Submitted by Ubirajara Cruz (ubirajara.cruz@gmail.com) on 2017-05-16T12:47:48Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertacao_Tatiele_Munbach.pdf: 829847 bytes, checksum: 819f6655b2f1238488bb374450563f95 (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2017-05-16T16:45:55Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertacao_Tatiele_Munbach.pdf: 829847 bytes, checksum: 819f6655b2f1238488bb374450563f95 (MD5) / Made available in DSpace on 2017-05-16T16:45:55Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertacao_Tatiele_Munbach.pdf: 829847 bytes, checksum: 819f6655b2f1238488bb374450563f95 (MD5) Previous issue date: 2015-02-11 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / O período de maior suscetibilidade a doenças em sistemas intensivos de produção de leite é durante o período de transição, o qual corresponde a três semanas antes até três semanas após o parto. O mesmo é caracterizado por um balanço energético negativo devido à mobilização de gordura das reservas corporais e por mudanças no ambiente ruminal pela introdução súbita de grãos na dieta. Com a finalidade de reduzir os efeitos do balanço energético negativo e promover o aumento na produção de leite, algumas medidas têm sido adotadas em relação ao manejo nutricional no período de transição. O uso de leveduras, por exemplo, têm sido uma boa alternativa para tentar melhorar o ambiente ruminal e facilitar às adaptações da microbiota a nova dieta. O objetivo do estudo foi avaliar os efeitos da suplementação de uma associação de levedura viva e hidrolisada nos marcadores metabólicos e do fluído ruminal de vacas leiterias durante o período de transição. O experimento foi conduzido em um sistema semi-extensivo, utilizando-se 20 vacas da raça Holandesa, divididas igualmente em grupo controle (GC) e grupo Suplementado (GS). O GS recebeu adição diária de 28g/animal/dia da associação de levedura S. cerevisiae desde os 20 ± 2 dias pré-parto até o início da lactação (18 ± 3 dias). As coletas de amostras de fluído ruminal e de sangue foram realizadas no pré-parto, e durante seis dias após o parto. No liquido ruminal foi avaliado o pH, Tempo de redução do azul de metileno, contagem de protozoários e avaliação morfo-tintorial das bactérias. No soro sanguíneo foram determinadas as concentrações de ácidos graxos não esterificados (AGNE’s), albumina e ureia. No metabolismo energético, os AGNE’s diminuíram durante o período do colostro nos animais suplementados, assim como a albumina e ureia no período da lactação. Houve mudança no ambiente bacteriano ruminal nos animais suplementados, ocorrendo um predomínio de bactérias Gram negativas. No ambiente ruminal as bactérias Gram negativas aumentaram significativamente no GS durante a lactação. Concluiu-se que a suplementação com uma associação de levedura viva e hidrolisada em vacas durante o período de transição pode influenciar positivamente o metabolismo energético e proteico assim como o ambiente bacteriano ruminal no início da lactação. / The period of greatest susceptibility to disease in intensive milk production systems is during the transition period, which corresponds to three weeks before up to three weeks after delivery. The same is characterized by a negative energy balance due to body fat mobilization and changes in the rumen by the sudden introduction of grain in the diet. In order to reduce the effects of negative energy balance and increase milk production, some measures have been taken into account in nutritional management during the transition period. The use of yeast, for example, have been a good alternative to improve the rumen and facilitating adjustment of the microbiotato the new diet. The aim of the study was to evaluate the effects of association of live and hydrolyzed yeast in the metabolic markers and ruminal fluid examination of dairy cows during the transition period. The experiment was conducted in a semi extensive system, using 20 Holstein cows, divided equally into control group (CG) and Supplemented group (SG). The SG received daily addition of 28 g / animal / day of S. cerevisiae yeast association from 20 ± 2 days pre-calving until early lactation (18 ± 3 days). The collections of rumen fluid and blood samples were performed during labor, and for six days after delivery. In ruminal fluid was evaluated pH, time of blue methylene reduction, protozoa count and morpho-dye bacteria evaluation. Serum concentrations of non-esterified fatty acids (NEFA's), albumin and urea were determined. There was a change in rumen bacterial environment in the supplemented animals, experiencing a predominance of gram negative. The number of Gram negative bacteria increased significantly in GS during lactation. In energy metabolism, the NEFA's decreased over the period of colostrum in the supplemented animals as well as albumin and urea in the period of lactation. We conclude that supplementation of an association of live and hydrolyzed yeast to cows during the transition period can positively influence the energy and protein metabolism as well as the rumen bacterial environment in early lactation.

The alarming rate of increase in incidence and prevalence of the type 2 diabetes mellitus has prompted intense research on understanding the pathogenesis of the type 2 diabetes. It is observed that the development of type 2 diabetes is preceded by a state of insulin resistance and obesity. Previous studies have suggested that the obesity induced insulin resistance may be mediated by elevated levels of circulating free fatty acids (FFAs). The increase in circulating levels of FFAs may be contributed by the release of FFAs from stored triglycerides (TG) in adipocytes via lipolysis. It is hypothesized that the decrease in levels of circulating FFAs by sequestration and storage of FFAs in adipocytes may prevent deleterious effects of FFAs on insulin sensitivity. Recently our lab and others have shown that the storage of TG in adipocytes is promoted by a novel protein, Fat Specific Protein 27 (FSP27). Although, these studies also revealed FSP27 to be a lipid droplet associated protein that suppresses lipolysis to enhance TG accumulation in adipocytes, the role of FSP27 in lipolysis remains largely undetermined. Therefore, this study investigates the role and regulation of FSP27 in adipocytes in both the basal state, as well as during lipolysis. The studies presented here show FSP27 to be a remarkably short-lived protein (half-life=15 min) due to its rapid ubiquitination and proteasomal degradation. Thus, I tested the hypothesis that lipolytic agents like the cytokine, TNF-α and the catecholamine isoproterenol modulate FSP27 protein levels to regulate FFA release. Consistent with this concept, TNF-α markedly decreased FSP27 mRNA and protein along with lipid droplet size as it increased lipolysis in cultured adipocytes. Similarly, FSP27 depletion using siRNA mimicked the effect of TNF-α to enhance lipolysis, while maintaining stable FSP27 protein levels by expression of HA epitope-tagged FSP27 blocked TNF-α mediated lipolysis. In contrast, the robust lipolytic action of isoproterenol is paradoxically associated with increases in FSP27 protein and a delayed degradation rate that corresponds to decreased ubiquitination. This catecholamine-mediated increase in FSP27 abundance, probably a feedback mechanism to restrain excessive lipolysis by catecholamines, is mimicked by forskolin or 8-Bromo-cAMP treatment, and prevented by Protein Kinase A (PKA) inhibitor KT5720 or PKA depletion using siRNA. These results show that isoproterenol stabililizes FSP27 via the canonical PKA pathway and increased cAMP levels. However, the work presented here also suggests that FSP27 does not get phosphorylated in response to isoproterenol treatment, and the stabilization of FSP27 is independent of isoproterenol mediated lipolysis. The data presented in this thesis not only identifies the regulation of FSP27 as an important intermediate in mechanism of lipolysis in adipocytes in response to TNF-α and isoproterenol, but also suggests that FSP27 may be a possible therapeutic target to modulate lipolysis in adipocytes.