Made available in DSpace on 2016-08-29T15:35:04Z (GMT). No. of bitstreams: 1 tese_5907_.pdf: 1034669 bytes, checksum: 7bf3706412d3c23a40de589e5805ec80 (MD5) Previous issue date: 2012-09-14 / O aumento da expectativa de vida atingido com a moderna terapia antirretroviral significa que a população infectada pelo HIV está exposta a diversas co-morbidades não infecciosas que se comportam como doenças que se agravam com a idade. O objetivo deste trabalho foi estudar a prevalência de alterações renais, ósseas e cardiovasculares, assim como os fatores associados com essas alterações, em pacientes infectados pelo vírus HIV atendidos em um serviço ambulatorial de AIDS. Exames de urina, dosagens de creatinina, glicose, lipídeos, avaliação de risco cardiovascular, determinação quantitativa de HIV-RNA, contagem de linfócitos TCD4/TCD8 foram realizados no serviço de assistência especializada da Santa Casa de Vitória. A medida da massa óssea por densitometria foi feita no CEDOES (Centro de Pesquisa e Diagnóstico de Osteoporose do Espírito Santo). Quatro trabalhos foram escritos com os resultados do projeto: Um relato de caso de uma fratura supracondilar espontânea de fêmur em um paciente com bom controle de carga viral do HIV; um artigo com dados de baixa massa óssea em 54,7% de 300 pacientes estudados, com associação com menopausa, sexo masculino e índice de massa corpórea menor que 25; outro artigo descrevendo 40,6% de anormalidades urinárias e 9,8% de redução da filtração glomerular em 254 pacientes estudados, sendo esta última associada com raça negra, hipertensão arterial, idade acima de 50 anos, proteinúria e hematúria; e um quarto artigo que identificou diabetes em 5,8%, intolerância a glicose em 28,5% e alterações de colesterol em 22,3% de 498 pacientes após terapia antirretroviral, sendo este último associado com uso de lopinavir/r. Nossa conclusão é que o manejo clinico adequado destas doenças que se agravam com a idade são essenciais no tratamento adequado de indivíduos HIV-AIDS. Palavras-chaves: co-morbidades, HIV, osteoporose, risco cardiovascular, alterações renais.

Background:Approximately 15% of the world’s population are affected bysome kind of disability where over 150 conditions may affect the human gaitpattern. The ability to ambulate with ease is important for overall well-being.Various assistive devices have been developed to improve mobility of theirusers. A lot of research is currently focused on ankle exoskeletons, showingpromising results in providing important assistance during stance phase of gait. Objective:To investigate how different combinations of active and passiveelements in an ankle exoskeleton affects the metabolic cost of walking. Methods:Musculoskeletal simulations were carried out in OpenSim Moco.Different assistive configurations were tested over one gait cycle using a pas-sive element, an active element, and a parallel connection of the both. Parame-ter values were modified to find the most optimal setup for reducing metaboliccost. Results:All assistive configurations were found successful in reducing bothwhole-body metabolic cost and the metabolic cost of the plantarflexors whencompared to the unassisted gait. Most whole-body metabolic cost reductionwas found when using a passive spring with resting length of 0.28 m and stiff-ness of 6 kN/m in parallel with an active motor capable of providing forceequal to 150% of body weight. The most reduction in metabolic cost of theplantarflexors was also found for a parallel connection of elements, but herewith a 100% body weight motor and spring with rest length of 0.19 m andstiffness of 10 kN/m. With higher assistance, more reduction in ankle mo-ment generated by the muscles was observed. Conclusion:Powered ankle exoskeletons are promising in terms of minimiz-ing metabolic cost during walking due to assistance during late stance phaseof gait for ambulators requiring plantarflexor assistance. Keywords:Simulation, exoskeleton, ankle, moco.

Neurons are specialized cells dedicated to transmit the nerve impulses throughout the human body across specialized structures called synapses. At the synaptic terminals, a crosstalk between multiple macromolecules regulates the structure and function of the presynaptic nerve endings and the postsynaptic recipient sites. Gephyrin is the central organizer at inhibitory postsynaptic specializations and plays a crucial role in the organization of these structures by anchoring GABAA receptors (GABAAR) and glycine receptors (GlyR) to the postsynaptic membrane. This 93 kDa protein features an N-terminal G domain and a C-terminal E domain and the latter interacts directly with the intracellular loop between transmembrane helices 3 and 4 of certain subunits of the GlyRs and GABAARs. Biochemical and structural analyses have already provided valuable insights into the gephyrin-GlyR interaction. Interestingly, biochemical studies on the gephyrin-GABAAR interaction demonstrated that the GABAARs also depend on the same binding site as the GlyRs for the interaction with the gephyrin, but the molecular basis for this receptor specific interaction of gephyrin was still unknown. Co-crystal structures of GephE-GABAAR α3- derived peptides with supporting biochemical data presented in this study deciphered the receptor-specific interactions of gephyrin in atomic detail. In its moonlighting function, gephyrin also catalyzes the terminal step of the evolutionarily conserved molybdenum cofactor biosynthesis. Molybdenum, an essential transition element has to be complexed with a pterin-based cofactor resulting in the formation of the molybdenum cofactor (Moco). Moco is an essential component at the active site of all molybdenum-containing enzymes with the exception of nitrogenase. Mutations in enzymes involved in this pathway lead to a rare yet severe disease called Moco deficiency, which manifest itself in severe neurodevelopmental abnormalities and early childhood death. Moco biosynthesis follows a complex multistep pathway, where in the penultimate step, the N-terminal G domain of gephyrin activates the molybdopterin to form an adenylated molybdopterin intermediate. In the terminal step, this intermediate is then transferred to the C-terminal E domain of gephyrin, which catalyzes the metal insertion and deadenylation reaction to form active Moco. Previous biochemical and structural studies provided valuable insights into the penultimate step of the Moco biosynthesis but the terminal step remained elusive. Through the course of my dissertation, I crystallized the C-terminal E domain in the apo-form as well as in complex with ADP and AMP. These structures shed lightonto the deadenylation reaction and the formation of a ternary E-domain-ADP-Mo/W complex and thus provide structural insight into the metal insertion mechanism. Moreover, the structures also provided molecular insights into a mutation leading to Moco deficiency. Finally, ternary complexes of GephE, ADP and receptor-derived peptides provided first clues regarding the integration of gephyrin’s dual functionality. In summary, during the course of the dissertation I was able to derive high resolution structural insights into the interactions between gephyrin and GABAARs, which explain the receptor-specific interaction of gephyrin and, furthermore, these studies can be extended in the future to understand GABAAR subunit-specific interactions of gephyrin. Finally, the understanding of Moco biosynthesis shed light on the molecular basis of the fatal Moco deficiency. / Neurone sind spezialisierte Zellen, die über die Synapsen Nervenimpulse im menschlichen Körper übertragen. An den synaptischen Enden reguliert ein Netzwerk aus einer Vielzahl von Makromolekülen die Struktur und die Funktion der präsynaptischen Nervenenden und der postsynaptischen Kontaktstellen. Gephyrin ist der Hauptorganisator an inhibitorischen, postsynaptischen Spezialisierungen und spielt durch die Verankerung von GABAA-Rezeptoren (GABAAR) und Glycinrezeptoren (GlyR) in der postsynaptischen Membran eine zentrale Rolle für den Aufbau dieser Strukturen. Dieses 93 kDa Protein enthält eine N-terminale G-Domäne (GephG) und eine C-terminale E-Domäne (GephE), wobei letztere direkt mit der intrazellulären unstrukturierten Region zwischen Transmembranhelices 3 und 4 bestimmter Untereinheiten der GlyR und GABAAR interagiert. Biochemische und strukturelle Analysen lieferten bereits wertvolle Erkenntnisse über die Gephyrin-GlyR Interaktion. Interessanterweise zeigten Versuche zur Gephyrin-GABAAR Interaktion, dass GABAARs die gleiche Bindungsstelle auf Gephyrin benutzen wie GlyRs, wobei die molekulare Basis für diese Interaktion nicht bekannt war. In dieser Arbeit zeige ich Co-Kristallstrukturen von GephE-GABAARα3 sowie unterstützende biochemische Daten, die die atomaren Details der rezeptorspezifischen Interaktionen von Gephyrin entschlüsseln. Als zweite Funktion katalysiert Gephyrin den terminalen Schritt der evolutionär konservierten Molybdän Cofaktor Biosynthese. Dabei muss das essentielle Übergangselement Molybdän mit einem Pterin-basierten Cofaktor komplexiert werden, um den Molybdän Cofaktor (Moco) zu bilden. Moco ist essentieller Bestandteil im aktiven Zentrum aller Molybdän-enthaltenden Enzyme mit Ausnahme der Nitrogenase. Mutationen in Enzymen, die in die Molybdän Cofaktor Biosynthese involviert sind, verursachen eine Moco Defizienz, eine seltene, jedoch schwere Erkrankung, die sich durch schwere neurologische Entwicklungsstörungen und Tod im frühen Kindesalter äußert. Die Moco Biosynthese folgt einem komplexen mehrstufigen Ablauf. Im vorletzten Schritt adenyliert GephG das Molybdopterin und ein Zwischenprodukt entsteht. Im letzten Schritt wird dieses Zwischenprodukt auf GephE übertragen, das die Insertion des Metalls und die Deadenylierungsreaktion katalysiert, wodurch der aktive Moco entsteht. Frühere biochemische und strukturelle Studien brachten wertvolle Erkenntnisse über den vorletzten Schritt der Moco Biosynthese, aber die Kenntnisse über den letzten Schritt blieben vage. Während meiner Dissertation kristallisierte ich GephE in der apo-Form sowie im Komplex mit ADP oder AMP. Diese Strukturen gaben Aufschluss über die Deadenylierungsreaktion und die Formation eines ternären GephE-ADP-Mo/W Komplexes und gewährten so einen strukturellen Einblick in den Mechanismus der Metallinsertion. Darüber hinaus ermöglichten die Strukturen eine Mutation, die zu Moco Mangel führt, auf molekularer Ebene zu verstehen. Schließlich lieferten ternäre Komplexe aus GephE, ADP und von Rezeptoren abgeleiteten Peptiden ersten Aufschluss bezüglich der Verflechtung von Gephyrins dualer Funktion. Zusammenfassend konnte ich während der Dissertation hochauflösende strukturelle Einblicke in den Komplex aus GephE und GABAAR α3 Untereineinheit gewinnen, die die rezeptorspezifische Interaktion von Gephyrin erklären. Weiterhin können diese Studien in der Zukunft ausgeweitet werden, um die GABAAR-untereinheitenspezifische Interaktion mit Gephyrin zu verstehen. Schließlich erlauben die Studien zur Moco Biosynthese die tödliche Moco Defizienz auf molekularer Ebene zu verstehen.

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Made available in DSpace on 2014-12-17T15:18:10Z (GMT). No. of bitstreams: 1 IvandilsonPPM.pdf: 2514597 bytes, checksum: 43b19b0aeeca04f9d3f3a1defc9d82cc (MD5) Previous issue date: 2009-02-26 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Brazil is one of the major centers of diversity for polyploid cotton plants; these plants belong to the genus Gossypium, which has three known species: G. hirsutum, G. barbadense and G. mustelinum. The Northeast is the only region where the three species occur, the last group being endemic. Northeast s cotton plants can be important sources of variability for genetic breeding. It is believed that great part of local diversity is being lost, due to economic, political, cultural and agricultural problems. In an attempt to mitigate this loss and delineate conservation strategies it is necessary to know how the species are found where they occur. The objective was to characterize and determine how plants are maintained in situ in the states of Maranh?o, Piau?, Cear?, Rio Grande do Norte and Para?ba at the beginning of the XXI century. The in situ characterization of G. hirsutum and G. barbadense was conducted through structured interviews with the cotton plants owners and through the analysis of the environment. The data were collected during expeditions undertaken between the years 2004 to 2005. Twenty-two plants were collected in the state of Para?ba, forty-four in the state of Rio Grande do Norte, one hundred and forty-six in the state of Cear?, forty in the state of Maranh?o and ninety-one plants in the state of Piau?. All plants collected in the states of Para?ba and Rio Grande do Norte belonged to moco type. Moco cotton plants also predominated in the other states, representing 92%, 62% and 78% of plants collected in Cear?, Piau? and Maranh?o, respectively. The other cotton plants collected belong to the species G. barbadense. The cotton plants were found in situ as dooryard plants, roads side, feral populations, cultivation or local varieties. Great part were dooryard plants (45.2%), being major in Piau? and Maranh?o. Cultivation predominated in Cear?; in Rio Grande do Norte feral populations were the most frequent and, in Para?ba, local varieties. The maintenance of moco plants is related, mainly, to the phytotherapic domestic use (20.9%) and to confection of lamp wicks (29.7%). Few inhabitants in Para?ba, Rio Grande do Norte, Piau? and none in Maranh?o used harvest the plants, storage the seeds or gin; however, in Cear?, 40.5% of owners affirmed that they harvested and commercialized the fiber. It was found that the maintenance of species is dependent of the fragile cultural habits of local inhabitants, therefore the maintenance in situ is not a suitable way to conservation of genetic resources. The efforts must be directed to the continuity of collections, maintenance and characterization ex situ / O Brasil ? um dos importantes centros de diversidade de algodoeiros polipl?ides pertencente ao g?nero Gossypium, com 3 esp?cies conhecidas: G. hirsutum, G. barbadense e G. mustelinum. O Nordeste ? a ?nica regi?o com ocorr?ncia das tr?s esp?cies, sendo a ?ltima end?mica. Os algodoeiros desta regi?o podem ser fontes importantes de variabilidade para o melhoramento gen?tico. Acredita-se que grande parte da diversidade local esteja sendo perdida, devido a problemas econ?micos, pol?ticos, culturais e agr?colas. Na tentativa de mitigar tal perda e delinear estrat?gias de conserva??o ? necess?rio conhecer como as esp?cies se encontram no local em que ocorrem. Objetivou-se caracterizar e determinar o modo com que as plantas s?o mantidas in situ nos estados do Maranh?o, Piau?, Cear?, Rio Grande do Norte e Para?ba no inicio do s?culo XXI. A caracteriza??o in situ de G. hirsutum e G. barbadense foi realizada por meio de entrevista estruturada com o propriet?rio e pela an?lise do ambiente. Os dados foram tomados durante expedi??es empreendidas entre os anos de 2004 a 2005. Foram coletadas 22 plantas no estado da Para?ba, 44 no estado do Rio Grande do Norte, 146 no estado do Cear?, 40 no estado do Maranh?o e 91 plantas no estado do Piau?. Todas as plantas coletadas nos estados da Para?ba e Rio Grande do Norte eram do tipo moc?. O algodoeiro moc? tamb?m predominou nos demais estados, representando 92%, 62% e 78% das plantas coletadas no Cear?, Piau? e Maranh?o, respectivamente. Os demais algodoeiros coletados pertencem a esp?cie G. barbadense. Os algodoeiros moc?s foram encontrados in situ na forma de planta de fundo de quintal, beira de estrada, feral, lavoura, variedade local. Em grande parte eram do tipo fundo de quintal (45,2%), sendo maioria no Piau? e Maranh?o. No Cear? predominou o tipo lavoura, no Rio Grande do Norte tipo feral e na Para?ba variedades locais. A manuten??o das plantas do tipo moc? est? ligada, principalmente, ao uso dom?stico fitoter?pico (20,9%) e confec??o de pavios para candeeiro (29,7%). Poucos moradores na Para?ba, Rio Grande do Norte, Piau? e nenhum no Maranh?o apresentaram o h?bito de realizar a colheita, armazenamento e beneficiamento das sementes, entretanto no Cear? 40,5% dos propriet?rios afirmaram realizar a colheita e comercializar a fibra. Verificou-se que a manuten??o da esp?cie ? dependente dos fr?geis h?bitos culturais da popula??o local, portanto a manuten??o in situ n?o ? um meio adequado ? conserva??o dos recursos gen?ticos. Os esfor?os devem ser direcionados para a continuidade das coletas, caracteriza??o e manuten??o ex situ

A previous study identified Enterococcus faecalis as one of two bacteria known to have the selD gene and other selenium related genes without having the genes necessary to make selenocysteine or selenouridine. EF2570, a gene in the cluster, was later shown to be upregulated during biofilm formation and also responsible for a selenite- and molybdate-dependent increase in biofilm formation in vitro. The protein encoded was identified as a selenium dependent molybdenum hydroxylase (SDMH), enzymes that contain a labile selenium atom required for activity. While the process of inserting selenocysteine into a protein is well known, the process by which a SDMH acquires a labile selenium atom has not yet been described. To begin unraveling this pathway, the nifS-like EF2568 from the gene cluster will be characterized. Some NifS-like proteins have been shown to have selenocysteine lyase activity, providing a source of selenium for selenophosphate synthetase, the selD gene product. Study of EF2568 has shown that it specifically reacts with L-selenocysteine to form selenide and alanine with L-cysteine inhibiting the reaction. Guided by homology to the well-characterized human and E. coli NifS-like proteins, mutants of the active site and substrate discerning residues were also characterized for activity with L-selenocysteine and L-cysteine. While mutation of the residue at position 112 thought to be responsible for substrate specificity did not affect reactivity of the enzyme with L-cysteine, it did affect reactivity with L-selenocysteine. Studying the characteristics of this novel group II selenocysteine lyase will provide a foundation for studying the remaining pathway.

Made available in DSpace on 2014-12-17T15:36:43Z (GMT). No. of bitstreams: 1 RuteAS.pdf: 1886240 bytes, checksum: 64a629cdddb6fa7a9a7cd0db535bac2c (MD5) Previous issue date: 2006-06-28 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Kerodon rupestris (rock cavy, moc?) is an endemic caviidae of Brazilian northeast that inhabits rocky places in the semi arid region. The aim of this study was to characterize the activity/rest rhythm of the rock cavy under 12:12 h LD cycle and continuous light. In the first stage, seven animals were submitted to two light intensities (LD; 250:0 lux and 400:0 lux; 40 days each intensity). In the second stage four males were kept for 40 days in LD (470:<1 lux), for 18 days in LL 470 lux (LL470) and for 23 days in red dim light below 1 lux (LL<1). In the third stage three males were initially kept in LD 12:12 h (450:<1 lux) and after that in LL with gradual increase in light intensity each 21 days (<1 lux LL<1; 10 lux-LL10; 160 lux LL160; 450 lux LL450). In the fourth stage it was analyzed the motor activity of 16 animals in the first 10 days in LD. Motor activity was continuously recorded by passive infrared movement sensors connected to a computer and totaled in 5 min bins. The activity showed circadian and ultradian rhythms and activity peaks at phase transitions. The activity and the rest occurred in the light as well as in the dark phase, with activity mean greater in the light phase for most of the animals. The light intensity influenced the activity/rest rhythm in the first three stages and in the first stage the activity in 400 lux increased in four animals and decreases in two. In the second stage, the tau for 3 animals in LL470 was greater than 24 h; in LL<1 it was greater than 24 h for one and lower for two. In the third stage the tau decreased with the light intensity increase for animal 8. During the first days in the experimental room, the animals did not synchronize to the LD cycle with activity and rest occurring in both phases. The results indicate that the activity/rest rhythm of Kerodon rupestris can be affected by light intensity and that the synchronization to the LD cycle results from entrainment as well as masking probably as a consequence of the action of two or more oscillators with low coupling strength / Kerodon rupestris ? um roedor cav?deo end?mico do nordeste do Brasil, que habita locais pedregosos da regi?o do semi-?rido. Este trabalho teve como objetivo caracterizar o ritmo de atividade motora e repouso do moc? em ciclo claro/escuro (CE) 12:12 h e em luz constante. Na 1a etapa, sete animais foram submetidos a duas intensidades de luz na fase de claro (CE 12:12 h 250:0 lux; 400:0 lux 40 dias em cada intensidade). Na 2a etapa, mantiveram-se 4 machos em ciclo CE 12:12 h (470:<1 lux) por 47 dias, em claro constante 470 lux (CC470) por 18 dias e sob luz vermelha constante de menos 1 lux (CC<1) por 23 dias. Na 3a etapa, 3 machos foram mantidos em CE 12:12 h (450:<1 lux) e depois em CC com aumento gradual da intensidade a cada 21 dias, de menos de 1 lux com luz vermelha (CC<1) para 10 (CC10), 160 (CC160) e 450 lux (CC450) respectivamente. Na 4a etapa foi analisada a atividade motora de 16 moc?s nos dez primeiros dias em CE 12:12 h. A atividade motora foi registrada continuamente por sensores de movimento por infravermelho, acoplados a um computador, com totaliza??es a cada 5 minutos. A atividade dos moc?s apresentou ritmos circadianos e ultradianos e picos de atividade nas transi??es de fase. A atividade e o repouso ocorreram tanto no claro quanto no escuro, com m?dia de atividade maior no claro para a maioria dos animais. A intensidade de luz afetou o ritmo de atividade/repouso dos animais nas tr?s primeiras etapas, de modo que na 1a etapa, a atividade em 400 lux aumentou em 4 animais e diminuiu em dois. Na 2a etapa, o tau de tr?s moc?s em CC470 foi maior do que 24 h, enquanto que em CC<1 foi menor em dois animais e maior em um. Na 3a etapa, o valor do tau diminuiu com o aumento da intensidade de luz no animal 8, variou pouco no animal 7. Durante os primeiros dias na sala de registro os animais n?o sincronizaram ao ciclo CE 12:12 h e tanto a atividade quanto o repouso ocorreram nas duas fases do ciclo. A an?lise dos resultados indica que o ritmo de atividade/repouso de Kerodon rupestris pode ser afetado pela intensidade luminosa e que a sincroniza??o ao ciclo CE ocorre devido ao arrastamento da atividade e tamb?m como conseq??ncia de mascaramento, provavelmente devido a atua??o de 2 ou mais osciladores com baixa for?a de acoplamento