Osteoarthritis (OA) is a chronic and highly prevalent degenerative disease of the synovial joint leading to cartilage destruction and bone remodelling. The current management of end-stage OA is joint replacement, however, this procedure is not suitable for a subset of patients hence there is a growing need for alternative treatments and technologies to address this limitation. One such approach to this problem is the application of cell-based therapies that regenerate areas of damaged cartilage. Recently discovered articular cartilage progenitor cells (CPC) have been hallmarked as a potential cell source for repair and/or regeneration of damaged articular cartilage. Initial focus was on the characterisation of human CPC isolated from healthy donors and compared with OA derived CPC and patient matched OA Bone Marrow Mesenchymal Stem Cells (BM-MSCs). Comparison of all cell types showed similar morphology and proliferative capacity. In addition, all cell types isolated showed positive expression of the putative mesenchymal stem cell makers; CD-90, CD-105 and CD-166 while lacking expression of CD-34. All cell types investigated showed successful osteogenic, chondrogenic and adipogenic differentiation, hence providing evidence of the mesenchymal stem cell properties of isolated CPC. A gene profiler array was used to identify the expression of Wnt pathway genes from RNA isolated from CPC cell lines originating from healthy and OA cartilage. Interestingly, the expression of Dkk-1 was observed to have the highest up-regulation in OA-derived CPC. The role of Dkk-1 was further studied in a number of CPC and chondrocyte cell lines from healthy and OA cartilage. It was found that normal CPC cell lines showed homogenously low expression and secretion of Dkk-1, however, OA-derived CPC cell lines exhibited a heterogeneous expression and secretion of Dkk-1. In a pellet culture model of chondrogenic differentiation, CPC cell lines secreting high levels of Dkk-1 failed to undergo chondrogenic differentiation, measured by diminished expression of chondrogenic differentiation markers, Type II collagen, ACAN and Sox-9 at both molecular and protein levels. Immunolocalisation of Dkk-1 in OA osteochondral plugs showed peri-cellular expression in chondrocytes located in all zones and around migratory endothelial cells invading articular cartilage where there was a quantifiable increase of blood vessel invasion. This later observation was further studied through a series of experiments to investigate the role of Dkk-1 in relation to endothelial cell migration and angiogenesis using an in vitro model of angiogenesis and migration/invasion assays. A novel finding emerged from these studies, which provides evidence for a pro-angiogenic and pro-migratory role of Dkk-1 and to a lesser extent Dkk-2 in human endothelial cell lines. A novel in vitro Transwell co-culture model was developed to study the interaction between chondrocytes and endothelial cells mimicking the osteochondral interface. A novel finding from these studies included the observation that normal or OA-derived chondrocytes appeared to induce an endothelial to mesenchymal transformation (EndMT) of the co-culture endothelial cells. This was assessed by a loss of the endothelial cobble stone morphology and a down-regulation of key factors implicated in endothelial cell phenotype, including VE-cadherin, Tie-2, e-NOS, PDGF-AA and PECAM-1. As endothelial cells lost their phenotype they adopted a spindle morphology and expressed mesenchymal cell markers including: Lumican, Snail, α-SMA, Vimentin and MMPs. Interestingly, this was also associated with an increase in Dkk-1 expression. To confirm a role for Dkk-1 in this process endothelial cells were cultured in the presence of Dkk-1 and were found to undergo EndMT when compared to the control. In summary, this thesis has uncovered several interesting differences in CPC phenotype. In addition, my results suggest that Dkk-1 has potential as a biomarker of OA pathology. This thesis highlights further the complex role of the Wnt Pathway and in particular Dkk-1 may play a role in the pathogenesis of osteoarthritis.

While the genetics of retinoblastoma and the implications of the retinoblastoma susceptibility gene, RB1, are well described, there is still scarce evidence to suggest why RB1 acts in such a cell-type specific manner. Using the murine cortex as a model, we examined the effects of RB1 deletion of cycling neural progenitors and post-mitotic neurons, in order to ascertain cell-type specific functions in the central nervous system. Using the previously identified cell-cycle independent role for Rb in tangential migration, we validated Rb/E2f regulation of neogenin and implicated it in this process. In quiescent cortical neurons, we identified a pivotal role for Rb in neuronal survival. Unlike in cycling progenitors, in post-mitotic neurons Rb specifically represses the expression of cell-cycle associated genes in an E2f-dependent manner. Finally, in cortical neurons in the absence of Rb, we observe an activation of chromatin at E2f associated promoters. To determine the role of direct interaction between Rb and chromatin modifying enzymes, we utilized an acute LXCXE-binding deficient mutant paradigm. We report that the LXCXE binding motif is dispensable in establishment and maintenance of cortical neuron quiescence and survival. The activation state of E2f-responsive promoters appears to be dependent on E2f-activity and not simply Rb-mediated repression. Taken as a whole, this thesis serves to support the hypothesis that Rb plays a diverse role in different cell-types by regulation of unique gene targets and regulatory mechanisms. Characterizing specific cancer-initiating populations and understanding the specific function of Rb will help in the treatment of many cancers resulting from RB1 mutation or mutation within the Rb/E2f pathway.

The origins of respiratory disease, such as asthma in childhood and COPD in later life are unclear. Maternal smoking during pregnancy and low birth weight is associated with increased risk of asthma, poor lung function in adults and COPD in old age. Exposure to oxidative stress and poor nutrition in utero is thought to cause damage to the lung and alter the normal course of lung development. Glutathione S-transferases (GST) are potent antioxidants. In this work, genetic polymorphisms that alter GST enzyme activity were genotyped in a family-based childhood asthma cohort (341 families, n = 1508) and analysed to investigate whether they alter the risk of developing asthma when individuals are exposed to environmental tobacco smoke. Real-time PCR based copy number variation methodology was developed to genotype the common gene deletion polymorphism of GSTT1 and GSTM1 genes, for other GST genes (GSTP1 and GSTO2) SNP haplotypes were constructed. A rare GSTO2 haplotype was negatively associated with asthma susceptibility, atopy severity, and FEV1 values. Asthmatic children with a GSTT1 gene deletion, or a common GSTP1 haplotype, developed more severe asthma compared to individuals with a GSTT1 gene or non-carriers of the GSTP1 haplotype. Total IgE levels were increased in GSTT1*0 individuals when exposed to tobacco smoke in early life, suggesting a gene-environment interaction. GSTO2 may be a shared susceptibility locus for asthma in childhood and COPD in later life. Animal models of maternal protein-restriction during pregnancy can induce hypertension, diabetes and endothelial dysfunction in offspring and in some of these models alterations to lung gene expression and lung architecture have been reported. This work established that a rat model of maternal dietary protein-restriction during pregnancy known to induce hypertension in the offspring, results in persistent alterations to the expression of genes in the lungs of adult offspring (120 days), including genes involved in glucocorticoid action (Hsd11b2), growth (Igf1 & 2 and Pcdh1) and alveolar development (Tp53). Lung microRNA expression profiles were also altered in response to exposure to protein restriction in utero. These findings suggest a role for nutritional programming in respiratory disease susceptibility in later life and a role for microRNAs in the study of the developmental origins of health and disease in general. Further work will include the investigation of epigenetic mechanisms that control nutritional programming in lungs of animals exposed to protein-restriction in utero. This work has demonstrated that GST polymorphism is a risk factor for childhood asthma and certain genotypes can offer some protection against the development of severe asthma. There was little evidence to suggest that GST polymorphism modulates the effects of smoke exposure in early life. In addition, we have demonstrated that maternal diets that are poor in nutrition could predispose her offspring to respiratory disease in later life by altering the course of normal lung development in early life or response to environmental stimuli in later life.

京都大学 / 新制・論文博士 / 博士(医学) / 乙第13418号 / 論医博第2226号 / 新制||医||1052(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 羽賀 博典, 教授 長船 健二, 教授 伊藤 貴浩 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

Den här uppsatsen behandlar de s.k. inställelsereglerna i RB:s femte kapitel. Frågan om hur rättegångens aktörer bör inställa sig ställdes på sin spets under coronapandemin och gav upphov till denna uppsats. I uppsatsen redogörs således för hur inställelsereglerna ser ut och hur de tillämpas. Uppsatsen är främst skriven med utgångspunkt i Ekelöfs teleologiska metod. Med andra ord har huvudsakligt fokus varit att utreda vilka de bakomliggande ändamålen med inställelsereglerna är. Inställelsereglerna har, för att söka svar på frågan om vilka ändamålen är, satts i sitt sammanhang, där jag utrett hur gällande rätt ser ut, hur de systematiskt förhåller sig till andra regler i RB samt hur reglerna förhåller sig till övriga grundläggande principer för den svenska rättegången. Då skillnader mellan fysiska och digitala möten belyses har även ett avsnitt tillägnats den psykologiska innebörden av digitalt deltagande. I arbetets slutskede har jag fört en diskussion kring hur jag anser att tillämpningen av reglerna i 5 kap. 10 § RB motsvarar det tilltänkta ändamålet bakom densamma och om det funnits utrymme för justering i detta avseende. Här har jag kommit fram till att, mot bakgrund av det jag kommit fram till under arbetet med uppsatsen, det finns skäl att se över dels hur tillämpningen av bestämmelsen i 5 kap. 10 § RB bör ske, dels hur utformningen av lagtexten bör vara. Den justering jag förespråkat är en mer extensiv tillämpning av reglerna samt en mer generös utformning av bestämmelsen vad gäller dels vilka aktörer som omfattas, dels hur lämplighetsbedömningen i 5 kap. 10 § fjärde stycket RB ska gå till. / The main subject for this essay has been the regulations concerning court appearance in accordance with the 5th chapter of the Swedish code of judicial procedure. The subject has been of interest concerning the trials the society had to endure during the corona pandemic. In this essay I analyze the rules and how they are practiced today. The essay has been written using the method of Ekelöf called “the teleological method” where the main focus is to find the purpose behind the rules. For the understanding of the purpose of the rules they have been put into context, systematically for instance, and how they relate to other rules and principles for the Swedish judicial procedure. One chapter has been dedicated to the study of psychology since it is an aspect of interest when examining the rules of court appearance, especially regarding the difference between physical and digital meetings. The last chapters are dedicated to the discussion of whether the rules as of now are compliant with its purpose and, if not, there is a need to review the law and how it is being interpreted. One of my conclusions is that there is a need and that the interpretation of the law needs to expand in a more generous way, both concerning what kind of participants who should be included and also when it is considered to be appropriate to participate using the alternative way with videoconference technic.

The level structures of some single closed shell nuclei (⁸⁷₃₇Rb₅₀ and ⁸⁸₃₈Sr₅₀) and nearly closed shell nuclei (⁸⁷₃₈Sr₄₉ and ⁸⁸₃₇Rb₅₁) have been determined by means of beta decay processes of ⁸⁷Kr, ⁸⁸Kr and ⁸⁸Rb to final states ⁸⁷Rb, ⁸⁸Rb and ⁸⁸Sr respectively, and by means of the thermal neutron capture reaction on strontium (⁸⁶Sr(n,ɣ)⁸⁷Sr and ⁸⁷Sr(n,ɣ)⁸⁸Sr). The gamma radiation was studied using a Ge(Li) spectrometer and a Ge(Li)-NaI(Tl) coincidence spectrometer. Beta groups were identified using a plastic scintillator and a plastic-NaI(Tl) coincidence spectrometer. Spins and parities were determined for some of the levels from the deduced decay schemes. / Thesis / Doctor of Philosophy (PhD)

No description available.

No description available.

Esta pesquisa foi realizada com o objetivo de identificar a zona do solo de onde o K foi absorvido e acumulado na soja (safra) e nas plantas em sucessão (segunda safra: milho, braquiária e milho consorciado com a braquiária). Para tanto, utilizou o elemento rubídio (Rb) como marcador do nutriente potássio (K), o qual foi posicionado na superfície do solo, a 30 cm e 60 cm de profundidade. A contribuição relativa de as camadas do solo para a ciclagem de K será assumida como proveniente dos primeiros 30 cm de solo (Rb na superfície), de 30 cm a 60 cm (Rb a 30 cm) e de 60 cm a 90 cm de profundidade (Rb a 60 cm). Ainda, a ciclagem de K foi obtida indiretamente, a partir da recuperação de Rb, com base na hipótese de que a mesma equivale à recuperação de K do solo. O experimento foi realizado na Universidade Federal de Goiás, Regional de Jataí, no ano agrícola 2015/2016. O delineamento experimental foi em blocos casualizados, com parcelas subdivididas e quatro repetições. As parcelas constituem os sistemas de produção (soja, soja/milho, soja/braquiária e soja/milho-braquiária), e as subparcelas correspondem às profundidades onde foi posicionado o Rb (superfície do solo, 30 e 60 cm de profundidade). Na soja, a maior parte do K (60%) acumulado foi absorvida da camada superficial do solo, e nas plantas em sucessão a contribuição foi das camadas subsuperficiais, abaixo de 30 cm de profundidade (braquiária - 74%, milho-braquiária - 71% e milho - 81%). A recuperação do K reciclado da parte aérea da soja foi maior para a forrageira (84%), seguida pelo consórcio milho-braquiária (13%) e milho (2,5%). / This research was carried out to identify the soil depth from where K was absorbed and accumulated in the soybean (first season) and in the plants of succession (second season: corn, brachiaria and corn-brachiaria intercropped). In order to do so, it was used the rubidium element (Rb) as marker of potassium nutrient (K), which was positioned on the soil surface, 30 cm and 60 cm depths. The relative contribution of the soil layers to the K cycling will be assumed as the amount coming from the first 30 cm of soil (Rb at the surface), from 30 cm to 60 cm (Rb at 30 cm) and from 60 cm to 90 cm depth (Rb at 60 cm). Also, the cycling of K was obtained indirectly, from the recovery of Rb based on the hypothesis that it has equal recovery of K from the soil. The experiment was carried out at the Federal University of Goiás, Jataí Regional, in the season of 2015/2016. The experimental design was split-plot with randomized blocks and four replicates. The plots constitute the production systems (soybean, soybean/corn, soybean/brachiaria and soybean/corn-brachiaria intercropped), and the subplots correspond to the depths where the Rb was placed (soil surface, 30 and 60 cm deep). In soybean, most of the K uptake (60%) was taken from the soil surface layer, and in the plants in succession the contribution was from the subsurface layers, below 30 cm depth (brachiaria - 74%, Brachiaria-corn - 71% and corn - 81%). The recovery of recycled K of the soybean above ground was higher in the forage (84%), followed by the corn-brachiaria intercropped (13%) and corn (2.5%).

Ce travail, à la fois expérimental et théorique, présente l’étude d’un processus d’excitation nucléaire, appelé NEET (Nuclear Excitation by Electron Transition), faisant intervenir un cou-plage entre le noyau et le cortège électronique. Dans celui-ci, une désexcitation électronique peut induire une excitation nucléaire si les transitions nucléaire et atomique sont résonantes et ont la même multipolarité. Le noyau de84Rbest un bon candidat pour mettre en évidence ce processus dans un plasma créé par laser car il possède une transition de basse énergie (environ 3 keV) entre l’état isomérique (Jπ= 6−,T1/2= 20,26min) et l’état Jπ= 5−(T1/2= 9ns). Afin d’évaluer un taux d’excitation par effet NEET, il est nécessaire de décrire les états atomiques dans un plasma et de caractériser précisément la transition nucléaire.Afin d’obtenir une description précise des états atomiques dans un plasma, une méthode de calcul de structure atomique a été développée. Cette méthode est basée sur le code de physique atomique MCDF (Multi-Configuration Dirac-Fock) et emploie une technique de sélection des configurations électroniques les plus probables en tenant compte des propriétés du plasma. Cette méthode de calcul a été validée par l’interprétation d’un spectre X émis par un plasma de Rb produit avec le laser PHELIX du GSI à une intensité de6×1014W/cm2.L’énergie de la transition nucléaire entre les états6−et5−n’était pas connue avec une précision suffisante (∼200eV) pour une évaluation précise du taux d’excitation par effet NEET.Deux expériences de spectroscopie γ ont alors été réalisées auprès des accélérateurs ELSA duCEA/DAM/DIF et Tandem de l’IPN d’Orsay. Elles ont permis d’améliorer de plus d’un ordre de grandeur la précision sur l’énergie de cette transition.A l’issue de ce travail, une évaluation du taux d’excitation par effet NEET dans un plasma a été réalisée afin de dimensionner une expérience. D’après celle-ci, le plasma doit avoir une température suffisamment élevée (de l’ordre de 400 eV) afin d’obtenir un nombre d’isomères excités suffisant pour être détectés. / This experimental and theoretical work deals with the Nuclear Excitation by Electron Tran-sition (NEET) process which involves a coupling between the nucleus and its electron cloud. In this process, an electron de-excitation can induce a nuclear excitation if the atomic and nuclear transitions are resonant and have the same multipolarity. This process could be observed in a laser created plasma of 84Rb because this nucleus has a low energy transition (around 3 keV) between the isomeric state (Jπ= 6−,T1/2= 20,26m) and theJπ= 5−(T1/2= 9ns) state. To evaluate a NEET excitation rate, the atomic states in plasma must be described and the nuclear transition must be precisely characterised.To describe the atomic states in plasma, a method based on a MCDF (Multi-ConfigurationDirac-Fock) code was developed. This one uses a procedure to select the most probable atomic configurations according to the plasma properties. This method was checked by the interpretation of a X-rays spectrum emitted by a Rb plasma. This plasma was produced by the PHELIXlaser of the GSI laboratory at an intensity of6×1014W/cm2.The energy of the nuclear transition between the states6−and5−was not accurate enough for the NEET rate evaluation. Twoγ-rays spectroscopy experiments were conducted at the ELSA accelerator from CEA/DAM/DIF and at the Tandem accelerator from the Orsay laboratory. The accuracy of the nuclear transition energy was improved of more than one order of magnitude. In this work, the NEET rate was evaluated to predict an experiment. The Rb plasma must have a high temperature (around 400 eV) to obtain a sufficient number of excited isomers and therefore to make possible the detection.