This PhD project explored the role of tricarboxylic acid (TCA) cycle enzymes in regulating Chinese hamster ovary (CHO) cell metabolism with respect to growth and recombinant protein expression. It was hypothesised that regulation of central metabolism can influence CHO cell productivity in terms of biomass and protein production. Thus the aim of these studies was to identify the key metabolic reactions of the TCA cycle associated with growth and protein expression in CHO cells. The gene expression of all TCA cycle genes was independently knocked-down using RNAi technology. The small interfering RNA (siRNA) mediated silencing of 11 TCA cycle genes significantly reduced cellular growth along with a decline in adenylate energy charges and an increase in catabolic reduction charges. The gene profiling of glucose and amino acid metabolism (not targeted by siRNA) suggested siRNA mediated knock-down of targeted TCA cycle genes led to cellular stress along with an enhanced rate of glycolysis leading to channelling of glucose for the generation of pyruvate. For the purpose of estimating intracellular metabolites, quenching and extraction method using ammonium bicarbonate and methanol was optimised to use with UCB CHO-K1 cell line and static transient siRNA transfections. A gas chromatography-mass spectrometry (GC-MS) analysis post-silencing of the aconitase gene, which catalyses the conversion of citrate to isocitrate in the TCA cycle, yielded higher MS peak intensities of at least four metabolites (gluconic acid, lysine, threonine and leucine) 72 h post-transfection in comparison to the controls. Transient knock-down of gene expression of seven TCA cycle genes in a recombinant stable cell line (expressing a rabbit monoclonal antibody) reduced cellular growth and altered the energy charges leading to a decline in antibody expression. Although silencing of the pyruvate dehydrogenase E1 gene, which is the component of the pyruvate dehydrogenase complex connecting glycolysis to the TCA cycle, did not affect cell viability, a reduction in antibody expression was recorded. Seven TCA cycle genes which demonstrated the most significant effect on cellular growth and energy charges were transiently over-expressed along with a monoclonal antibody in CHO-K1 cells with addition of their corresponding preceding intermediates. No differences in protein expression and cell specific productivity were observed compared to the control transfections. These results could be due to limitations of the effects of transient transfections for enhancing the metabolic activity of CHO cells. The aconitase gene demonstrated the most significant effect on CHO cell growth and proliferation in this study, therefore this gene was proposed as a novel selection marker for a metabolic selection system for the generation of recombinant therapeutics. This PhD project also established metabolite analysis tools and siRNA protocols for future metabolomic studies for investigating the intracellular CHO metabolism. The findings validated the hypothesis that TCA cycle plays an important role in CHO cell growth and recombinant protein production. The key metabolic genes affecting cellular growth and altering energy metabolism can be further explored for generation of an energy efficient CHO host-cell line (by over-expression of key TCA cycle genes) for enhanced recombinant protein production.

Syfte – Examensarbetets syfte är att skapa förståelse kring ett outsourcingbeslut genom att utveckla en beslutsmodell för outsourcing. För att uppnå syftet ska följande frågeställningar besvaras: Vilka faktorer bör påverka ett outsourcingbeslut? Vilka kostnader bör analyseras för att ett outsourcingbeslut ska bli väl avvägt? Vilka komponenter bör ingå i en beslutsmodell för outsourcing? Metod – Examensarbetet bygger på en analytisk konceptuell studie med ett deduktiv och induktiv förhållningssätt, där påverkansfaktorer togs fram genom litteraturstudier. Jämförelser och analyser av redan befintliga teorier gjordes för att identifiera påverkansfaktorer, utifrån dessa faktorer skapades en beslutsmodell för outsourcing. Resultat – Resultatet från studien visar att det finns ett flertal olika faktorer som påverkar ett outsourcingbeslut och kan sammanfattningsvis placeras som påverkansfaktorer, strategiska- och konkurrensfaktorer samt kostnadsfaktorer. Den utvecklade beslutsmodellen för outsourcing består av sju steg där varje steg innefattar olika analyser och beslut, de sju stegen baseras på de faktorer som bör påverkas vid outsourcing. Förslag till fortsatta studier – För att utveckla beslutsmodellen skulle fler teorier och faktorer som möjligen kan beröras i samband med outsourcing adderas. En ytterligare studie skulle kunna vara att utveckla ett verktyg där organisationen kan gradera hur organisationen förhåller sig till de faktorer som påverkas vid ett outsourcingbeslut och därmed utöka beslutsunderlaget. Praktiska konsekvenser – För att underlätta för beslutsfattaren och få en helhetsbild över faktorer och kostnader som påverkas vid outsourcing kan beslutsmodellen användas. Originalitet – Det finns enligt författarnas vetskap ingen fullständig beslutsmodell som belyser alla faktorer som kan påverkas vid ett outsourcingbeslut. / Purpose – The purpose of the thesis is to create understanding of the outsourcing decision making by developing a decision model for outsourcing. To achieve this, the following questions are answered: What factors should influence the outsourcing decision? What costs should be analyzed so that an outsourcing decision should be balanced? What components should be included in a decision model for outsourcing? Method – The thesis is based on an analytical conceptual study with a deductive and inductive approach, where the influencing factors was generated through literature reviews. Comparison and analysis of existing theories were made to identify the influencing factors, based on the generated influencing factors a decision model for outsourcing was created. Findings – Results from the study shows that there are several factors that influence the outsourcing decision and can in summary be positioned as strategic and competitive factors and cost factors. The developed decision model for outsourcing consists of seven steps, where each step includes various analyzes and decisions, the seven steps are based on the factors that should be affected by outsourcing. Research limitations/implications – The results in this thesis are based only on theory. The model can still be generalized by being adaptable for different Organization. Practical implications – Decision model can be used to aid and support in an outsourcing decision. To facilitate the decision-maker and get an overview of the factors and costs that are affected by the outsourcing decision model can be used. With the decision model, the organization can analyze factors and costs to obtain a comprehensive decision-making. Originality – Since many organizations are making outsourcing decision wrongly without taking into account all the factors affected by the outsourcing decision. There is, in the authors' knowledge no complete decision model that highlights all the factors that may be affected by an outsourcing decision.

OBJETIVOS:- Estimar la prevalencia de trastornos de la conducta alimentaria (TCA) según diferentes niveles de severidad (diagnóstico de anorexia y bulimia nerviosa, de trastornos de la conducta alimentaria no especificados (TCANE) y síntomas de TCA) en adolescentes del municipio de Tarragona y de zona rural de la misma provincia.- Describir y analizar según los diferentes niveles de severidad de alteración de la conducta alimentaria.- Valorar la influencia de los diferentes factores sobre el riesgo de desarrollo de TCA.MATERIAL Y MÉTODOS:Estudio epidemiológico transversal en dos fases con adolescentes de ambos sexos que cursaban 1º y 3º de ESO (12-16 años) en 26 centros públicos y privados de la provincia de Tarragona. Primera fase (n=2967): se valoró la prevalencia de síntomas de alteración de la conducta alimentaria mediante dos test de cribado: Eating Attitudes Test (EAT) y Youth´s Inventory. Segunda fase: se estudió sólo el sexo femenino (n=295). Con los resultados de los test se seleccionaron dos grupos de individuos en función de la presencia (EAT>25 y Youth´s (+) para TCA) o ausencia de riesgo (controles) de TCA; valorandose: satisfacción y autopercepción corporal, influencia sociocultural, clima familiar, síntomas psicopatológicos, actividad física, conocimientos, preferencias y hábitos alimentarios mediante cuestionarios. Se valoró individualmente: el consumo de alimentos mediante el recordatorio de 24 horas, la composición corporal y se realizó la entrevista "Schedules for Clinical Assessment in Neuropsychiatry" (SCAN) en versión DSM-IV, CIE-10 para la obtención de diagnósticos.RESULTADOS:El 13,3-17,1% de los participantes fueron considerados a riesgo de padecer un TCA (18,9-22,5%) mujeres, (7,6-11,7%) hombres dependiendo del test de cribado considerado. El nivel educativo de los padres es más bajo en las adolescentes con síntomas de TCA. Las situaciones familiares estresantes se asocian a una mayor prevalencia de síntomas de TCA. La prevalencia estimada de TCA en el sexo femenino es de 5,2% (IC 95%: 5-5,4), de anorexia nerviosa (0,53%) (IC 95%: 0,5-0,55), bulimia (0,66%) (IC 95%: 0,63-0-68) y TCANE (4 %) (IC 95%: 3,83-4,17 ). El nivel socioeconómico y educativo de los padres no esta significativamente asociado con TCA. En las adolescentes con TCANE se producen más situaciones familiares estresantes: divorcio padres, convivir con la nueva pareja del progenitor y/o con otras personas ajenas al núcleo familiar y no convivir con la madre. A medida que aumenta la severidad de la alteración de la conducta alimentaria, las adolescentes describen a sus familias con menor cohesión, mayor conflictividad y control; se evidencia también un aumento progresivo de síntomas psicopatológicos.Las adolescentes con TCANE son más corpulentas que el resto, el 77,3% de ellas y el 40% de las adolescentes a riesgo refieren elevada insatisfacción corporal. Hay una disminución progresiva en el consumo de alimentos, energía y macronutrientes en relación a la severidad de la alteración de la conducta alimentaria. Las adolescentes con riesgo de TCA y las adolescentes con TCANE consumen respectivamente alrededor de 500 y 900 Kcal. menos que las controles. La probabilidad de ingesta inadecuada es superior en las adolescentes a riesgo y las adolescentes con TCANE. El 67% y el 86% respectivamente realizan ingestas de calcio inferiores a las recomendaciones. Hay una mayor probabilidad de ingesta vitamínica inadecuada en adolescentes con TCANE que puede estar relacionado con las alteraciones mentales asociadas y podría ser un factor más que contribuyera a dar continuidad a los TCA.La restricción calórica es un factor de riesgo para cualquier nivel de severidad de alteración de la conducta alimentaria. El riesgo de síntomas de TCA se relaciona con las variables ansiedad y que algún familiar haga dieta, mientras que el riesgo de TCANE se relaciona con un IMC elevado, insatisfacción corporal y una situación de estrés (divorcio padres). / OBJETIVES:- To estimate the prevalence of eating disorders (ED) at a different levels of severity (diagnosed as anorexia and nervous bulimia and eating disorders not otherwise specified (EDNOS) and symptoms of ED) in teenagers from the town of Tarragona and surrounding areas of Tarragona.- To describe and analysed the varying degrees of severity of the EDs.- To evaluate the influence of the different factors which contribute to the risk of contracting EDs.METHODS AND MATERIAL: A cross sectional study in two phases of teenagers of both sexes (12-16 years) who study (1st ,2nd ESO) in 26 both publics and privates schools in the Tarragona region. First phase (n=2967): To evaluate the prevalence of EDs symptoms using two screening test: Eating Attitudes Test (EAT) and Youth's Inventory. Second phase: only females studied (n=295). With the results of the test two groups were selected those which fell into the risk category (EAT>25 or Youth's (+) for ED) and non-risk category of ED (controls); evaluating: body satisfaction and autoperception, sociocultural influence, family climate, psychopathologic symptoms, physical activity, knowledge, preferences and dietary habits using written questionnaires. Individually evaluated was: food consumption using 24-h recall, body composition and the interview "Schedules for Clinical Assessment in Neuropsychiatry" (SCAN) in version DSM-IV, ICD-10 was administered to obtain the diagnoses.RESULTS:13,3-17,1% of participants were considered at risk of suffering from an ED (18,9-22,5% females, 7,6-11,7% males) depending on the screening test used. The education level of parents is lower in parents of those female teenagers with ED symptoms. Stressful family backgrounds are also associated with a greater risk of ED symptoms. The estimate prevalence of ED in females is 5,2% (CI 95%: 5-5,4), nervous anorexia (0,53%) (CI 95%: 0,5-0,55), bulimia (0,66%) (CI 95%: 0,63-0-68) and EDNOS (4 %) (CI 95%: 3,83-4,17 ). Socioeconomic and educational stages of the parents were not associated with ED. EDNOS were more prevalent in teenagers with stressful family situations: divorce, living with a step-parent or unrelated people, and not living with the birth mother. As well as those factors affecting the severity of eating disorders, other factors include teenagers descriptions of the closeness of family, family conflicts and control; the evidence also notes the increase of psychopathologic symptoms. EDNOS are more frequently found among heavily-built teenage girls, 77,3% of those and the 40% of the teenagers at risk of ED reported high body dissatisfaction. There is a steady decline in the daily intake of food, energy and nutrients directly related to the severity of ED. The teenagers at risk of ED y the adolescents with EDNOS consume respectively around 500 to 900 Kcal. less than the controls. The probability of unsuitable intake is higher in those teenagers at risk of EDs and those with EDNOS. 67% and 86% respectively consume less than the recommended amount of calcium. There is a higher probability of unsuitable vitamin intake in adolescents with EDNOS that could be related to mental factors and this in itself could be another factor which contributes to the continuation of EDs. Caloric restraint is a risk factor that applies to all levels of ED. The risk of symptoms of EDs are directly related to factors such as anxiety, and to have any relative on diet; while the risk of EDNOS is related to a high body mass index (BMI), body dissatisfaction and stressful events (divorced parents).

Companies are increasingly outsourcing business critical activities to suppliers of outsourcing services. As the complexity and business  criticality of the outsourced activities increases, the risk of poor performance increases. This thesis studies large scale outsourcing in the telecom industry where a recent trend is to transfer the development, operation and maintenance of the telecom infrastructure to telecom equipment suppliers. The significance of this type of outsourcing is that the outsourced activity is the revenue generating part of the telecom operators business. Part 1 discusses the purpose and research questions followed by the theoretical underpinning in the research. The research strategy is to study the outsourcing relationship in three distinct stages of its development and the theoretical underpinning applies transaction costs analysis in the Scoping & Search stage and Das &Teng’s (2001) framework of trust and control for managing risks in the Negotiation and Transition stages. This design is in response to calls for a more detailed understanding of how organizations manage risks, it therefore takes the perspective of both the outsourcer and the supplier in the research. Part 2 is a multiple case study of telecom operators in Holland, Sweden and Australia where the supplier in all three cases is Ericsson Global Services organization. The study is further supplemented by mini-cases of large scale IS/IT infrastructure outsourcing. Part 3 has three main parts. Firstly, a cross case analysis of the cases in Part 2; secondly, a discussion of the findings linked to the research questions resulting in a set of propositions. The third and final part covers additional insights and learnings from studying business critical outsourcing and suggestions for further research. The main contributions in the research can be summarised as: Physical asset specificity follows transaction costs logic, however human asset specificity is largely ignored by both outsourcer and supplier Business critical outsourcing by its nature faces a limited market for capable suppliers. This results in single-source negotiations followed by a cooperative stance and open book negotiations. Das & Teng’s (2001) framework for management of risks has been found to have specific directions, some bi-directional and others uni-directional. Furthermore, different dimensions in the framework operate at different managerial levels. Goodwill trust-building operate at the corporate executive level, competence trust-building, output and behavioural control at the level of the negotiation team, and the research indicates that the social control dimension is not applied in business critical outsourcing negotiations. A further finding is that goodwill trust-building precedes all other dimensions of trust and control, and is a pre-requisite for establishing a cooperative stance in the negotiations.

This article presents a study conducted on data containing drug concentrations. The data was obtained from femoral venous blood samples collected at medico legal autopsies in Sweden. Cases positive for antidepressant drugs were scrutinized and divided in to two groups for 15 antidepressant drugs: B‑cases, where the cause of death was intoxication with more than one drug detected in the blood sample. C‑cases, where the cause of death was NOT intoxication and at least one drug (the antidepressant) was detected in the blood sample. This data was then processed to find frequencies of concomitant drugs taken together with the antidepressant drugs. Frequencies of the most common concomitant drugs were then compared between B-cases and C-cases for each antidepressant drug. This revealed that the drugs dextropropoxyphene, ethanol, codeine, flunitrazepam, paracetamol, propiomazine and alimemazine were signifcantly more common as concomitant drugs in B-cases (intoxications) than in C‑cases (non‑intoxications). With regards to unknown interactions the most interesting combinations were: Propiomazine with mirtazapine, venlafaxine, citalopram or fluoxetine; Paracetamol with paroxetine; Flunitrazepam with mirtazapine, venlafaxine or citalopram; Codeine with mirtazapine or sertraline. These combinations should be further investigated.

Thesis (D.W.S.)--Institute for Worship Studies, 2007. / Abstract. Includes bibliographical references (leaves 203-212).

Thesis (D.W.S.)--Institute for Worship Studies, 2007. / Abstract. Includes bibliographical references (leaves 203-212).

Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Jurídicas. Programa de Pós-Graduação em Direito / Made available in DSpace on 2012-10-22T23:20:26Z (GMT). No. of bitstreams: 1 248842.pdf: 1585576 bytes, checksum: c14577e63a4fa73dd71d9e251cd5927d (MD5) / A presente dissertação de Mestrado tem como tema central o Tratado de Cooperação Amazônica (TCA) e a sustentabilidade da Região MAP, no período compreendido entre 1992 a 2002. O objetivo é o de verificar se, no período estudado, o Tratado de Cooperação Amazônica, assinado por oito países sul-americanos, dentre eles Peru, Brasil e Bolívia, apresentou-se como um mecanismo efetivo de promoção da sustentabilidade da Região MAP # uma região-fronteira, situada na Amazônia Sul-Ocidental e que compreende o Estado do Acre (BRA) e os Departamentos de Madre de Dios (PER) e Pando (BOL). Para a realização do trabalho, foi feita uma ampla pesquisa monográfica compreendendo, sobretudo, a questão da sustentabilidade amazônica e a evolução institucional do Tratado. A partir da década de 1990, o Tratado de Cooperação Amazônica, adota o desenvolvimento sustentável como tema estratégico, constituindo-se a sua promoção em sua grande missão. Contudo, em razão da sua fragilidade institucional, das políticas nacionais e regionais dos países signatários, contraditórias em relação ao princípio da sustentabilidade, bem como da realidade socioambiental da Região MAP no início do século XXI - que gerou iniciativas locais de cooperação, alternativas ao modelo tradicional de cooperação representado pelo TCA - o discurso de sustentabilidade adotado pelo TCA é um discurso que se apresenta contraditório em relação às práticas das políticas de cooperação amazônica e dos seus países signatários. La presente disertación de Maestría tiene como tema central el Tratado de Cooperación Amazónica (TC 1992 a 2002. El objetivo es verificar se, en el período estudiado, el Tratado de Cooperación Amazónica, asignado por ocho países suramericanos, entre ellos Perú, Brasil y Bolivia, se mostró como un mecanismo efectivo de promoción de la sustentabilidad de la Región MAP # una región-frontera, situada en la Amazonía Sur-Occidental y que comprende el Estado de Acre (BRA) y los Departamentos de Madre de Dios (PER) e Pando (BOL). Para la realización del trabajo, se ha hecha una amplia pesquisa monográfica comprendiendo, sobretodo, a la cuestión de la sustentabilidad amazónica y la evolución institucional del Tratado. A partir de la década de 1990, el Tratado de Cooperación Amazónica adopta el desarrollo sostenible como tema estratégico, constituyendo su promoción en su gran misión. Sin embargo, en razón de su fragilidad institucional, de las políticas nacionales y regionales de los países signatarios, pontradictorias en relación al principio de la sustentabilidad, así como de la realidad socioambiental de la Región MAP en el inicio del siglo XXI - que ha generado iniciativas locales de cooperación, alternativas al modelo tradicional de cooperación representado por el TCA # el discurso de sustentabilidade adoptado por el TCA es un discurso que se muestra contradictorio en relación a las prácticas de las políticas de cooperación amazónica e de sus países signatarios.

Edwardsiella ictaluri is the causative agent of enteric septicemia of catfish (ESC), which is the most economically important disease of farm-raised channel catfish. E. ictaluri is considered a facultative intracellular pathogen like other well-known species in the Enterobacteriaceae, and it is capable of surviving inside channel catfish neutrophils and macrophages. Its ability to survive inside neutrophils and macrophages has made the development of an effective vaccine against ESC particularly challenging. The goal is to develop a safe, efficacious live attenuated ESC vaccine that is practical and economically beneficial to catfish producers. In this study, single and combination of mutations in genes encoding TCA cycle enzyme and C-1 metabolism proteins were constructed using inrame mutagenesis. The virulence, vaccine efficacy, and tissue persistence of the constructed single and combination mutants were determined in channel catfish. The constructed mutants EideltasdhC, Eideltamdh, EideltafrdA, EideltaglyA, EideltasdhCdeltamdh, EideltasdhCdeltafrdA, and EideltasdhCdeltagcvP were significantly attenuated and showed 100% protection against E. ictaluri 93-146 infection in juvenile channel catfish. However, when tested in 15-d old catfish fry, mutant EideltasdhCdeltagcvP and EideltafrdA were found to provide good protection (99% and 60%, respectively) against E. ictaluri 93-146 infection. The tissue persistence study indicated higher tissue concentration in mutants EideltasdhCdeltagcvP and EideltafrdA relative to the tissue concentration in EideltasdhC and EideltasdhCdeltafrdA mutants.

Serial passaging of primary human fibroblasts leads to the formation of non-dividing senescent cells by a process termed replicative senescence. This tissue culture-based methodology is currently used as a model system to determine the underlying mechanisms of in vivo cellular aging and tumor suppression. Senescence is regarded as an alternative pathway to apoptosis, where cells undergo multiple changes in metabolic and cellular signaling pathways in order to prevent proliferation but still maintain a metabolically-active cell. Whether or not this model accurately reflects in vivo processes is presently controversial; however, replicative senescence is currently the most applicable model through which one can investigate the underlying causes of human cellular aging in the context of controlled environmental stress over time. This work was directed at understanding the molecular processes involved in replicative senescence with specific emphasis on the role of the mitochondria. A series of experiments were performed to assess changes during the induction of replicative senescence under conditions of low (3%) and high (20%) oxygen levels. Measurements were made at the transcriptional, protein, and metabolite levels. Microscopy wasalso utilized to monitor changes in mitochondrial morphology and volume. While previous studies have evaluated specific pathways and/or products; this work combines a more complete metabolomic, genomic, proteomic, and morphological picture of cells undergoing senescence and oxidative stress. Considering the low cell population densities of primary adherent fibroblasts and the subsequent low concentrations of small polar metabolites involved in glycolysis and the TCA cycle, methodologies needed to be developed in order to optimize metabolite extraction and liquid chromatography-mass spectrometric analysis. Protein kinase and transcriptional microarrays were also performed in order to quantify the changes in activated/deactivated signaling cascades as well as gene expression and relate these findings to metabolomic data. Mitochondrial dynamics of cells at different age time points and under different oxygen conditions were also assessed including mitochondrial size, shape, membrane potential, and percent volume per cell volume using confocal microscopy. The results obtained not only confirm the major pathways involved in senescence (p53/p21, PTEN/p27, and RTK/Raf/MAPK) but also provide evidence at both the transcriptional and protein levels for additional senescence-associated pathways. The majority of the changes observed were related to pathways involved in cellular stress, cell cycle control, and the survival response. Metabolic data suggested a –pooling effect– of glycolysis and TCA precursor molecules due to attenuation in enzyme function; this theory was also supported by an observed up regulation of gene expression as a compensatory mechanism. Mitochondria exhibited changes in membrane potential as well as volume and percent volume per cell which suggested compensatory hypertrophy and/or attenuation of mitochondrial fission processes. When the aforementioned analyses are tied together, a “theoretical model of senescence” can be formulated and is characterized by increased metabolic protein and associated metabolite levels due to attenuation in their respective enzyme function, resulting in increases in expression of their associated genes as a compensatory mechanism. / Ph. D.