The impact of nutritional intervention on affective states has largely been ignored in the exercise-affect literature. For decades the impact of such interventions on perceptions of exertion has been well documented. However, Hardy and Rejeski (1989) assert that `what' a person feels, as measured by the rating of perceived exertion (RPE) scale, may be very different from `how' they feel, and that on its own the RPE provides limited information about the subjective experiences of individuals during exercise. This thesis describes a series of studies that assess the influence of various fluid ingestion regimes on both `how' and `what' a person feels. Seven studies were undertaken, incorporating a variety of exercise modes, including prolonged running (Study 1,3 & 7), prolonged cycling (Study 2& 4) and prolonged intermittent, high intensity exercise (Study 5,6 & 7). The relationship between fluid ingestion during exercise and affective states during and following exercise proved to be a complex one. The initial investigation (Study 1) showed that the ingestion of water during prolonged running resulted in an overall improvement in valence during the recovery period. A significant increase in activation was also noted in the water trial only, from pre to post exercise. Furthermore, subjective ratings of energy post-exercise were higher in the water trial, compared to the no water trial. In study 2 the beneficial effects observed in study 1 were not so apparent. In this instance the only significant change of interest was in energetic arousal, which was found to be higher 5 min post exercise in the water trial compared to the no water trial. When the ingestion of a CHO solution during exercise was compared to a placebo or flavoured water solution (Studies 3-7) the findings also varied. However, the observation of an enhanced affective profile following CHO ingestion in Study 4 and Study 5 highlights the importance of considering nutritional status and intervention when investigating the exercise-affect relationship. These studies have highlighted some important aspects in our understanding of the exercise-affect relationship alone. Firstly, a robust finding across all the studies was the observation of an almost uniformly positive shift in valence from the final within-exercise assessment to the post exercise assessments. Thus emphasising the dynamic nature of affect and the importance of repeated within exercise assessments. Secondly, moderate intensity exercise of a fixed duration was marked by highly variable inter-individual differences in the response of participants to the valence and activation dimensions. However, exercise to fatigue elicited a homogenous valence response as participants came closer to reaching their exercise capacity.

Thesis (D. Miss.)--Trinity Evangelical Divinity School, 1994. / Includes abstract. Includes bibliographical references (leaves 190-198).

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Background Timing and quantity of protein (PRO) consumption are important considerations for muscle protein synthesis (MPS), fat-free mass (FFM) accretion, and body fat % (BF%) reduction. The effect of PRO ingestion on changes in FFM is mediated by many variables. Past studies have focused on specific composition of carbohydrate (CHO) and PRO consumption (CHO vs. PRO + CHO), and have also investigated PRO intake timing at pre-exercise, post-exercise, or both. Other studies have investigated FFM maintenance and growth with increased PRO consumption during catabolic or anabolic phases of energy balance (EB). These mechanisms have been studied in various populations, including healthy untrained individuals, overweight and obese people, and endurance athletes. However, studies have not explored relationships between the amount and timing of PRO ingested, and the state of EB, as it relates to FFM%. Method/Design A retrospective analysis design was used to assess relationships between PRO ingestion, timing, and EB on FFM in collegiate football players. Subjects were members of an intercollegiate Division 1 football team, had completed a one-day food and activity record, and had body composition assessed as part of a regular team screening procedure. Data acquisition was supervised by a PhD/Registered Dietitian. Food and activity records were analyzed using NutriTiming®, which predicts RMR via the Harris-Benedict equation, uses a MET-based relative intensity activity scale, and accesses the USDA Nutrient Database for Standard Reference, Release 26 to predict hourly EB and PRO consumption. EB was assessed as ±400 kcal EB (EBR), < 0 kcal EB (NEGEB), and > 0 kcal EB (POSEB). Total useable PRO (TUP) was defined as the sum of PRO consumed in units up to 30g max/meal, a value also assessed relative to EB at the time of ingestion. The goal was to assess the amount and timing of PRO intake with EB as these factors relate to FFM. Results Pearson's correlations found that BF% was negatively associated with TUP while in EBR (r=-.253; p=0.049), and FFM% was positively associated TUP in EBR (r=0.279; p=0.030) and in POSEB (r=0.282; p=0.028). NEGEB was positively associated with BF% (r=0.325; p=0.011), and negatively associated with FFM% (r=-0.322; p=0.011). Conclusions Results elucidate that players who ingest PRO in a relatively good energy-balanced state had higher FFM% and a lower BF%. Further, those players consuming TUP while in POSEB had an even stronger positive association with FFM% and a stronger inverse association with BF%. These data reject the null hypothesis that football players who consume PRO in POSEB have less FFM% than those who consume PRO in NEGEB.

La sialylation des N-glycanes du fragment Fc des immunogobulines G (IgG) est une modification peu fréquente des IgG humaines. Pourtant, elle est l’objet de beaucoup d’attention depuis que deux articles fondateurs ont été publiés, qui montrent l’un que la sialylation des IgG diminue leur capacité à déclencher la cytotoxicité cellulaire dépendant de l’anticorps (ADCC), et l’autre que les IgG sialylées en α2,6 seraient la fraction efficace des IgG intraveineuses (IgIV) anti-inflammatoires. Les anticorps monoclonaux thérapeutiques, qui sont le plus souvent des IgG recombinantes produites en culture de cellules de mammifère, connaissent depuis la fin des années 90 un succès et une croissance phénoménaux sur le marché pharmaceutique. La maîtrise de la N-glycosylation du Fc des IgG est une clé de l’efficacité des anticorps monoclonaux. Si les IgG sialylées sont des molécules peu fréquentes in vivo, elles sont très rares en culture cellulaire. Dans cette étude, nous avons développé une méthode de production d’IgG avec une sialylation de type humain en cellules CHO. Nous avons travaillé principalement sur la mise au point d’une stratégie de production d’IgG sialylées par co-expression transitoire d’une IgG1 avec la β1,4-galactosyltransférase I (β4GTI) et la β-galactoside-α2,6-sialyltransférase I (ST6GalI). Nous avons montré que cette méthode permettait d’enrichir l’IgG1 en glycane fucosylé di-galactosylé mono-α2,6-sialylé G2FS(6)1, qui est le glycane sialylé présent sur les IgG humaines. Nous avons ensuite adapté cette méthode à la production d’IgG présentant des profils de glycosylation riches en acides sialiques, riches en galactose terminal, et/ou appauvris en fucosylation. L’analyse des profils de glycosylation obtenus par la co-expression de diverses combinaisons enzymatiques avec l’IgG1 native ou une version mutante de l’IgG1 (F243A), a permis de discuter des influences respectives de la sous-galactosylation des IgG1 en CHO et des contraintes structurales du Fc dans la limitation de la sialylation des IgG en CHO. Nous avons ensuite utilisé les IgG1 produites avec différents profils de glycosylation afin d’évaluer l’impact de la sialylation α2,6 sur l’interaction de l’IgG avec le récepteur FcγRIIIa, principal récepteur impliqué dans la réponse ADCC. Nous avons montré que la sialylation α2,6 augmentait la stabilité du complexe formé par l’IgG avec le FcγRIIIa, mais que ce bénéfice n’était pas directement traduit par une augmentation de l’efficacité ADCC de l’anticorps. Enfin, nous avons débuté le développement d’une plateforme d’expression stable d’IgG sialylées compatible avec une production à l’échelle industrielle. Nous avons obtenu une lignée capable de produire des IgG enrichies en G2FS(6)1 à hauteur de 400 mg/L. Cette étude a contribué à une meilleure compréhension de l’impact de la sialylation sur les fonctions effectrices des IgG, et a permis d’augmenter la maîtrise des techniques de modulation du profil de glycosylation des IgG en culture cellulaire. / Only a fraction of the N-glycans present on the Fc fragment of the human IgGs is sialylated. However, a new interest for sialylation has risen since two major articles were published, one showing that sialylation reduces the capacity of the antibody to trigger antibody-dependent cell cytotoxicity (ADCC), whereas the other showed that the IgGs carrying α2,6-sialic acids on their Fc N-glycans were responsible for the anti-inflammatory activity of intravenous immunoglobulins (IVIGs) injected at high doses. Therapeutic monoclonal antibodies (mAbs) are in majority recombinant IgGs produced in mammalian cell culture. Since the end of the nineties, mAbs have become a major class of pharmaceutical products, and their success is still growing. The control of Fc N-glycosylation is a key parameter for the improvement of the therapeutic efficacy of mAbs. Sialylated IgGs are found only as traces in the classic CHO cell culture processes. In this study, we developed a method for the production of IgGs with a human-like sialylation in CHO cells. We focused on a production strategy relying on the transient co-expression of an IgG1 with the β1,4-galactosyltransferase I (β4GTI) and the β-galactoside-α2,6-sialyltransferase I (ST6GalI). We showed that this method allowed the enrichment of the IgG1 glycoprofile in the fucosylated di-galactosylated mono-α2,6-sialylated glycane G2FS(6)1, which is the main sialylated glycan found in human IgGs. We then adapted this method to the production of highly galactosylated or highly sialylated IgGs with and without core-fucosylation. The analysis of the glycosylation profiles obtained using the various enzyme combinations co-expressed with the native IgG1 or the mutant IgG1 F243A allowed us to discuss the influence of the under-galactosylation found in IgGs produced in CHO cells versus the Fc structural constraints on the limitation of IgG sialylation in CHO cells. We used the IgG1 glycovariants produced with our method to assess the impact of Fc α2,6-sialylation on the interaction of the IgG with the receptor FcγRIIIa, which is the main receptor mediating the ADCC response. We showed that the presence of α2,6-sialylation in the Fc increased the stability of the IgG-FcγRIIIa complex. This benefit however did not translate into an improved ADCC capacity. Finally, we initiated the development of a stable expression platform for the production of sialylated IgGs at yields relevant for the industry. We obtained a cell line capable of producing IgGs enriched in G2FS(6)1 at 400 mg/L. This may eventually represent a novel approach to manufacture a recombinant IVIG surrogate. With this work, we contributed to a better understanding of the impact of sialylation on the effector functions of IgGs. We also improved our understanding of the techniques allowing for the modification and control of the glycosylation profile of IgGs in cell culture.

Therapeutic proteins, a.k.a. biopharmaceuticals, are most commonly produced in expression systems derived from Chinese Hamstery Ovary (CHO) cells, thanks to great capacity of post-translational modifications like secretation, folding and glycosylation. The engineering of cells for regulation of protein expression has many options including knock-in and knock-out of genes, epigenetic studies or improvement of the expression casette of the protein of interest by e.g. promotor variants or modifications of the 5’ and 3’ untranslated region (UTR). The 3’-UTR is therefore a good optimization candidate for attempting to achieve increased expression of therapeutic proteins. The final aim of this study was to identify and design 3’-UTRs for improved expression of therapeutic proteins in HyClone™ CHO cells from GE Healthcare Bio-Sciences AB (GEHC). The impact goal is to increase the efficiency and lower the costs for pharmaceutical companies when producing biopharmaceuticals in the HyClone™ CHO cell line, leading to increased accessibility of monoclonal antibodies (mAbs) on the pharmaceutical market. The study was initiated with bioinformatic analysis of the CHO cell transcriptome from a set of RNA-seq data of HyClone™ CHO to find high expressing, context independent genes. The 3’-UTRs from the best candidate genes were used for construction of plasmids for expression of a Fc-eGFP fusion protein. Nine selected 3’-UTRs were designed, synthesized and cloned into a parent plasmid (pGE0520) creating nine plasmid variants (pGE0523-531). The constructed plasmids were used for evaluation with site directed integration (SDI) into the HyClone™ CHO cell line and expression analysis were performed by flow cytometry and antibody titer measurements from cells with successfully integrated plasmid sorted by fluorescence-activated cell sorting (FACS).   Result show a significant effect on protein expression when using different variants of 3’-UTRs. Two variants, pGE0524 and pGE0526, competing with the parent plasmid in expression levels and integration efficiency from SDI, making them candidates for further investigations against the parent plasmid. Results also show good correlation between flow cytometry data from pre- and post-sorting, which can make research for further 3’-UTRs more efficient by evaluations and prediction of expression levels before cell sorting.

Submitted by PPG Hist?ria (historia-pg@pucrs.br) on 2017-11-03T16:34:10Z No. of bitstreams: 1 TESE - LUCIANA DA COSTA DE OLIVEIRA - VERS?O MENOR.pdf: 14177891 bytes, checksum: 99e2fd93dd19fb5b7b8d954e21be268a (MD5) / Rejected by Caroline Xavier (caroline.xavier@pucrs.br), reason: Devolvido devido a 1) Diferen?a de entre o t?tulo da capa/t?tulo da publica??o cadastrada, e o t?tulo do restante do material (folha de rosto, ficha catalogr?fica, folha de aprova??o da banca). 2) Data de defesa cadastrada na publica??o (31/08/2017), est? diferente da data do arquivo PDF (22/08/2017) on 2017-11-16T17:39:40Z (GMT) / Submitted by PPG Hist?ria (historia-pg@pucrs.br) on 2017-11-20T19:47:40Z No. of bitstreams: 1 final_corrigido.pdf: 13712674 bytes, checksum: 0fae3ff53f2742992410ac18fecb8139 (MD5) / Approved for entry into archive by Caroline Xavier (caroline.xavier@pucrs.br) on 2017-11-28T18:59:51Z (GMT) No. of bitstreams: 1 final_corrigido.pdf: 13712674 bytes, checksum: 0fae3ff53f2742992410ac18fecb8139 (MD5) / Made available in DSpace on 2017-11-28T19:05:50Z (GMT). No. of bitstreams: 1 final_corrigido.pdf: 13712674 bytes, checksum: 0fae3ff53f2742992410ac18fecb8139 (MD5) Previous issue date: 2017-08-22 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / The main objective of this doctoral thesis is to analyze the way in which the image of the gaucho and the ga?cho was constructed and consecrated in the Argentine, Uruguayan and Brazilian painting of the 20th century, especially in the painting by Ces?reo Bernaldo de Quir?s (1879-1968), Pedro Figari (1861-1938) and Pedro Weing?rtner (1853-1929). In this perspective, starting from theoretical-methodological issues that include not only the dialogues that the images interlock, but, especially, their temporal and mnemic links, it was also aimed to establish a relation between the work that celebrated the gaucho and the ga?cho in the twentieth century with those that preceded it. Thus combining the nascent image of the eighteenth and nineteenth centuries with that which was consecrated in the twentieth century, it was intended to define the diverse and multiple threads that, intertwined, produced a wide and complex network of meanings around the image of the gaucho and the ga?cho. These same threads, woven by times and memories, by yearnings and desires, besides particularizing the image of the man from the pampa, also reveals their uses and their functions. / La presente tesis doctoral, tiene como objetivo principal analizar la forma con lo cual la imagen gaucha y del ga?cho fue construida y consagrada en la pintura argentina, uruguaya y brasile?a del siglo XX, en especial en la pintura de Ces?reo Bernaldo De Quir?s (1879-1968), Pedro Figari (1861-1938) y Pedro Weing?rtner (1853-1929). En ese sentido, y partiendo de las cuestiones te?rico metodol?gicos que perciben no solamente los di?logos que las im?genes se traban entre s?, pero, sobre todo, en sus encadenamientos temporales y mn?micos, se ha objetivado, tambi?n, establecer una relaci?n entre la obra que ha celebrado el gaucho y el ga?cho en el siglo XX junto aquellas que los precedieron. As?, conjugando la imagen de los siglos XVIII y XIX, se pretendi?, pues, definir los diversos y m?ltiples hilos que entrelazados, produjeron una amplia y compleja red de significados acerca de la imagen del gaucho y del ga?cho. Esos mismos hilos, tramados por el tiempo y por memorias, por anhelos y deseos, aparte particularizar la imagen del hombre del pampa, igualmente revela sus usos y funciones. / A presente tese de doutorado tem como objetivo central analisar a forma com a qual a imagem do gaucho e do ga?cho foi constru?da e consagrada na pintura argentina, uruguaia e brasileira do s?culo XX, especialmente na pintura de Ces?reo Bernaldo de Quir?s (1879- 1968), Pedro Figari (1861-1938) e Pedro Weing?rtner (1853-1929). Nesse sentido, partindo de quest?es te?rico-metodol?gicas que percebem n?o apenas os di?logos que as imagens travam entre si, mas, sobretudo, seus encadeamentos temporais e mn?micos, objetivou-se, tamb?m, estabelecer uma rela??o entre a obra que celebrou o gaucho e o ga?cho no s?culo XX junto das que lhe antecederam. Assim conjugando a imagem nascente dos s?culos XVIII e XIX junto daquela que foi consagrada no XX, pretendeu-se, pois, definir os diversos e m?ltiplos fios que, entrela?ados, produziram uma ampla e complexa rede de significa??es no entorno da imagem do gaucho e do ga?cho. Esses mesmos fios, tramados por tempos e mem?rias, por anseios e desejos, afora particularizar a imagem do homem do pampa, igualmente descortina seus usos e suas fun??es.