Thesis (Ph.D.)--Medical University of Ohio, 2006. / "In partial fulfillment of the requirements for the degree of Doctor of Philosophy in Medical Sciences." Major advisor: Han-Fei Ding. Includes abstract. Document formatted into pages: iv, 163 p. Title from title page of PDF document. Title at ETD Web site: NF-KappaB2 is an autoimmunity regulator and its mutation leads to lymphomagenesis in mice. Bibliography: pages 67-77,106-112,134-161.

Orientadores: Anibal Eugenio Vercesi, Helena Coutinho Franco Oliveira / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-08T02:32:32Z (GMT). No. of bitstreams: 1 Alberici_LucianeCarla_D.pdf: 5846392 bytes, checksum: a41c4e31bbe1d1fa3a670c7a9956806e (MD5) Previous issue date: 2006 / Resumo: Alterações no metabolismo energético mitocondrial promovidas por proteínas desacopladoras (UCPs) são frequentemente encontradas em desordens metabólicas. Recentemente demonstramos que camundongos hipertrigliceridêmicos (HTG) apresentam uma respiração mitocondrial de repouso elevada, não relacionada a UCPs. Neste trabalho, nós elucidamos o mecanismo responsável por esta elevação da respiração de repouso e demonstramos algumas conseqüências dessa resposta mitocondrial à hiperlipidemia no fígado e no metabolismo corporal total. Resultados: Mitocôndrias isoladas de fígados e de células mononucleares de baço de camundongos HTG apresentaram velocidades respiratórias elevadas comparadas aos camundongos controles. Mudanças no consumo de oxigênio em mitocôndrias de fígados de camundongos HTG foram sensíveis a ATP, diazóxido e ácido 5-hidroxidecanóico (5-HD) indicando que o consumo pode ser atribuído à atividade dos canais de K+ sensíveis a ATP (mitoKATP). Do mesmo modo, as mitocôndrias HTG apresentaram um maior inchamento na presença de íons K+, sensível aos agonistas e antagonistas do mitoKATP. Além disso, a ligação de glibenclamida marcada às mitocôndrias indica que os camundongos HTG expressaram maiores quantidades de receptores de sulfoniluréias, um componente os mitoKATP. Aumento da velocidade de metabolismo foi evidenciado por um aumento no consumo de oxigênio no fígado (sensível ao tratamento agudo in vivo pela administração de 5-HD), elevada temperatura retal e maior produção corporal de CO2 nesses camundongos. De acordo com a velocidade metabólica elevada, a ingestão alimentar foi significantemente maior em camundongos HTG, sem concomitante aumento de peso. Assim como verificado em camundongos HTG, mitocôndrias de fígados de animais submetidos à dietas ricas em substratos energéticos apresentaram também elevação da respiração mitocondrial de repouso. Conclusões: Esses resultados demonstram que a hiperlipidemia primária leva ao aumento da atividade dos mitoKATP em fígados, o que pode representar uma adaptação regulada para oxidar o excesso de ácidos graxos em camundongos HTG. Além disso, nossos resultados indicam que os mitoKATP, além das UCPs, podem estar envolvidos no controle do metabolismo energético e do peso corporal / Abstract: Changes in mitochondrial energy metabolism promoted by uncoupling proteins (UCPs) are often found in metabolic disorders. We have recently shown that hypertriglyceridemic (HTG) mice present higher mitochondrial resting respiration unrelated to UCPs. Here, we disclose the underlying mechanism and consequences, in tissue and whole body metabolism, of this mitochondrial response to hyperlipidemia. Results: As observed in HTG mice, liver mitochondria from animals submitted to the rich energy diets presented high resting respiration. Mitochondria isolated from the livers and spleen mononuclear cells of HTG mice presented enhanced respiratory rates compared to those from wild-type mice. Changes in oxygen consumption of liver mitochondria from HTG mice were sensitive to ATP, diazoxide and 5-hydroxydecanoate (5-HD), indicating they can be attributable to mitochondrial ATP-sensitive K+ channel (mitoKATP) activity. Indeed, mitochondria from HTG mice presented enhanced swelling in the presence of K+ ions, sensitive to mitoKATP agonists and antagonists. Furthermore, mitochondrial binding to fluorescent glibenclamide indicates that HTG mice expressed higher quantities of sulfonylurea receptors, a component of mitoKATP. An overall faster metabolic state was evidenced by increased liver oxygen consumption (sensitive to acute in vivo 5-HD administration), higher body CO2 release and temperature in these mice. In agreement with higher metabolic rates, food ingestion was significantly larger in HTG mice, without enhanced weight gain. Liver mitochondria isolated from rats fed glucose rich diet or from mice fed fat rich diet also presented higher resting respiration rates. Conclusions: These results demonstrate that primary hyperlipidemia leads to an elevation in liver mitoKATP activity, which may represent a regulated adaptation to oxidize excess fatty acids in HTG mice. Furthermore, our data indicate that mitoKATP, in addition to UCPs, may be involved in the control of energy metabolism and body weight / Doutorado / Ciencias Biomedicas / Doutor em Ciências Médicas

Orientador: Sara Teresinha Olalla Saad / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-25T10:07:43Z (GMT). No. of bitstreams: 1 Costa_LauremiliaRiconG.R.da_D.pdf: 3335092 bytes, checksum: 1187b50ec4df532e027b86fabe029bc9 (MD5) Previous issue date: 2014 / Resumo: Os eventos de migração e adesão das células progenitoras hematopoéticas são cruciais para a sua manutenção nos nichos da medula óssea e são regulados por diversos fatores, dentre eles as RhoGTPases. ARHGAP21 é uma proteína RhoGAP (RhoGTPase activating protein, que regulam negativamente as RhoGTPases), e atua na migração e adesão celulares. O papel da Arhgap21 na hematopoese ainda é desconhecido. O objetivo geral do presente estudo foi estudar a função da Arhgap21 na migração e adesão das células progenitoras hematopoéticas. Neste trabalho, mostramos que ainda não foi possível a obtenção do camundongo nocaute para Arhgap21, pois parece que morrem antes do nascimento. O estudo foi realizado com os camundongos Arhgap21+/-, cuja expressão gênica e protéica de Arhgap21 está reduzida. As células progenitoras hematopoéticas Arhgap21+/- apresentaram menor quimiotaxia induzida por CXCL-12 e redução da adesão à fibronectina, o que provavelmente levou a um homing ineficiente para a medula óssea e para o baço. Observou-se também que o microambiente da medula óssea e do baço com redução de Arhgap21 leva à ineficiência do homing das células progenitoras hematopoéticas normais. Não foi observada alteração na expressão gênica e na freqüência de células expressando CXCR-4, bem como não houve diferença na secreção de CXCL-12. As células da medula óssea Arhgap21+/- apresentaram maior expressão de Cdc42 e parecem possuir maior atividade desta RhoGTPase. Pela primeira vez se descreve o papel da Arhgap21 na hematopoese normal e ela parece ser um importante regulador da migração, adesão e homing das células progenitoras hematopoéticas, provavelmente pela regulação negativa de Cdc42, através da sua função GAP / Abstract: Migration and adhesion of hematopoietic progenitor cells are crucial events for their maintenance on bone marrow niches. The role and mechanisms underlying hematopoietic progenitors migration and adhesion has been extensively studied and RhoGTPases have been implicated in these events. Regulators and effectors of RhoGTPases functions also have been studied in biology of hematopoietic progenitor cells. ARHGAP21 is a RhoGAP member, which function as negative regulators of RhoGTPases, and plays role in cell migration and adhesion. The aim of this study was to investigate the role of Arhgap21 in migration and adhesion of hematopoietic progenitor cells. It was not possible to generate a knockout mouse until now, because they die before birth, as results indicate. Heterozygous mice (Arhgap21+/-)v have reduction in Arhgap21 gene and protein expression and we used it as model. Hematopoietic Progenitor cells from Arhgap21+/- mice showed reduction in CXCL-12- induced chemotaxis as well as in fibronectin adhesion, which probably leaded to inefficient homing to wild-type bone marrow and spleen observed. In addition, homing of wild-type hematopoietic progenitor cells to Arhgap21+/- bone marrow and spleen was also reduced. It was not observed differences in CXCR-4 gene expression and frequency of progenitor hematopoietic cells expressing this receptor as well as in CXCL-12 secretion in Arhgap21+/- bone marrow or serum. It is the first time that the role of Arhgap21 in hematopoiesis is described and it seems to be an important regulator of hematopoietic progenitor cell migration, adhesion and homing, probably due to Cdc42 negative regulation by its RhoGAP function / Doutorado / Biologia Estrutural, Celular, Molecular e do Desenvolvimento / Doutora em Fisiopatologia Médica

Abstract Mammalian extracellular matrix metalloproteinases, MMPs, are a family of enzymes capable of degrading components of the connective tissue. The in vivo regulation of the cell lineage-specific expression of MMPs, however, is not well known. This study used transgenic mice to identify cell-specific elements in the upstream regulatory regions of MMP-2 and MMP-9. Transgenic mice were generated by pronuclear microinjections into fertilised oocytes using lacZ as a reporter gene. The reporter gene constructs containing varying lengths of the MMP-9 5'-upstream region revealed an area that allowed for expression in osteoclasts and migrating keratinocytes, the cells that also express MMP-9 in vivo. The sequence driving the cell specific expression included the nucleotides from -2722 to -7745. When the same upstream regulatory fragment of MMP-9 was used to drive the expression of the human tissue specific inhibitor of MMPs, TIMP-1, instead of lacZ, the transgenic mice developed normally and the animals were fertile with normal post-embryonic growth. However, cutaneous wound healing was remarkably retarded, but not totally prevented, and the migration of keratinocytes over the wound was slow. The mice expressed the human TIMP-1 in keratinocytes during wound healing and in situ zymography revealed a total blockage of the gelatinolytic activity of MMP-2 and MMP-9, the main gelatinases active in the healing wound tissues. By using a sequence of 6500 base pairs from the 5'-upstream regulatory region of the MMP-2 gene it was possible to drive the expression of lacZ in mesenchymal cells of the developing transgenic mouse embryo. The expression pattern was similar to that found in previous in situ hybridization studies, following the different stages of tissue morphogenesis and being present in the areas of basement membrane degradation and epithelial cell invasion. Computer analyses of the sequence revealed three regulatory upstream regions conserved between human, mouse, and rat, and possibly responsible for the cell-and tissue specificity. New transgene constructs containing fragments of the conserved regions will provide a more detailed profile of the in vivo MMP-2 regulation in the future. This study defined a fragment in the upstream regulatory region of MMP-9 that is essential for expression in osteoclasts and migrating keratinocytes. Furthermore, the keratinocyte derived MMPs, including MMP-9, were found to play important role in epithelial cell migration in the area of the healing wound.

No description available.

Dissertation (Ph.D.)--University of South Florida, 2008. / Includes vita. Includes bibliographical references.

FGFR 3 is one of a family of four high affinity receptors for FGF ligands. Activating mutations in FGFR 3 result in skeletal dysplasias that vary in severity from undetectable to neonatal lethal. Mice with congenital deficiency of FGFR3 develop severe kyphosis and skeletal overgrowth. FGFR3 is also expressed in calvarial pre-osteoblasts, osteoblast and articular chondrocytes, although it biological role in these cells remains undefined. By changing the genetic background of the Fgfr3-/- mice we were able to extend their lifespan and examine its impact on post-natal skeletal growth. To investigate the implication of FGFR 3 in post-natal cartilage and bone metabolism we used a combination of imaging, classic histology, molecular biology and biomechanical testing. The results demonstrated that the synovial joints of young adult Fgfr3-/- mice revealed a progressive deterioration, loss of the joint space width and changes in the subchondral bone. These alterations were accompanied by an increase of cartilage matrix degradation. Increased aggrecan and collagen type II degradation products, generated by MMPs were detected with DIAPEN and COL2-3/4C antibodies. Increased collagen type X, cellular hypertrophy and loss of proteoglycan at the articular surface were also demonstrated. A novel micro-mechanical indentation protocol revealed that the humeral heads of Fgfr3-/- mice were less stiff than those of wild type littermates. On the other hand, young adult Fgfr3-/- mice are osteopenic due to reduced cortical bone thickness and defective trabecular bone mineralization. The reduction in mineralized bone and lack of trabecular connectivity observed by micro-computed tomography were confirmed by histological and histomorphometric analyses, which revealed a significant decrease in calcein labeling of mineralizing surfaces and a significant increase in osteoid in the long bones of 4-month-old Fgfr3-/- mice. Primary cultures of adherent bone marrow-derived cells from Fgfr3-/- mice expressed markers of differentiated osteoblasts but developed fewer mineralized nodules than Fgfr3+/+ cultures of the same age. These data point to a major role for FGFR3 signaling in development and homeostatic maintenance of cartilage and bone post-natally and identify FGFR3 as a potential target for intervention in degenerative disorders of cartilage, osteopenia and those associated with defective bone mineralization.

Thesis (M.Sc.). / Written for the Dept. of Human Genetics. Title from title page of PDF (viewed 2008/12/09). Includes bibliographical references.

Thesis (Ph.D. in Immunology) -- University of Colorado Denver, 2008. / Typescript. Includes bibliographical references (leaves 138-153). Free to UCD Anschutz Medical Campus. Online version available via ProQuest Digital Dissertations;

Thesis (Ph. D.)--University of Oklahoma. / Bibliography: leaves 155-161.