και πως λειτουργεί αυτός ως φυσικό φαινόμενο. Επίσης περιγράφονται οι διάφοροι μηχανισμοί δημιουργίας υπερτάσεων από τους κεραυνούς. Στα επόμενα τρία κεφάλαια παρουσιάζονται ενδελεχώς τα τρία βασικά στοιχεία του δικτύου το οποίο τυγχάνει προσομοίωσης, δηλαδή οι γραμμές μεταφοράς, οι πυλώνες και οι αλυσοειδείς μονωτήρες ανάρτησης. Συγκεκριμένα, για τις μεταφοράς γίνεται αναφορά στα γενικά τους στοιχεία, στα χαρακτηριστικά τους αλλά και στις καταπονήσεις που υφίστανται. Στους πυλώνες παρουσιάζεται η καταπόνηση τους από κεραυνούς με απότομο μέτωπο, το πως εκτιμάται η αντίσταση γείωσης τους και τι ρόλο παίζει η ειδική αντίσταση εδάφους. Όσο αφορά τους μονωτήρες ανάρτησης προβάλλονται γενικά κατασκευαστικά τους στοιχεία, οι συνθήκες στις οποίες λαμβάνει χώρα η υπερπήδηση(διάσπαση) τους και οι μέθοδοι υπολογισμού του χρόνου επιφανειακής διάσπασης των. Στο έβδομο κεφάλαιο παρουσιάζεται το λογισμικό ATP- EMTP μέσω του οποίου γίνεται η μοντελοποίηση των στοιχείων του δικτύου και η προσομοίωση τους. Γίνεται αναφορά στη δομή του προγράμματος, τα ιστορικά του στοιχεία, τις διάφορες βιβλιοθήκες μοντέλων που εμπεριέχονται σε αυτό, τα διάφορα προγράμματα υποστήριξης του και περισσότερο το ATP – Draw όπου παρέχεται η δυνατότητα για σχεδίαση των στοιχείων και εισαγωγή των παραμέτρων τους. Στο όγδοο κεφάλαιο παρουσιάζεται η μοντελοποίηση των προαναφερθέντων στοιχείων της διάταξης και ενσωμάτωση τους στο ATP – EMTP. Τέλος, στο ένατο και τελευταίο κεφάλαιο παρατίθενται τα αποτελέσματα της προσομοίωσης, η εξαγωγή των αποτελεσμάτων, οι παρατηρήσεις και τα συμπεράσματα. / The aim of the work described in this thesis is the study of the induced overvoltages on high voltage transmission lines (132kV) of the network of the Electricity Authority of Cyprus. Every network element is separately described, ways for the modeling of each element are explored and the behavior of each element during a 100kA lightning strike using three different signals is observed. Lastly, the results of the simulation are presented by using ATP/EMTP software, observations are made and findings are determined. The first chapter introduces the thesis and consists of a historical overview of electrical power systems and an overview of the revolution brought through their development. Thereafter the development of electricity in Cyprus as part of the electricity supplier “Electricity Authority of Cyprus” is analyzed. A detailed review of the system used by the “Electricity Authority of Cyprus” (Generation, Transmission, Distribution, Supply) constitutes the second chapter. Further, a brief reference to the climate of Cyprus is made and on how it favors the formation of lightning. The third chapter presents and describes in detail what might cause extraneous voltages, i.e. lightning and how it works as a natural phenomenon. Plus the different generation mechanisms of lightning overvoltages are described. In the next three chapters the main three elements of the network being simulated are presented in detail, i.e. transmission lines, pylons and catenary suspension insulators. Namely, with regard to transmission lines general information is given, their characteristics are described along with the stresses they face. As long as the pylons are concerned, the stresses they face by steep-front lightning are described, information on how to assess their grounding system resistance is given and it is also discussed whether Soil Resistivity plays a dominant role. Regarding suspension insulators their parts are described, the circumstances under which their disruptive discharge takes place and the methods of calculating surface discharges. In the seventh chapter the ATP/EMTP software used for network modeling and simulation is presented. Reference is made to the program structure, the historical data, the various model- libraries included in it, the various supporting programs and more specifically ATPDraw which creates/draws the elements and which does the parameter fitting. The eighth chapter contains the analysis of the modeling of the aforementioned parts of the assembly and their integration in the ATP/EMTP. Lastly, in the ninth and final chapter the results of the simulation are presented, observations are made and findings are determined.

Lignin is a key component of plant secondary cell walls, providing strength to the plant and allowing water transport. Lignin is a polymer of monolignols that are synthesized in the cell and transported into the cellulose rich cell wall. The primary goal of this thesis is to understand the mechanism(s) of monolignol deposition during xylogenesis. The currently accepted theory is that monolignols are exported by Golgi-mediated vesicle delivery to the secondary cell wall. When this theory was re-examined using cryofixed developing pine, quantitative autoradiography showed that monolignols did not accumulate in Golgi but were rapidly translocated from cytosol to cell wall. This suggests alternative mechanisms, such as membrane transporters, work in monolignol export. ATP binding cassette (ABC) transporters were chosen because they transport other secondary metabolites and some ABC transporter encoding genes are highly expressed in lignifying cells. Four candidate ABC transporters were selected in Arabidopsis (ABCB11, ABCB14, ABCB15 from the ABCB/MDR subfamily and ABCG33 from the ABCG/PDR subfamily) and shown to have overlapping, high vasculature expression patterns. Mutants with T-DNA insertions in single ABC transporter genes had no change in lignification of inflorescence stems. However, a reduced polar auxin transport phenotype was detected in mutants of ABCB11, ABCB14 and ABCB15. An additional approach was the use of inhibitors of ABC transporters. A new assay, which was developed to quantify lignification in primary xylem of Arabidopsis roots, demonstrated that ABC inhibitors did not change lignin deposition. Monolignols are exported and polymerized in the polysaccharide matrix of the cell wall, which includes hemicelluloses that may organize monolignols during polymerization. Since diverse lignified cell types are enriched in either G- or S-lignin, I hypothesized that this pattern could reflect different hemicellulose distributions, which was examined using antibody labeling of xylans or mannans in hybrid poplar xylem. While xylans were generally distributed in all secondary cell walls, mannans were enriched in fibers but not in the ray and vessel walls. In summary, during secondary cell wall deposition, monolignols are exported by unknown transporter(s) rather than Golgi vesicles. In developing poplar wood, the monolignols are deposited into diverse hemicellulose domains in different cell types.

Untersucht wurde die mögliche Rolle von Adenosin-5´-Triphosphat (ATP) als extrazelluläres Signalmolekül im Neostriatum der Ratte. Zum Einsatz kam die patch-clamp Methode, adaptiert für Ableitungen aus akuten Hirnschnitten. Bereits bekannt war, dass ATP exzitatorische postsynaptische Ströme an GABAergen, striatalen Projektionsneuronen („medium spiny“ Neurone) hemmen konnte. Nun sollten die verantwortlichen Mechanismen hinter diesem Effekt aufgeklärt werden. Es zeigte sich, dass exogen zugeführtes ATP zunächst zu Adenosin metabolisiert werden musste, um seine Wirkung ausüben zu können. Ein Teil dieses Effektes war, vermittelt über präsynaptische Adenosin A1-Rezeptoren, einer Hemmung der striatalen Glutamat-Freisetzung geschuldet. Neu war, dass auch die „medium spiny“ Neurone selbst funktionelle A1-Rezeptoren exprimierten. Aktiviert durch lokal gebildetes Adenosin vermittelten diese eine Hemmung der Leitfähigkeit von Glutamat-Rezeptoren des N-Methyl-D-Aspartat (NMDA) Subtyps. Unter physiologischen Bedingungen mag dieser Mechanismus der Begrenzung der Informationsweiterleitung über die GABAergen Projektionsneurone dienen. Striataler Glutamat-Exzess mit Überaktivierung von NMDA-Rezeptoren ist ein Charakteristikum der Huntington´schen Erkrankung. Eine Adressierung der A1 Rezeptoren als therapeutische Option im Rahmen dieser Basalganglienerkrankung scheint daher prinzipiell möglich.

Protein synthesis is a highly conserved process across all domains of life, both structurally and functionally. This cyclic process is catalyzed by numerous soluble protein factors that interact with the ribosome to facilitate efficient protein synthesis. Many canonical translation factors bind and hydrolyze GTP to induce conformational changes that facilitate translation. For example, GTP hydrolysis by EF-Tu is required for the release of aminoacyl-tRNA into the ribosomal A site; GTP hydrolysis by EF-G facilitates the movement of tRNA and mRNA from the A site to the P site of the ribosome. However, protein synthesis seems to also have a requirement for ATP; the essential yeast protein eEF-3 facilitates release of deacyl-tRNA from the ribosomal E site. In Escherichia coli, the protein product of the open reading frame yhih has been suggested to have a similar function. However, the role of this unique prokaryotic protein is not understood. Preliminary characterization of this protein suggests a nucleotide-dependent conformational change occurs in a truncated form of the protein, ΔP541 Yhih. Interestingly, this phenomenon is not observed in ΔL432 Yhih. Both ΔP541 Yhih, and to a lesser extent ΔL432 Yhih, exhibit a ribosome-dependent ATPase activity, suggesting the primary region for binding with the ribosome lies between Leu432 and Pro541. / x, 101 leaves : ill. ; 29 cm.

No description available.

Glial-derived lipoparticles facilitate the transport of cholesterol and lipids between cells within the CNS and have been shown to support neuronal growth and synaptogenesis. Partial deafferentation of the hippocampus by unilateral entorhinal cortex lesioning (uECL) induces well-described cytoarchitectural reorganisation and reactive sprouting in the dentate gyrus (DG). Previous studies have demonstrated a dynamic regulation of cholesterol homeostasis in the hippocampus following deafferentation, and suggest that mechanisms facilitating cholesterol transport are important during reinnervation. Furthermore, there is growing evidence that statins, a family of cholesterol-lowering drugs which inhibit the rate-limiting enzyme of cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCoA-R), may confer neuroprotection following trauma. / The ATP binding cassette transporters (ABC) A1 and G1 assist the generation of lipoparticles by mediating cholesterol and phospholipid efflux to extracellular apolipoprotein E (APOE), the brain's primary lipoprotein. To examine a role for these transporters in the regulation of cholesterol efflux during synaptic remodelling, and the effects of low-dose pravastatin (a potent HMGCoA-R inhibitor) on such intercellular transport mechanisms, we measured the expression of ABCA1, ABCG1, APOE, apoE(LDL)R and HMGCoA-R in the hippocampus of saline and pravastatin treated mice over time following uECL. It is shown here that ABCA1 and not ABCG1 is up-regulated at the level of mRNA and protein expression, along with APOE, in the hippocampus during active regeneration (14DPL) as determined by histochemical analysis of acetylcholinesterase staining density in the DG. While pravastatin treatment was observed to differentially influence the expression of ABCA1 mRNA and protein over time, no effects on APOE or ABCG1 mRNA expression were observed following uECL. Additionally, HMGCoA-R mRNA expression was significantly down-regulated at 21 DPL in the deafferented hippocampus in pravastatin-treated animals. While the low-dose pravastatin treatment applied here was sufficient to inhibit HMGCoA-R activity in the liver, enzymatic activity was unaffected in the cortex. / These findings suggest that ABCA1 and not ABCG1 may be important in the APOE-mediated cholesterol recycling observed during the active phase of neural reinnervation in response to uECL. In addition, the results presented here suggest that the administration of clinically-relevant statin therapy may be sufficient to influence the regulation of cerebral cholesterol homeostasis following trauma in the adult mouse brain.

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We have employed a differential detector combined with dielectrophoretic (DEP) translation in a microfluidic channel to monitor dielectric response of single cells and particularly to track phenomenon related to apoptosis. Two different cell lines were studied; Chinese hamster ovary cells (CHO) and MRC5 cells. Dielectric response was quantified by a factor called Force Index. Force Index was studied statistically to identify apoptotic subpopulations. Another direction of this work was to monitor changes in the cytoplasm conductivity following inhibition of mitochondrial ATP production by Oligomycin. To make the DEP response mostly sensitive to the cytoplasm conductivity, medium conductivity and DEP frequency were adjusted such that Clausius Mossotti factor and hence DEP response become less sensitive to cell radius. Chinese hamster ovary cells were used in this work and the impact of different concentrations of Oligomycin has been studied. We show that following exposure to Oligomycin at 8 μg/ml, cytoplasm conductivity drops. The majority of the changes takes place within one hour of exposure to the drug. Furthermore, double shell models has been used to estimate cytoplasm conductivity in a medium with conductivity of 0.42 S/m and the drop in the cytoplasm conductivity following treatment with Oligomycin was estimated to be ≈ 0.16 S/m. The magnitude of the decrease in the cytoplasm conductivity is evidence that Glycolysis is active as an energy production pathway within the cell. This approach can be used to quantify Glycolysis versus mitochondria ATP production which has an application in Warburg effect in cancer cells and monitoring bioprocesses.

Glucose metabolism in pancreatic α- and β-cells is believed to regulate secretion of glucagon and insulin, respectively. In β-cells, ATP links glucose metabolism to electrical activity and insulin secretion. In α-cells, ATP has been attributed various roles in glucose-regulated glucagon release, but the underlying mechanisms are poorly understood. Despite its importance in insulin and glucagon secretion little is known about ATP kinetics in α- and β-cells. In this thesis, the novel fluorescent ATP biosensor Perceval was used to monitor physiologically relevant ATP concentrations with little influence of ADP. Glucose stimulation of β-cells within mouse and human pancreatic islets induced pronounced rise of ATP with superimposed oscillations. Simultaneous measurements of the sub-plasma membrane ATP and Ca2+ concentrations revealed glucose-induced oscillations in opposite phase. ATP increased further and the oscillations ceased when voltage-dependent Ca2+ influx was prevented. In contrast, ATP promptly decreased in response to K+-depolarization-induced elevation of Ca2+. Also mobilization of Ca2+ from intracellular stores lowered ATP, but the negative effect was not due to increased ATP consumption by the sarco/endoplasmic reticulum Ca2+-ATPase. Store-operated Ca2+ entry alone had little effect but markedly elevated ATP when combined with muscarinic receptor activation. When comparing ATP and Ca2+ responses in α- and β-cells within the same islet, glucose-induced ATP generation was much less pronounced and the dose-response relationship left-shifted in the α-cells. At basal glucose, individual α-cells showed Ca2+ and concomitant ATP oscillations in opposite-phase with variable frequency. These oscillations largely cancelled out when averaging data from several α-cells. At high glucose, the Ca2+ and ATP oscillations in α-cells tended to synchronize with the corresponding β-cell oscillations. Since β-cell Ca2+ oscillations drive pulsatile insulin secretion, which is antiparallel to pulsatile glucagon secretion, there seems to be an inverse relationship between changes in α-cell Ca2+ and glucagon release. This paradox is attributed to paracrine inhibition overriding Ca2+ stimulation, since somatostatin receptor blockade potently stimulated glucagon release with little effect on α-cell Ca2+ signalling. The data indicate that complex ATP-Ca2+ interactions in α- and β-cells underlie cell-intrinsic regulation of glucagon and insulin secretion and that paracrine inhibition of glucagon release becomes important in hyperglycaemia.