Made available in DSpace on 2015-04-14T13:35:46Z (GMT). No. of bitstreams: 1 450447.pdf: 3645797 bytes, checksum: abdc582487fc7c81fd903cc19cb4f28a (MD5) Previous issue date: 2013-03-25 / Objective: to evaluate the expression of AMP-activated protein kinase (AMPK) and Sirtuin-1 (SIRT-1) in liver tissue samples from patients during bariatric surgery and one year after the procedure. To evaluate the association between the expression of these proteins with weight, Body Mass Index (BMI) and biochemical parameters (glucose, total cholesterol, HDL and triglycerides). Methods: the expression of the proteins was verified by immunohistochemistry and quantified by digital image analysis. All other data were accessed in the database. Results: liver biopsies of 44 patients submitted to bariatric surgery were analyzed. 31 (70.5%) were women and 13 (29.5%) were men. The mean age was 37.18 ? 11.26 years. The mean preoperative weight was 127.29 ? 27.10kg, decreasing to 82.00 ? 19.94kg postoperatively. With this significant reduction, the mean BMI also reduced by removing the patient's classification from obesity grade III to overweight. The mean biochemical parameters also showed significant improvement, with the exception of HDL, which showed no significant change. SIRT-1 expression had a mean value of 128.80μm2 in the intraoperative biopsies and 306.68μm2 in the biopsies performed one year after (p = 0.0069). And AMPK had mean values of 4166.10μm2 and 2718.70μm2 respectively (p = 0.0028). Conclusion: the present study showed an increased expression of SIRT-1 and a decrease in the expression of AMPK in postoperative liver biopsies. This pattern of expression accompanied improvements in weight, BMI and the profile of biomarkers that are associated with the development of co-morbidities also verified one year after surgery. / Objetivo: avaliar a express?o da prote?na quinase ativada por AMP (AMPK) e sirtuina-1 (SIRT-1) em amostras histol?gicas hep?ticas de pacientes no transoperat?rio e um ano ap?s a realiza??o da cirurgia bari?trica. Avaliar a associa??o entre a express?o dessas prote?nas com o peso, ?ndice de Massa Corporal (IMC) e par?metros bioqu?micos (glicose, colesterol total, HDL e triglicer?deos). Metodologia: a express?o das prote?nas foi verificada atrav?s de imunoistoqu?mica e quantificada por m?todo de microscopia por an?lise de imagem digital. As demais informa??es foram acessadas em banco de dados. Resultados: as bi?psias hep?ticas de 44 pacientes submetidos ? cirurgia bari?trica foram analisadas. 31 (70.5%) eram de mulheres e 13 (29.5%) eram de homens. A m?dia de idade foi de 37.18 ? 11.26 anos. A m?dia do peso pr?-operat?rio foi de 127.29 ? 27.10kg, reduzindo para 82.00 ? 19.94kg no p?s-operat?rio. Com essa redu??o significativa, o IMC m?dio tamb?m reduziu, retirando os pacientes da classifica??o de obesidade grau III para sobrepeso. As m?dias dos par?metros bioqu?micos tamb?m apresentaram melhora importante, com exce??o do HDL, que n?o teve altera??o significativa. Em rela??o ? SIRT-1 foram encontradas m?dias de 128.80μm2 nas bi?psias do transoperat?rio e de 306.68μm2 nas bi?psias realizadas um ano ap?s (p= 0.0069). J? a AMPK teve m?dias de 4166.10μm2 e 2718.70μm2, respectivamente (p=0.0028). Conclus?o: a express?o da SIRT-1 aumentou e a da AMPK diminuiu nas bi?psias hep?ticas p?s-operat?rias, ambas de forma significativa. O padr?o de express?o encontrado acompanhou os resultados de melhorias no peso, IMC e perfil de biomarcadores que s?o associados ao desenvolvimento de co-morbidades, verificados tamb?m verificados ap?s o per?odo de um ano.

Made available in DSpace on 2015-04-14T13:35:47Z (GMT). No. of bitstreams: 1 450585.PDF: 3679647 bytes, checksum: 449520a022e29d4951604d60809e331f (MD5) Previous issue date: 2013-08-09 / Attention deficit/hyperactivity disorder (ADHD) is one of the most prevalent neuropsychiatric disorders among children. Although the clinical presentation and treatment of ADHD are well established, its etiology is not yet known. Recent functional neuroimaging techniques may help increase knowledge about the pathophysiology of the disorder, allowing for the empirical testing of theoretical hypotheses on brain networks in ADHD. In this study, we analyzed a group of 23 treatment na?ve boys with ADHD, aged between 8 and 10 years old, who underwent a protocol of resting-state functional magnetic imaging before and after six months of treatment with methylphenidate. Analyses of functional connectivity were performed between regions of interest (seeds) placed within the default mode network (DMN). Subsequently, changes in the DMN connectivity before and after treatment were investigated with an independent component analysis (ICA). Results of the seeds analysis showed no significant changes in connectivity between regions of the DMN following treatment, with a relatively small increase in the anterior-posterior connectivity of the network. The ICA revealed a significant increase in the connectivity between the left putamen and the DMN (p <0.01, corrected). There was also a positive correlation between the decrease of symptoms and the connectivity between the putamen and the DMN after treatment (rho = 0.65, p = 0.17). These findings indicate that treatment with methylphenidate modifies the connectivity between the DMN and subcortical nuclei. Dysfunctions in corticalsubcortical circuits have often been associated with the pathophysiology of ADHD. The effect of treatment with methylphenidate may in part be associated with elevated dopamine levels in subcortical nuclei, modulating its connectivity with the DMN. / O transtorno de d?ficit de aten??o/hiperatividade (TDAH) ? um dos transtornos neuropsiqui?tricos mais prevalentes entre crian?as. Embora a apresenta??o cl?nica e o tratamento do TDAH estejam bem estabelecidos, suas causas ainda n?o s?o conhecidas. T?cnicas recentes de neuroimagem funcional podem ampliar o conhecimento acerca da fisiopatologia do transtorno, permitindo a testagem emp?rica de hip?teses conceituais sobre o funcionamento de redes cerebrais no TDAH. Neste trabalho, analisamos um grupo de 23 meninos com diagn?stico de TDAH, com idades entre 8 e 10 anos, virgens de tratamento, que foram submetidos a um protocolo de resson?ncia funcional de repouso antes e ap?s seis meses de tratamento com metilfenidato. Foram realizadas an?lises de conectividade funcional entre regi?es de interesse (seeds) demarcadas sobre ?reas da rede default mode network (DMN). Posteriormente, a conectividade da DMN antes e ap?s o tratamento foi mapeada atrav?s de uma an?lise de componentes independentes (ICA, independent component analysis). Os resultados da an?lise de seeds n?o demonstraram modifica??es significativas na conectividade entre regi?es da DMN com o tratamento, havendo apenas um pequeno aumento da conectividade ?ntero-posterior da rede. A an?lise por ICA revelou um aumento significativo de conectividade da DMN com o put?men esquerdo (p<0,01 corrigido). Observou-se ainda uma correla??o positiva entre a redu??o dos sintomas e a conectividade entre o put?men e a DMN ap?s o tratamento (rho=0,65, p=0,17). Esses achados indicam que o tratamento com metilfenidato modifica a conectividade entre a DMN e n?cleos subcorticais. Disfun??es em circuitos corticosubcorticais foram frequentemente associados ? fisiopatologia do TDAH. O efeito do tratamento com metilfenidato pode, em parte, estar associado ? eleva??o dos n?veis dopamin?rgicos em n?cleos subcorticais, modulando a sua conectividade com a DMN.

Made available in DSpace on 2015-04-14T13:35:48Z (GMT). No. of bitstreams: 1 451277.pdf: 1746632 bytes, checksum: e0650e210318a78c825ed4397d3cdad3 (MD5) Previous issue date: 2013-07-12 / To meet socio-cultural demands imposed by the new millennium and its implications for education, it was necessary for teachers and students to participate actively in the teaching-learning process. In this context the course evaluation by students has been a tool widely used in various medical schools in different countries. For institutions and teachers, these assessments allow to know and measure the results, further analysis of the institutional status, review projects, adjust goals, diagnose weaknesses and correct possible deviations. For the monitoring and improvement of this process, it was necessary to create instruments that aimed to evaluate the performance of the proposed changes. Due the importance of veracity and reliability that should have this information, the search for valid and reliable instruments is essential. The present study aimed to analyze the evaluation questionnaire answered by the the undergraduate students of the Faculty of Medicine PUCRS in 2011, perform validation of the questionnaire, check the level of students satisfaction in relation to the evaluated subjects and the association among the level of satisfaction, the percentage of practical activities of the discipline, student performance and the level of absenteeism. It is cross-sectional study, exploratory and contemporary, with qualitative and quantitative evaluation. About statistics, categorical variables were described by absolute and relative frequencies, and continuous variables were described by mean and standard deviation or median and interquartile range, as the results distribution. The validity of the instrument was assessed by psychometric properties of reliability and convergent validity. 1205 responses were studied, 70.6% of students reported satisfaction with the courses taken, however a significant number of students reported dissatisfaction with assessment practices and the dynamics of the course subjects. When evaluating the convergent validity it was observed that the domain dynamics of discipline showed an association between the qualitative and quantitative questions. The reliability presented Cronbach alpha=0.753and all domains had the floor effect below 20%. The ceiling effect showed above 20% in all domanies evaluated. There was a significant association between the total score obtained by the discipline and practice workload percentage above 60%. There was no association between total score and the degree obtained and absenteeism. Stimulus should be made to ongoing training group of teachers for correction of minor routes that do not allow the enchantment of the student by course and teacher for teaching. / Para atender as demandas s?cio - culturais impostas pelo novo mil?nio e suas implica??es na educa??o, foi necess?rio que professores e alunos participassem ativamente do processo ensino-aprendizagem. Neste contexto, a avalia??o do curso pelos alunos tem sido uma ferramenta utilizada em v?rias escolas m?dicas nos diversos pa?ses. Para as institui??es e para os docentes, estas avalia??es permitem conhecer e medir os resultados obtidos, aprofundar a an?lise da realidade institucional, rever projetos, diagnosticar fragilidades adequar metas e corrigir poss?veis desvios. Para acompanhamento e aprimoramento deste processo, foi necess?ria a cria??o de instrumentos que visassem avaliar o desempenho das mudan?as propostas. Pela import?ncia da veracidade e da confiabilidade que devem ter estas informa??es, a busca de instrumentos v?lidos e confi?veis ? imprescind?vel. O presente estudo teve como objetivo geral analisar o question?rio de avalia??o das disciplinas respondido pelos alunos do curso de gradua??o da Faculdade de Medicina da PUCRS no ano de 2011, realizar a valida??o do question?rio, verificar o n?vel de satisfa??o dos alunos em rela??o ?s disciplinas avaliadas do curso e a associa??o existente entre o n?vel de satisfa??o e o percentual de atividades pr?ticas da disciplina, o desempenho do aluno e o n?vel de absente?smo. Trata-se de estudo de delineamento transversal, explorat?rio e contempor?neo, com avalia??o quali-quantitativa. Quanto ao tratamento estat?stico, as vari?veis categ?ricas foram descritas por frequ?ncias absolutas e relativas e as vari?veis continuas foram descritas mediante m?dias e desvio padr?o ou mediana e intervalo interquartil, conforme a distribui??o dos resultados. A validade do instrumento foi avaliada mediante as propriedades psicom?tricas validade convergente e confiabilidade. Foram estudadas 1205 respostas; 70,6% dos alunos referiram satisfa??o em rela??o ?s disciplinas cursadas, por?m um n?mero expressivo de alunos relatou insatisfa??o em rela??o ?s pr?ticas avaliativas e ? din?mica das disciplinas do curso. Ao avaliar a validade convergente observou-se que o dom?nio din?mica da disciplina apresentou uma associa??o entre as quest?es qualitativas e as quantitativas. A confiabilidade apresentou alfa de Cronbach =0,753 e todos os dom?nios apresentaram o efeito ch?o inferior a 20%. O efeito teto apresentou valor superior a 20% em todos os dom?nios avaliados. Houve associa??o significativa entre o escore total obtido pela disciplina e percentual de carga hor?ria pr?tica acima de 60%. N?o houve associa??o do escore total e o grau obtido e absente?smo. Est?mulo dever? ser feito para capacita??o permanente do grupo de professores para corre??o de pequenas rotas que n?o permitem o encantamento do aluno pelo seu curso e do professor pelo ensino.

Made available in DSpace on 2015-04-14T13:35:48Z (GMT). No. of bitstreams: 1 451791.pdf: 11131527 bytes, checksum: 73b1d535d96f83c30dd5fb46e7f2af5d (MD5) Previous issue date: 2013-09-11 / Tuberculosis (TB) remains a leading infectious killer and its causative agent, Mycobacterium tuberculosis, infects one third of the world population. Despite 50 years of available drug treatments, TB continues to cause considerable mortality worldwide. The TBHIV co-infection and the emergence of multidrug and extensively-resistant TB have provided a very alarming challenge to global health and led us to focus on the research for new and more effective therapeutics against the disease. Advances in the identification of new TB drug targets have been driven by the availability of the genome sequence of M. tuberculosis H37Rv, and include elucidation of the role played by proteins of essential biochemical pathways for mycobacterial growth. Inosine Monophosphate Dehydrogenase (IMPDH, EC 1.1.1.205) catalyzes the penultimate and rate-limiting step in guanine nucleotide biosynthesis, the oxidation of inosine 5 -monophosphate (IMP) to xanthosine 5 - monophosphate (XMP), with concomitant reduction of nicotinamide adenine dinucleotide (NAD+) to NADH. This reaction controls the guanine nucleotide pool and IMPDH inhibitors are used as immunosuppresive, anticancer, and antiviral agents. Recently, IMPDH from M. tuberculosis (MtIMPDH) has gained attention as a promising anti-mycobacterial target, associated with a number of distinct inhibitor scaffolds. In the present work, the guaB2- encoded MtIMPDH has been cloned, expressed and purified to homogeneity. The recombinant MtIMPDH has a subunit molecular weight of 54,775 Da and Inductively-Coupled Plasma Optical Emission Spectroscopy (ICP-OES) and Flame Atomic Absorption Spectroscopy (FAAS) identified a K+ ion per subunit. Glutaraldehyde cross-linking analysis suggests that MtIMPDH predominates as a tetramer. Steady-state kinetics showed that MtIMPDH optimal activity is dependent on the presence of a monovalent cation, mainly K+. Initial velocity and product inhibition patterns suggested a steady-state ordered Bi Bi kinetic mechanism in which IMP binds first followed by NAD+, and hydride transfer is not the ratelimiting step in the MtIMPDH catalyzed reaction. In addition, NAD+ substrate inhibition implies that product release is ordered, with NADH released first. The pH-rate profile indicated one deprotonated group essential for catalysis and that groups with pK values of 7.5 and 9.0 are important for NAD+ binding. The data presented here are discussed in light of the kinetic and structural information available for IMPDHs investigated to date and should inform us on how to better design inhibitors targeting this enzyme. Additionaly, a vital part of drug development is the identification of gene products that are critical for bacterial growth and survival. In this regard, we have performed site-directed mutagenesis by gene replacement in order to evaluate the importance of the guaB2 gene for mycobacteria growth. Our results suggest that this gene is essential for M. tuberculosis H37Rv in vitro growth, and the confirmation of gene essentiality will point out the guaB2-encoded IMPDH as a potential drug target. Data resulting from enzyme s characterization and gene replacement were the starting point for MtIMPDH inhibitors planning, selection and testing. A compound was identified as promising lead molecule, with Ki values in nanomolar range and it was characterized as noncompetitive and uncompetitive inhibitor towards MtIMPDH substrates NAD+ and IMP, respectively. MtIMPDH characterization and results on primary inhibitors identification are crucial step towards a potential novel drug development for TB treatment. Altogether, these data may be useful for a better understanding about the biology of M. tuberculosis and as a framework on which to base efforts towards the development of efficient therapeutic strategies, aiming TB control. / A tuberculose (TB) permanece entre as principais causas de morte por doen?as infectocontagiosas e estima-se que um ter?o da popula??o mundial esteja infectada pelo Mycobacterium tuberculosis. Apesar de existirem tratamentos dispon?veis h? mais de 50 anos, a TB ? respons?vel por causar altas taxas de mortalidade em todo o mundo. A coinfec??o com o HIV e a emerg?ncia de TB resistente a m?ltiplas drogas representam um desafio ? sa?de p?blica e t?m estimulado a pesquisa por novos e mais efetivos agentes terap?uticos contra a doen?a. Avan?os na identifica??o de novos alvos para drogas contra a TB t?m sido poss?veis gra?as ao sequenciamento do genoma do M. tuberculosis H37Rv e incluem a elucida??o da fun??o de prote?nas envolvidas em rotas bioqu?micas essenciais para o crescimento micobacteriano. A enzima Inosina Monofosfato Desidrogenase (IMPDH, EC 1.1.1.205) ? uma enzima chave na bioss?ntese de nucleot?deos de purina, pois participa de uma etapa limitante na s?ntese de novo de nucleot?deos de guanina, a partir de inosina 5 -monofosfato (IMP). Ela catalisa a oxida??o de IMP a xantosina 5 -monofosfato (XMP), com concomitante convers?o de nicotinamida adenina dinucleot?deo (NAD+) ? sua forma reduzida, NADH. Essa rea??o controla os n?veis de nucleot?deos de guanina e inibidores da IMPDH t?m sido utilizados como agentes imunossupressores, antic?ncer e antivirais. Recentemente, a IMPDH de M. tuberculosis (MtIMPDH) foi descrita como um alvo antimicobacteriano promissor e inibidores desta enzima t?m sido reportados. Neste trabalho, foi realizada a clonagem da regi?o codificante do gene guaB2, a express?o e a purifica??o da prote?na MtIMPDH. A MtIMPDH recombinante apresentou uma massa molecular m?dia de 54.775 Da e a an?lise atrav?s de Espectroscopia de Absor??o At?mica em Chama (FAAS) e Espectroscopia de Emiss?o ?ptica com Plasma Indutivamente Acoplado (ICPOES) identificou um ?on K+ por subunidade. Os experimentos de rea??o cruzada, utilizando glutaralde?do, mostraram que a MtIMPDH pode existir como um tetr?mero em solu??o. Os estudos de cin?tica em estado estacion?rio revelaram que a atividade catal?tica da MtIMPDH ? dependente de c?tions monovalentes, principalmente K+. Os estudos de velocidade inicial e inibi??o pelos produtos sugeriram que a cat?lise procede atrav?s de um mecanismo cin?tico sequencial Bi Bi ordenado, no qual o IMP associa-se primeiramente ? MtIMPDH, seguido pela liga??o do NAD+, e que a transfer?ncia do hidreto n?o ? a etapa limitante da rea??o catalisada pela enzima em estudo. Al?m disso, a inibi??o pelo substrato NAD+ indica que a libera??o dos produtos ? ordenada, onde o NADH ? o primeiro produto a ser liberado. Os estudos de perfil de pH indicaram que a desprotona??o de um grupamento com valor de pK em torno de 8,2 ? essencial para a atividade catal?tica da MtIMPDH e, que amino?cidos cujas cadeias laterais possuem valores de pK pr?ximos de 7,5 e 9,0 est?o envolvidos na liga??o ao NAD+. Os dados aqui apresentados s?o discutidos com base em estudos cin?ticos e estruturais dispon?veis de IMPDHs de diferentes organismos e podem auxiliar no desenvolvimento de compostos inibidores para a enzima em estudo. Al?m disso, a identifica??o de genes essenciais para o crescimento bacteriano representa uma etapa importante no desenvolvimento de drogas. Assim, foi realizada a mutag?nese s?tio-dirigida por substitui??o g?nica, a fim de avaliar a import?ncia do gene guaB2 no crescimento do M. tuberculosis H37Rv. Nossos resultados sugerem que este gene ? essencial para o crescimento in vitro do M. tuberculosis H37Rv e a confirma??o da essencialidade permitir? a valida??o do produto deste gene como um alvo para inibidores. Os dados obtidos a partir da caracteriza??o cin?tica da enzima e da substitui??o g?nica representaram o ponto de partida para o planejamento, sele??o e testes de poss?veis inibidores da MtIMPDH. Entre os compostos qu?micos sintetizados, um deles apresentou valores de Ki na faixa de nanomolar e apresentou perfis de inibi??o n?o-competitivo e incompetitivo em rela??o aos substratos NAD+ e IMP, respectivamente. A caracteriza??o da MtIMPDH e os resultados preliminares de inibi??o desta enzima representam passos cruciais no desenvolvimento de novas drogas para o tratamento da TB. Assim, estes dados podem ser ?teis para um melhor entendimento sobre o metabolismo do M. tuberculosis e podem servir como base para o desenvolvimento de estrat?gias terap?uticas objetivando o controle da TB.

Made available in DSpace on 2015-04-14T13:35:49Z (GMT). No. of bitstreams: 1 452038.pdf: 5926182 bytes, checksum: 5375eb92e47f0a2a30ef9fa65dc549c5 (MD5) Previous issue date: 2013-08-08 / Objective: To evaluate the effect of platelet-rich plasma (PRP) and platelet-rich fibrin (PRF) on cutaneous wound healing in rats by comparing treated and control wounds. Material and Methods: Forty-eight Wistar rats were used in the study. Two wounds with 1-cm diameter, of total skin thickness, were excised from the back of the animals and treated with either PRP or PRF and saline (paired model). The wounds were evaluated immediately after the procedure and after 7 and 14 days using macroscopic (photographs) and microscopic (histology) methods. The photographs were used to measure the area of wounds in order to analyse wound contraction (morphometric analysis). The histological images permitted a count-analysis of collagen concentration (histomorphometric analysis) while the histological slides providet the description of the wounds characteristics. Results: Morphometric analysis revealed statistically significant time effect (P<0.001) and treatment group effect (P=0.022) over the observation period. The rate of wound contraction was higher in PRF-treated wounds than in PRP-treated wounds (P=0.030). In the histomorphometric analysis, mean collagen concentration, on the images of the experimental groups, increased significantly (P<0.001) from day 7 to day 14. There was wide variation in the percentage of collagen concentration: 129% in the PRF group vs. 22% in the PRP group; however, this difference did not achieve statistical significance (P=0.380). Histology of the wounds suggested a more accelerated healing process in the PRF group vs. the PRP group, despite the lack of statistical significance in the characteristics analyzed. PRP-treated and PRF-treated groups were not statistically superior to their controls in any of the studied variables. Conclusion: The present results suggest that treatment with PRP and PRF were not more effective than saline for cutaneous wound healing in rats. However, PRF proved to be superior to PRP. / Objetivo: Avaliar o efeito do Plasma Rico em Plaquetas (PRP) e da Fibrina Rica em Plaquetas (PRF) na cicatriza??o de feridas cut?neas em ratos, comparando os tratamentos entre si e com seus respectivos grupos controle. Material e M?todos: Foram utilizados 48 ratos Wistar. Duas feridas de 1,0 cm de di?metro, de espessura total da pele, foram excisadas do dorso dos animais e tratadas com PRP ou PRF e soro fisiol?gico (modelo emparelhado). As feridas foram avaliadas no p?s-operat?rio imediato e depois de 7 e 14 dias, de forma macrosc?pica (fotografias) e microsc?pica (histologia). As fotografias forneceram medidas da ?rea das feridas, possibilitando analisar sua contra??o (an?lise morfom?trica). As imagens histol?gicas permitiram calcular a concentra??o de col?geno nas feridas (an?lise histomorfom?trica) e as l?minas histol?gicas a descrever suas caracter?sticas. Resultados: Quanto ? an?lise morfom?trica, o efeito do tempo (P<0,001) e dos grupos (P=0,022), ao longo do per?odo de observa??o, foi estatisticamente significativo. A taxa de contra??o das feridas tratadas com PRF foi maior do que as tratadas com PRP (P=0,030). Na an?lise histomorfom?trica, a m?dia da concentra??o de col?geno, nas imagens dos grupos experimentais, apresentou um aumento do 7? para o 14? dia, considerado significativo (P<0,001). Houve grande varia??o percentual na concentra??o de col?geno: 129% para a PRF contra 22% para o PRP, por?m, sem signific?ncia estat?stica (P=0,380). A descri??o histol?gica das feridas sugeriu que o grupo PRF apresentou maior acelera??o do reparo quando comparado ao grupo PRP, apesar da aus?ncia de signific?ncia estat?stica nas caracter?sticas avaliadas. Os grupos de tratamento PRP e PRF n?o foram estatisticamente superiores aos seus grupos controle nas vari?veis estudadas. Conclus?o: Os dados do presente estudo sugerem que o PRP e a PRF n?o apresentaram resultados mais eficazes do que o soro fisiol?gico na cicatriza??o de feridas cut?neas em ratos. No entanto, a PRF mostrou-se superior ao PRP.

Made available in DSpace on 2015-04-14T13:35:49Z (GMT). No. of bitstreams: 1 452036.pdf: 1676220 bytes, checksum: 099b6754a6aea6b267cc1b8e2ae71a36 (MD5) Previous issue date: 2013-07-24 / Molecular Imaging is a technique that allows visualization, characterization and quantification of biochemical processes in a molecular and cellular level, in humans and other live organisms. Among all the technics available, it s possible to highlight PET (Positron Emission Tomography), which needs the administration of a radiotracer in the organism to be studied. Radiotracers or radiopharmaceuticals are molecules that possess a radioactive element in their composition. Nowadays, the most used PET radiopharmaceutical is fluorodeoxyglucose (18F) or [18F]FDG. This molecule is a glucose analogue that accumulates inside the cell, allowing glucose metabolism visualization. However, considering the use of glucose by the brain as its main energy source, this radiopharmaceutical accumulates in high rates in the normal brain, making difficult to see pathological processes. In order to visualize more specific alterations in the brain, many others radiotracers can be used, for example flumazenil (18F). Flumazenil (FMZ) shows high affinity for the benzodiazepine site in the GABAA receptor, acting as competitive antagonist. It s already known that GABAA receptors play a key role in neuronal excitability control, and expression deficiencies in these receptors are involved in many neurological and psychiatric disorders, such as epilepsy, anxiety, depression, schizophrenia, etc. Regarding epilepsy, it is believed that in the epileptogenic foci the GABAA receptor expression is reduced, the brain region where seizures start. For this reason, the aim of this work was to study flumazenil (18F) synthesis, as well as its purification and quality control analysis, in order to produce a molecule that can be used to localize the epileptogenic foci. The fluoride ion (18F-) was produced using PET Trace 16 MeV cyclotron from GE Healthcare, through the nuclear reaction 18O(p,n)18F, through the irradiation of enriched 18O water with accelerated protons. The flumazenil (18F) synthesis was performed in the automated synthesis module TRACERlab FX F-N from GE Healthcare, by nucleophilic aromatic substitution reaction. The established ideal reaction conditions were 145?C for 15 minutes, and 6,2mg precursor mass. The 18F- incorporation degree in the flumazenil molecule was 72?6% (n = 5), verified by reaction mixture analysis. In the last step flumazenil (18F) was purified through high performance liquid chromatography (HPLC) and reserve-phase tC18 cartridge, obtaining a product with a purity higher than 99%. The radionuclidic purity and identity were analyzed by gamma ray spectroscopy and half-life evaluation. Radiochemical purity was verified using thin layer chromatography (TLC) and HPLC. The chemical purity tested the presence of kryptofix 2.2.2, through a colorimetric test, and residual solvents (ethanol and acetonitrile) through gas chromatography (GC). The pH was verified using strips. All the results complied with the pharmacopeia using [18F]FDG as reference. Synthesis time was 80 minutes including purification steps and the overall yield was 9.3% (decay corrected). The radiopharmaceutical stability was analyzed for 8 hours, and no impurities were detected in this period. The method developed showed to be viable to produce flumazenil (18F), which can be used in pre-clinical and clinical studies in the future. The knowledge acquired with this work will allow the improvement of this technology in the country, and the research of new radiotracers for PET/CT imaging. / A Imagem Molecular ? uma t?cnica que permite a visualiza??o, caracteriza??o e quantifica??o de processos bioqu?micos a n?vel molecular e celular, em humanos e outros organismos vivos. Dentre as tecnologias dispon?veis, destaca-se o PET (Positron Emission Tomography - Tomografia por Emiss?o de P?sitrons) que necessita da administra??o do radiotra?ador ao organismo a ser estudado. Radiotra?adores ou radiof?rmacos s?o mol?culas que possuem um elemento radioativo em sua composi??o. Atualmente, o radiof?rmaco mais utilizado em PET ? o fludesoxiglicose (18F) ou [18F]FDG. Esta mol?cula ? um an?logo da glicose, que se acumula no interior da c?lula, permitindo a visualiza??o do metabolismo da glicose. Entretanto, considerando que o c?rebro utiliza basicamente a glicose como fonte de energia, este radiof?rmaco se acumula em altas taxas no c?rebro normal, dificultando a visualiza??o de processos patol?gicos. A fim de visualizar altera??es patol?gicas mais espec?ficas no c?rebro, v?rios outros radiotra?adores podem ser utilizados, como por exemplo o flumazenil (18F). O flumazenil (FMZ) apresenta alta afinidade pelo local de liga??o de benzodiazep?nicos do receptor GABAA, atuando como antagonista competitivo. Sabe-se que os receptores GABAA possuem um papel chave no controle da excitabilidade neuronal e que defici?ncias na express?o destes receptores est?o envolvidas em um grande n?mero de patologias neurol?gicas e psiqui?tricas, como epilepsia, ansiedade, depress?o, esquizofrenia e etc. Com rela??o a epilepsia, acredita-se que exista uma diminui??o da express?o dos receptores GABAA no foco epileptog?nico, regi?o do c?rebro geradora das crises epil?pticas. Desta forma, o objetivo deste trabalho foi estudar a s?ntese do flumazenil (18F), bem como sua purifica??o e as an?lises de controle de qualidade, visando produzir uma mol?cula que auxilie na localiza??o do foco epileptog?nico. O ?on fluoreto (18F-) foi produzido no c?clotron PET Trace 16 MeV da GE Healthcare, atrav?s da rea??o nuclear 18O(p,n)18F, decorrente da irradia??o da ?gua enriquecida com 18O pelos pr?tons acelerados. A s?ntese do flumazenil (18F) foi realizada no m?dulo automatizado TRACERlab FX F-N da GE Helthcare, atrav?s de uma rea??o de substitui??o nucleof?lica arom?tica. As condi??es ideais de rea??o foram estabelecidas em 145?C durante 15 minutos, sendo que a massa do precursor foi de 6,2mg. A taxa de incorpora??o do 18F- na mol?cula do flumazenil foi de 72?6% (n = 5), verificado atrav?s da an?lise da mistura de rea??o. Na fase final o flumazenil (18F) foi purificado atrav?s de cromatografia l?quida de alta efici?ncia (CLAE) e cartucho de fase-reversa tC18, obtendo-se um produto com grau de pureza superior a 99%. A pureza e identidade radionucl?dica foram avaliadas atrav?s de espectroscopia de raios gama e do tempo de meia-vida. A pureza radioqu?mica foi verificada por cromatografia em camada delgada (CCD) e CLAE. Na an?lise da pureza qu?mica verificou-se a presen?a de kryptofix 2.2.2, atrav?s de teste colorim?trico, e solventes residuais (etanol e acetonitrila) por cromatografia gasosa (CG). O pH foi analisado utilizando fitas. Os resultados obtidos foram dentro dos limites estabelecidos pela farmacop?ia levando em considera??o o [18F]FDG. O tempo total de s?ntese foi de 80 minutos e o rendimento total foi de 9,3% (corrigido pelo decaimento). A estabilidade do radiof?rmaco foi analisada durante 8 horas, sendo que nenhuma impureza foi detectada neste per?odo. O m?todo desenvolvido mostrou ser vi?vel para produ??o do flumazenil (18F), podendo este ser futuramente utilizado em estudos pr?-cl?nicos e cl?nicos. Os conhecimentos adquiridos com este trabalho permitir?o o avan?o desta tecnologia no pa?s, e a pesquisa de novos radiotra?adores para a realiza??o de exames PET/CT.

Made available in DSpace on 2015-04-14T13:35:50Z (GMT). No. of bitstreams: 1 452619.pdf: 1154303 bytes, checksum: 1c5fd674a322085ab06b90525da4401a (MD5) Previous issue date: 2013-07-22 / Introduction: Coxa brevis is a deformity of the proximal femur caused by a decrease or absence of growth of the physis of the proximal femur, usually caused by avascular necrosis present condition in conditions such as developmental dysplasia of the hip, proximal femoral epiphysiolysis, septic arthritis or disease Legg-Calv?-Perthes disease. Several surgeries have been proposed to lower the greater trochanter and femoral neck lengthening reestablishing the anatomy. Objective: To compare the mechanical tests by polyurethane in bone models the compressive strength of two different assemblies of plates for fixing osteotomy "L" of the proximal femur. Material and Methods: We used anatomical models of the proximal femur polyurethane which underwent an osteotomy longitudinal and divided into two groups with one group fixed with 4.5 DCP plate 10 holes with four screws proximal and three distal screws and fixed with a group 4.5 DCP plate 6 holes with two proximal and two distal screws. The anatomical models were tested for flexo in mechanical universal testing machine and was evaluated relative stiffness and linear movement. Results: By comparing the two groups in flexo tests, the average value of the relative stiffness in group 1 set with the long plate was 386.17 x10-3 N / m. In the group with the smaller plate fixed average value of the relative stiffness was 261,89 x10-3 N / m. The average linear displacement observed for an applied force of 1000 N in group 1 was 2.53 x10-3. In group 2, the mean value of linear displacement was 3.49 x 10-3m. Conclusion: Based on the parameters measured in the tests flexo, stability obtained by fixing the first group with a plate of ten holes was significantly higher than that obtained by fixing the second group with six holes of the plate. / Introdu??o: Coxa brevis ? uma deformidade do f?mur proximal causada pela diminui??o ou aus?ncia de crescimento da fise da extremidade proximal do f?mur, geralmente causada por necrose avascular condi??o presente nas patologias como displasia do desenvolvimento do quadril, epifisi?lise femoral proximal, artrite s?ptica ou doen?a de Legg-Calv?-Perthes. Diversas cirurgias tem sido propostas para rebaixar o grande troc?nter ou alongar o colo femoral reestabelecendo a anatomia. Objetivo: comparar atrav?s de testes mec?nicos em modelos ?sseos de poliuretano a resist?ncia ? compress?o entre duas montagens diferentes de placas para fixa??o osteotomia em L do f?mur proximal. Material e M?todos: Foram utilizados modelos anat?micos de f?mur proximal em poliuretano no qual foi realizado uma osteotomia longitudinal e divididos em dois grupos sendo um grupo fixado com placa 4,5 DCP 10 furos com quatro parafusos proximais e tr?s parafusos distais e um grupo fixado com placa 4,5 DCP 6 furos com dois parafusos proximais e dois distais. O modelos anat?micos foram submetidos a teste de flexocompress?o em m?quina universal de ensaios mec?nicos e foi avaliado a rigidez relativa e o deslocamento linear. Resultados: Ao comparar os dois grupos nos ensaios de flexocompress?o, o valor m?dio da rigidez relativa no grupo 1 fixado com a placa longa foi de 386,17 x10-3 N/m. No grupo fixado com a placa menor o valor m?dio da rigidez relativa foi de 261,89 x10-3 N/m. O valor m?dio do deslocamento linear, observado para uma for?a aplicada de 1000 N, no grupo 1 foi de 2,53 x10-3. No grupo 2 o valor m?dio do deslocamento linear foi de 3,49 x10-3m. Conclus?o: Com base nos par?metros analisados nos ensaios de flexocompress?o, a estabilidade obtida pela fixa??o do grupo 1 com placa de dez furos foi significativamente maior que a obtida pela fixa??o no grupo 2 com a placa de seis furos.

Made available in DSpace on 2015-04-14T13:35:51Z (GMT). No. of bitstreams: 1 452950.pdf: 1650025 bytes, checksum: d2d9176381fad308a3ded6adffbce4e4 (MD5) Previous issue date: 2013-07-30 / Background: Fluid overload (FO) is common in patients on PD and the combination with inadequate nutritional status causes increased mortality in peritoneal dialysis (PD) patient. Bioimpedance spectroscopy is a precise, sensitive and reliable tool for determining the fluid volume status and body composition of PD patients. The main aims of this study were: to compare body composition and volume variables, measured with the dialysis fluid inside the peritoneal cavity (CC) and after its drainage (EC), using the Body Composition Monitor (BCM); and to evaluate associations of the nutritional status and body composition of patients receiving PD, with their volume status. Methods: A cross-sectional study involving 37 stable adult patients (>18 years) on PD. A BCM report was used for the analysis of fluid status and body composition, and was conducted with both a CC and EC. Results: The study sample had the following characteristics: 62.5% female, 68.8% Caucasian, 75.0% on continuous ambulatory peritoneal dialysis (CAPD) and 25.0% on peritoneal dialysis automatized (APD). Evaluation of 32 patients showed no statistical difference between CC and EC in regard to overhydration (OH), total body water, extracellular water, intracellular water and their corrections for height and weight, lean tissue mass, fat tissue mass and their indices, adipose tissue mass, and body cell mass. Pearson's correlation coefficient of OH between CC and EC was r=0.989 (P<0.001). Bland-Altmann plot for OH full and OH empty had a line of bias of -9 mL and 95% limits of agreement from -603 to 585 ml. In another analysis with 37 patients serum albumin was higher in euvolemic (EV) patients (P=0.013), and the Subjective Global Assessment and Malnutrition Inflamation Score were higher in OH patients (P=0.002 and P=0.004, respectively). All hypertensive patients were diagnosed as OH. Several correlations were disclosed between nutritional markers and body composition markers and volemia. Conclusion: The presence of intraperitoneal fluid does not interferes with the evaluation of hydration status using BCM, or in the analysis of corporal composition - lean tissue mass, fat tissue mass, adipose tissue mass and their indices, suggesting that the BCM methodology can be applied in both conditions, with or without drainage of the dialysate solution. Besides the volume status of PD patients has a strong association with nutritional assessment variables. Consequently, a detailed evaluation of volume status should be part of a complete nutritional assessment, aiming an appropriate nutritional state. / Introdu??o: A sobrecarga h?drica (SH) ? um achado comum em pacientes em di?lise peritoneal (DP) e a combina??o com a inadequa??o do estado nutricional aumenta a mortalidade desta popula??o. A bioimped?ncia espectrosc?pica ? uma ferramenta precisa, sens?vel e confi?vel para determinar o estado vol?mico e a composi??o corporal de pacientes em DP. O principal objetivo deste estudo foi: comparar a composi??o corporal e as vari?veis de volume, medidas com o l?quido de di?lise presente na cavidade peritoneal (CC) e ap?s este ser drenado (EC), atrav?s do Body Composition Monitor (BCM); e avaliar as rela??es do estado nutricional e composi??o corporal de pacientes em DP de acordo com a volemia. M?todos: Estudo transversal, envolvendo 37 pacientes adultos (>18anos de idade) est?veis em DP. Os dados gerados pelo BCM foram utilizados para an?lise da volemia e da composi??o corporal, realizados em dois momentos, CC e EC. Resultados: As caracter?sticas da amostra em estudo foram: 62,5% mulheres, 68,8% caucasianos, 75,0% em CAPD e 25,0% em APD. A avalia??o de 32 pacientes mostrou que n?o h? diferen?a estat?stica entre CC e EC em rela??o ? hiper-hidrata??o (OH), ?gua corporal total, intra e extracelular e suas corre??es por altura e peso, massa magra, massa gorda e seus ?ndices, massa de tecido adiposo e massa celular corporal. A correla??o de Pearson entre OH-CC e OH-EC apresentou valor de r=0,989 (P<0,001). Bland-Altmann plot para OH-CC e OH-EC mostrou linha de vi?s de -9mL e 95% de limites de concord?ncia (-603 a 585 mL). Em outra an?lise com 37 pacientes a albumina s?rica foi maior em pacientes euvol?micos (EV) (P=0,013), e a Avalia??o Subjetiva Global e o Escore de Desnutri??o-Inflama??o foram maiores em pacientes hiperhidratados (P=0,002 e P=0,004, respectivamente). Todos hipertensos foram diagnosticados como hiperhidratados. Diversas correla??es foram encontradas entre marcadores de estado nutricional e composi??o corporal com volemia. Conclus?o: A presen?a de l?quido intraperitoneal n?o interfere na avalia??o do estado de hidrata??o atrav?s do uso do BCM, assim como a an?lise da composi??o corporal massa magra, massa gorda, massa de tecido adiposo e seus ?ndices sugerindo que a metodologia do BCM pode ser aplicada em ambas as condi??es, com e sem o l?quido presente na cavidade peritoneal. Al?m disso, as vari?veis de volemia estudadas apresentam forte associa??o com vari?veis de estado nutricional. Com isso, observa-se que uma avalia??o detalhada do estado de hidrata??o deve fazer parte da avalia??o nutricional criteriosa, visando determinar corretamente o estado nutricional destes pacientes.

Made available in DSpace on 2015-04-14T13:35:52Z (GMT). No. of bitstreams: 1 455924.pdf: 866376 bytes, checksum: 11973f76ada2aeeeb6a5dcd6c7c51f89 (MD5) Previous issue date: 2014-01-02 / Paraquat (PQ) is an agrochemical agent commonly used worldwide, which is allied to potential risks of intoxication. This herbicide induces the formation of reactive oxygen species (ROS) that ends up compromising various organs, particularly the lungs (by the polyamine uptake system), and the brain (by dopaminergic neuronal cell loss). This study evaluated the deleterious effects of paraquat on the central nervous system (CNS) (changes in physiologic parameters, nociceptive responses, locomotor activity and motor coordination, and expression profile of some inflammatory markers), or peripherally (inflammatory cells in the blood and lungs), with special attempts to assess the putative protective effects of the selective CXCR2 receptor antagonist SB225002 on these parameters. PQ-toxicity was induced in male Wistar rats, in a total dose of 50 mg/kg, given by the intraperitoneal (i.p.) route (10 mg/kg each three days). Control animals received saline solution (10 ml/kg) at the same schedule of administration. Separate groups of animals were treated with the selective CXCR2 antagonist SB225002 (1 or 3 mg/kg, i.p.), administered 30 min before each paraquat injection. The major changes found in paraquat-treated animals were: decreased body weight and hypothermia, nociception behavior, impairment of locomotor and gait capabilities, enhanced TNF-&#945; and IL-1&#946; expression in the striatum, and cell migration to the lungs and blood. Some of these parameters were reversed when the antagonist SB225002 was administered, including recovery of physiological parameters, decreased nociception, improvement of gait abnormalities, modulation of striatal TNF-&#945; and IL-1&#946; expression, and decrease of neutrophil migration to the lungs and blood. Taken together, our results demonstrate that damage to the central and peripheral systems elicited by paraquat can be prevented by the pharmacological inhibition of CXCR2 chemokine receptors. The experimental evidence presented herein extends the comprehension on the toxicodynamic aspects of paraquat, and opens new avenues to treat intoxication induced by this herbicide. / O paraquat (PQ) ? um composto qu?mico bastante utilizado no mundo, o qual ? aliado a potenciais riscos de intoxica??o. Este herbicida induz a forma??o de esp?cies reativas de oxig?nio (ROS) que podem comprometer v?rios ?rg?os, especialmente os pulm?es (atrav?s da recapta??o de poliaminas), e o c?rebro (pela perda de neur?nios dopamin?rgicos). Este trabalho avaliou os efeitos delet?rios do paraquat frente ao Sistema Nervoso Central (SNC) (como mudan?as nos par?metros fisiol?gicos, respostas nociceptivas, atividade locomotora e coordena??o motora, e o perfil de express?o de alguns marcadores moleculares de inflama??o), e perif?rico (como c?lulas inflamat?rias no sangue e pulm?es), com o objetivo de avaliar o poss?vel papel protetor do antagonista de receptores de quimiocinas CXCR2, o SB225002, nestes par?metros. A toxicidade ao paraquat foi induzida em ratos machos Wistar, em uma dose total de 50 mg/kg, administrada intraperitonealmente (i.p.) (10 mg/kg a cada tr?s dias). Animais do grupo controle receberam solu??o salina (10 ml/kg) no mesmo protocolo de administra??o. Diferentes grupos de animais foram tratados com o antagonista SB225002 (1 ou 3 mg/kg, i.p.), administrado 30 minutos antes de cada aplica??o do paraquat. As prinicpais mudan?as encontradas nos animais tratados com paraquat foram: diminui??o do peso corporal e hipotermia, aumento da resposta nociceptiva, diminui??o da capacidade de marcha e locomo??o, aumento da express?o de TNF-&#945; e IL-1&#946; no estriado, e a migra??o de c?lulas inflamat?rias no sangue e pulm?es. Alguns destes par?metros foram revertidos quando administrado o antagonista SB225002, como recupera??o dos par?metros fisiol?gicos, diminui??o da nocicep??o, melhora na atividade de marcha, modula??o da express?o estriatal de TNF-&#945; e IL-1&#946;, e diminui??o da migra??o neutrof?lica para o sangue e pulm?es. Em conjunto, nossos resultados demonstram que os danos causados pelo paraquat ao sistema central e perif?rico podem ser prevenidos atrav?s da inibi??o farmacol?gica dos receptores de quimiocinas CXCR2. A evid?ncia experimental aqui apresentada extende a compreens?o dos efeitos toxicodin?micos do paraquat, e proporciona novas possibilidades para tratar a intoxica??o causada por este herbicida.

Made available in DSpace on 2015-04-14T13:35:52Z (GMT). No. of bitstreams: 1 455930.pdf: 2412584 bytes, checksum: f510a7887fb6fa3e2f006d9db557d511 (MD5) Previous issue date: 2014-01-24 / Spinal voltage-gated calcium channels (VGCC) are pivotal regulators of painful and inflammatory alterations, representing attractive therapeutic targets. We examined the effects of epidural administration of the selective P/Q- and N-type VGCC blockers Tx3-3 and Ph&#945;1&#946;, respectively, isolated from the spider P. nigriventer, on symptomatic, inflammatory and functional changes allied to cyclophosphamide (CPA)-induced hemorrhagic cystitis (HC) in mice. The effects of P. nigriventer-derived toxins were compared to those displayed by MVIIC and MVIIA, extracted from the cone snail C. magus. HC was induced by a single intraperitoneal injection of CPA (300 mg/kg). The spinal blockage of P/Q-type VGCC by Tx3-3 and MVIIC, or N-type VGCC by Ph&#945;1&#946; attenuated nociceptive and inflammatory events associated with HC, including bladder oxidative stress and cytokine production. Moreover, CPA produced an evident increase of bladder TRPV1 and TRPA1 mRNA expression, which was virtually reversed by all the tested toxins. Noteworthy, Ph&#945;1&#946; strongly prevented bladder neutrophil migration, besides HC-related functional alterations. Finally, the spinal co-administration of the selective NK1 receptor antagonist CP-96345 heightened Ph&#945;1&#946; antinociceptive effects. Our results shed new lights on the role of spinal P/Q and N-type VGCC in bladder dysfunctions, pointing out Ph&#945;1&#946; as a promising alternative for treating complications associated to CPA-induced HC. / Os canais de c?lcio voltagem-dependentes (CCVDs), ao n?vel medular, s?o um dos principais reguladores das altera??es inflamat?rias e dolorosas, representando um interessante alvo terap?utico. Com isso, avaliamos os efeitos da administra??o intratecal dos bloqueadores seletivos dos CCVD do subtipo P/Q e N, Tx3-3 e Ph&#945;1&#946;, respectivamente, isolados do veneno da aranha P. nigriventer, nas mudan?as sintom?ticas, inflamat?rias e funcionais, relacionadas com a cistite hemorr?gica (CH), induzido pelo quimioter?pico ciclofosfamida (CPA) em camundongos. Os efeitos dos pept?deos provenientes da aranha P. nigriventer foram comparados com os exibidos pelas toxinas MVIIC e MVIIA, obtidas do caramujo C. magus. A CH foi induzida por uma ?nica administra??o intraperitoneal de CPA (300 mg/kg). O bloqueio medular dos CCVD do subtipo P/Q atrav?s da Tx3-3 e MVIIC ou, o bloqueio dos CCVD do subtipo N pela toxina Ph&#945;1&#946; atenuou as respostas nociceptivas e inflamat?rias associadas ? CH, incluindo estresse oxidativo e produ??o de citocinas na bexiga. Al?m disso, a CPA produziu um aumento evidente na express?o do RNAm de TRPV1 e TRPA1 na bexiga, o qual foi virtualmente revertido por todas as toxinas. Notavelmente, a isoforma Ph&#945;1&#946; reduziu a migra??o de neutr?filos para a bexiga, al?m de reverter as disfun??es da bexiga relacionadas ? CH. Finalmente, a co-administra??o medular do antagonista seletivo dos receptores NK1, CP-96345, com o pept?deo Ph&#945;1&#946;, intensificou o efeito antinociceptivo do mesmo. Nossos resultados trazem novas evid?ncias da fun??o dos CCVD do subtipo P/Q e N ao n?vel medular na disfun??o da bexiga, apontando a Ph&#945;1&#946; como uma poss?vel alternativa de tratamento para complica??es associadas ? CH, induzida por CPA.