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L'incidence des maladies cardiovasculaires (MCV) dans la population âgée et particulièrement chez les femmes postménopausées en surpoids se montre sans cesse grandissante au fil des années. Cette augmentation de la survenue des MCV serait particulièrement due à une diminution d'oestrogène pendant la ménopause, à une redistribution de la masse grasse au niveau abdominal et à une augmentation de la production des radicaux libres au cours du vieillissement. Bien que les femmes postménopausées utilisent souvent des oestrogènes de synthèse pour soulager les symptômes de la ménopause, plusieurs études prospectives ont montré que la thérapie hormonale de substitution (THS) entraînait également une diminution significative des facteurs de risque des MCV et que des effets synergiques sur l'amélioration du profil lipidique étaient aussi observés lorsque la THS était associée à un programme d'exercices aérobies. Cependant, quelques études longitudinales récemment publiées ont montré que la THS entraînait non seulement une augmentation de l'incidence de cancers, mais aussi que celle-ci augmenterait la survenue d'attaques cardiaques et de maladies coronariennes. Les phytoestrogènes semblent être une alternative intéressante à la THS. En effet, plusieurs études longitudinales et in vitro ont montré que les phytoestrogènes diminuaient l'incidence des MCV en améliorant le profil lipidique et en augmentant la résistance des lipoprotéines de faible densité (LDL) à l'oxydation. Cet effet protecteur des phytoestrogènes à l'égard des LDL semble se faire par l'amélioration de l'activité de l'enzyme paraoxonase 1 (PON1) présente à la surface des lipoprotéines de haute densité (HDL). Ainsi, le but de notre étude est d'évaluer l'effet d'une supplémentation en phytoestrogènes associés à l'exercice physique sur les marqueurs du stress oxydatif et l'activité de la PON1 chez les femmes postménopausées présentant un gain pondéral. Pour ce faire, nous avons recruté des femmes âgées entre 50 et 75 ans que nous avons aléatoirement réparties en deux groupes, 25 femmes dans le groupes phytoestrogènes et 25 femmes dans le groupe placebo. Ces sujets devaient prendre des phytoestrogènes (70 mg/jour d'isoflavones) ou le placebo pendant une période de six mois, puis les deux groupes, en plus de ces suppléments, étaient soumis à des séances d'exercices aérobies pendant six mois additionnels. Des prélèvements sanguins étaient effectués au début de l'étude, après 6 mois et à la fin de la période de suivie (12 mois). Un certain nombre de marqueurs sanguins du stress oxydatif étaient mesurés, notamment l'activité antioxydante totale (AAOT) et résiduelle (AoGap), les diènes conjugués, les dialdéhydes maloniques (MDA), mais aussi la vitamine C, la vitamine E, l'activité paraoxonase et la leptine. Nos résultats montrent une augmentation significative de la vitamine C et de l'activité paraoxonase dans les deux groupes au bout des 12 mois de suivi et également une diminution significative de la vitamine E dans les deux groupes à l'issue de la même période. Aucune variation significative n'a été observée lors du dosage des autres variables. Ces résultats suggèrent que les phytoestrogènes n'ont aucun effet sur l'ensemble des marqueurs du stress oxydatif, mais que les variations significatives obtenues lors des dosages des vitamines C et E ainsi que de l'activité paraoxonase seraient dues à une réponse adaptative de l'organisme au stress induit par l'exercice physique. L'absence d'effets des phytoestrogènes pourrait aussi s'expliquer par le haut niveau d'abandons observé tout au long de l'étude ou encore par le fait que la dose prescrite n'est pas nécessairement suffisante pour avoir une influence positive sur les différentes variables étudiées.

Atherosclerotic cardiovascular disease (ASCVD) is currently the most common cause of death in the United States. Some dietary factors contribute importantly to ASCVD and other factors can reduce risk of ASCVD. Oleic acid is a monounsaturated fatty acid (MUFA). Dietary patterns in which oleic acid contributes to a majority of dietary fatty acids are associated with reduced ASCVD risk. These beneficial effects are due to MUFA-induced lipoprotein profile changes such as decreases in low density lipoprotein (LDL) and increases in high density lipoprotein (HDL). LDL oxidation plays a central role in atherosclerosis development as it both initiates and propagates atherosclerosis. HDL is anti-atherogenic as it can attenuate LDL oxidation. HDLs are a class of diverse lipoprotein that varies in protein and enzymatic composition. The paraoxonase (PON) family of enzymes, especially PON1, is primarily expressed in the liver; PON activity in the circulatory system is associated with HDL. Both PON and HDL have been documented to be anti-atherogenic. Other factors such as homocysteine and C-reactive protein (CRP) can also be considered risk factors for ASCVD. However, studies of risk factors in healthy men who consume ground beef with a different content of MUFA are lacking; hence, no conclusive evidence has established whether consuming a high amount of MUFA in the form of ground beef alters the development of atherosclerosis. The overall purpose of this study was to investigate whether the provision of ground beef with a fractionally higher MUFA content could lower or improve several ASCVD risk factors in men who consume ground beef. These risk factors include the metabolic indices of glucose, insulin and homeostasis model assessment (HOMA), inflammation risk factors of CRP and homocysteine and anti-risk factor of paraoxonase. The concentration of homocysteine was determined spectrophotometrically following separation by high pressure liquid chromatography (HPLC). Enzyme-linked imminosorbent assay kits that measured the CRP and insulin concentration in plasma. The significance of the results was determined by subjecting the data to ANOVA using the general linear model for repeated measurement (P<0.05). From this study, it can be concluded that MUFA has a beneficial effect of lowering risks as determined by metabolic indices and lipoprotein profile. Moreover, our study showed that different concentrations of MUFA in ground beef has no effect on PON1 activity, but that increased beef consumption generally reduces PON1 in association with increases in homocysteine concentration while improving indicators of glucose tolerance.

Polychlorinated biphenyls (PCBs) are ubiquitous environmental pollutants that have been associated with various adverse health effects in humans and wildlife. Dioxin-like PCBs elicit a broad spectrum of biochemical and toxic changes including cardiovascular disorders. Paraoxonase 1 (PON1), an antioxidant enzyme, prevents oxidative stress and plays key roles in the pathogenesis of atherosclerosis. The overall goal of this dissertation is to investigate the mechanism and role of PON1 in the antioxidant defense to the exposure of 3, 3', 4, 4', 5-pentachlorobiphenyl (PCB 126), the most potent congener in PCB family. My overall hypothesis is that: The up-regulation of PON1 is an antioxidant response to PCB 126 exposure, which involves the Ah receptor (AhR) and results in changes in the PON1 protein level and activity which in turn has an influence on the oxidative stress status. First, the responses of PON1 gene expression and activities in serum and liver upon the PCB126 treatment were evaluated in the rat model. I found that PCB 126 up-regulated PON1, gene expression and activities, in a time and dose dependent manner. Next, I investigated the molecular mechanism of this response. My results show that the up-regulation of PON1 by PCB 126 involves the AhR and the XRE-like sequence in the gene promoter region. The up-regulation of PON1 was tissue specific, and this response probably protected liver and serum from lipid oxidation to some extent. The structure-activity relationship studies with PCB congeners indicate that the up-regulation of PON1 was specific to dioxin-like PCB 126. Other different AhR ligands displayed different PON1 induction capabilities. Also, reduction of PON1 activity was found in male rats dosed with non-dioxin-like PCBs. Finally I investigated the interaction, and possible protection, of other antioxidants (Se and NAC) on the response to PCB 126. I found that these antioxidants reduced the magnitude of the response of PON1 to the PCB 126 exposure in liver. Both the increase of PON1 activities and addition of antioxidants may be the reason for the lack in increase of lipid peroxidation. In total, these findings support my hypothesis and suggest that up-regulation of PON1 by PCB 126 may be an adaptive antioxidant mechanism that is involved in the body's antioxidant system.

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Submitted by Ana Maria Fiscina Sampaio (fiscina@bahia.fiocruz.br) on 2012-08-03T19:33:51Z No. of bitstreams: 1 Rogerio Jorge Barbosa Oliveira Incorporação e esterificação do colesterol na... 2012.pdf: 2467697 bytes, checksum: 40447b73cfb90f7a212555b74a8d0091 (MD5) / Made available in DSpace on 2012-08-03T19:33:51Z (GMT). No. of bitstreams: 1 Rogerio Jorge Barbosa Oliveira Incorporação e esterificação do colesterol na... 2012.pdf: 2467697 bytes, checksum: 40447b73cfb90f7a212555b74a8d0091 (MD5) Previous issue date: 2012 / Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, Bahia, Brasil / As complicações da aterosclerose agregam as principais causas de morte no mundo relacionada a problemas cardiovasculares. É caracterizada por distúrbio da condição mecânica e fisiológica, que promove espessamento e endurecimento nas ar tér i as coronárias, cerebrais e periféricos, ocasionando lesão inflamatória crônica, rica em lipídios e células características do processo inflamatório nas paredes vasculares. Objetivos: Avaliar concentrações plasmáticas de biomarcadores do metabolismo lipídico e resultados de doppler de carótidas, relacionando-os com a doença aterogênica de carótidas. Delineamento: Estudo de Corte Transverso. Casuística e Métodos: 66 pacientes com idade 57,5 ± 15,5 anos, (20 a 77), 63% mulheres. Na análise estatística utilizaram-se testes paramétricos e nãoparamétricos, valores de p < 0,05 foram considerados estatisticamente significantes. Resultados: Verificou-se DCA na população masculina a partir da faixa 36-45 anos; nas mulheres, a doença pôde ser detectada a partir de 46-55 anos, elevando-se em ambos gêneros conforme aumento da idade. Tivemos maior prevalência de DCA no gênero feminino (33% contra apenas 15%), na faixa de 56-65 anos, mostrando RR de 1,56 para o gênero feminino (p<0,002; teste de Fisher, com aproximação de Katz). Quanto ao perfil clínico, houve prevalência de 53% pacientes com DCA; 82% pacientes estavam em uso de medicação regular para quaisquer das doenças pregressas; 68% hipertensos; 73% dislipidêmicos; 11% coronariopatas; 4% apresentaram AVE e 64% faziam uso regular de hipolipemiantes. O teste de Fisher evidenciou diferentes RR em função da presença ou não de DCA: Hipotireoidismo (DCA vs NDCA, p< 0,555; RR=1,0); (DCA vs EMI, p < 0,0001; RR=5,2); Dislipidemia (DCA vs NDCA, p< 0,001; RR=1,62); HAS (DCA vs NDCA, p< 0,003; RR=1,76); DM (DCA vs NDCA, p< 0,001; RR=3,6); AVE (DCA vs NDCA, p< 0,0001; RR=2,03); DAC (DCA vs NDCA, p< 0,0001; RR=6,14). Predominaram estenoses leves em 44%, estenoses moderadas em 9%, não foram observados pacientes com estenose severa ou oclusão. A avaliação dos dados de colesterol livre e total de colesterol marcado na fração HDL diferiu entre os grupos estratificados por HDL-C maior e menor do que 40mg/dl e para as condições DCA e sem DCA (p = 0,0409 e, p = 0,0475 teste t), respectivamente. A análise de correlação linear de Pearson entre os dados de atividade de PON1 quando HDL-C < 40mg/dl e incorporação de colesterol livre e, também entre percentual de esterificação CL/CE foi significativa (r = -0,6; p = 0,048; n = 20 e r = -0,7; p = 0,006; n = 12), respectivamente. Conclusões: Evidenciado presença da DCA com aumento da idade conforme literatura; prevaleceu DCA no perfil clínico dos participantes do estudo; as doenças de base identificadas elevam o risco para DCA; o transporte reverso de colesterol é prejudicado na DCA. / Atherosclerosis is the leading cause death worldwide related from cardiovascular. Atherosclerosis is characterized by disturbance of mechanical and physiological condition, which promotes thickening and hardening of the coronary vessels, cerebral and peripheral inflammatory lesion causing chronic high-fat and abnormal cells in vascular walls. Objectives: Assess plasma concentrations of biomarkers of lipid metabolism and vascular echocardiography results of carotid artery linking them with the disease of the carotid atherogenic. Delineation: Cross-sectional observational study. Casuistry and Methods: 66 patients aged 57.5 ± 15.5 years (20-77), 63% female. Were used in the statistical analysis parametric tests and non-parametric, p-values <0.05 was considered significant. Results: It was found in the DCA from the male population age 36-45 years, in women, the disease could be detected from 46-55 years, rising in both genders as age increases. We had a higher prevalence of DCA among females (33% versus 15%), range 56-65 years, showing an RR of 1.56 for females (p <0.002, Fisher's test with approximation of Katz). As the clinical profile, there was prevalence from 53% patients with DCA, 82% patients were taking regular medication for any of the previous diseases, 68% hypertensive, 73% had dyslipidemia, and of those, 53% had evidence of DCA associated with. Coronary artery disease 11%, 4% had stroke and 64% were regularly taking lipid-lowering. The Fisher test showed different RR due to the presence or absence of DCA: The Fisher test showed different RR due to the presence or absence of DCA: Hypothyroidism (DCA vs NDCA, p <0.555, RR = 1.0), (DCA vs. EMI, p <0.0001, RR = 5.2) ; Dyslipidemia (DCA vs NDCA, p <0.001, RR = 1.62), hypertension (DCA vs NDCA, p <0.003, RR = 1.76), DM (DCA vs NDCA, p <0.001, RR = 3.6 ), stroke (DCA vs NDCA, p <0.0001, RR = 2.03), DAC (DCA vs NDCA, p <0.0001, RR = 6.14). Mild stenosis predominated in 44%, moderate stenosis in 9%. Were not seen patients with severe stenosis or occlusion. The evaluation of data from free cholesterol and total cholesterol HDL fraction differed marked on stratified groups and increased HDL-C less than 40mg/dl and conditions and without DCA (p = 0.0409 and p = 0, 0475 t-test), respectively. The analysis of Pearson's linear correlation between the data of PON1 activity when HDL-C <40mg/dl and incorporation of free cholesterol, and also between percentage of esterification CL / CE was significant (r = -0.6, p = 0.048 n = 20 and r = -0.7, p = 0.006, n = 12), respectively. Conclusions: Evidenced presence of DCA with increasing age according to the literature, DCA prevailed in the clinical profile of study participants, underlying diseases increase the risk identified for DCA, the reverse is impaired in DCA.

Introduction: Cholinesterases and Paraoxonase 1 are mediators of poisoning process by organophosphate (OP). Monitor the activities of these enzymes and know the genetic variability of the agricultural population is of great importance in the evaluation of possible risk groups for poisoning OP. Objective: This case-control study aimed to investigate the association of genetic markers (tag SNPs) in BchE and PON1 genes with characteristic symptoms of exposure to OP pesticides in rural workers.Methodology: 427 patients, both sexes, mean age 40.96 years old, divided into 226 with and 201 without characteristic symptomatology of exposure to pesticides, were genotyped for three tag SNPs, rs1803274 (BChE gene), rs662 e rs854560 (PON1 gene). Also, socio-demographic and economic parameters were analyzed. BChE activity was demonstrated, as well as the profile of workers' health was evaluated. For the test of association of the categorical variables, the Chi-square test and Fisher's exact test. For the genetic models tests the Binary Logistic Regression was used in the Additive Model and Fisher's Exact Test and Chi-SquareTest in the Dominant and Recessive Models (p < .05). Results: Associationstatistically significant for place of residence with symptoms presentation; between the rs1803274 and symptoms, in the Recessive model. Muscular weakness was the most representative symptom, presenting a statistical association with rs1803274, in the Additive and Dominant model. BChE activity was associated with rs662 in theRecessive model, demonstrating that individuals with this SNP have a higher chance of presenting reduced activity for this enzyme. Conclusion: The results demonstrate that the intrinsic factors (SNPs) and extrinsic (residence) studied can modulate theprocess of intoxication by OR, increasing or reducing the susceptibility of the individuals involved. / Introdução: As colinesterases e a Paraoxonase 1 atuam como mediadores do processo de intoxicação por organofosforado (OF). Monitorar as atividades destas enzimas e conhecer a variabilidade genética da população agrícola é de grande importância na avaliação de possíveis grupos de risco para intoxicação por OF. Objetivo: Investigar associação dos polimorfismos genéticos rs1803274 (geneBChE) e rs662 e rs854560 (gene PON1) com sintomatologia característica de exposição a agrotóxicos OF em trabalhadores rurais. Metodologia: Estudo Casocontrole com abordagem transversal. Composto por 427 pacientes, ambos os gêneros, média de idade de 40,96 (±12,6) anos, divididos em: G1-226 indivíduos com presença de sintomas característicos de intoxicação por OF e G2-201 indivíduos sem presença de sintomas; foram genotipados para 3 SNPs: rs1803274 (gene BChE), rs662 e rs854560 (gene PON1). Além disso, os parâmetros socioeconômicos, sociodemográficos, perfil de saúde e a atividade de BChE foram analisados. Para teste de associação das variáveis categóricas, foi utilizado o teste Qui-Quadrado e Exato de Fischer. Para os testes dos modelos genéticos a Regressão Logística Binária foi utilizada no Modelo Aditivo e o Teste Exato de Fisher e teste Qui-Quadrado nos Modelos Dominante e Recessivo (p<0,05). Resultados: Associação estatisticamente significante para local de residência com apresentação de sintomas; entre o rs1803274 e sintomas, no modelo Recessivo. A fraqueza muscular se mostrou o sintoma mais representativo, apresentando uma associação estatística com o rs1803274, no modelo Aditivo e Dominante. A atividade de BChE apresentou associação com o rs662, no modelo Recessivo, demonstrando que indivíduos com este SNP tem maior chance de apresentar atividade reduzida para esta enzima. Conclusão: Os resultados demonstram que os fatores intrínsecos (SNPs) e extrínsecos (local de residência) estudados podem modular o processo de intoxicação por OF, aumentando ou reduzindo a susceptibilidade dos indivíduos envolvidos. / Lagarto, SE

INTRODUÇÃO. Os genes da família paraoxonase (PON1, PON2 e PON3) apresentam grande homologia estrutural. PON1 está associada à molécula de HDL e possui funções fisiológicas, sendo a principal a de lactonase. PON1 também pode proteger as moléculas de LDL de modificações oxidativas. Embora o papel biológico mais conhecido das paraoxonases seja a prevenção da aterosclerose, elas também atuam sobre o estresse oxidativo envolvido na patogênese de outras condições como doenças inflamatórias, infecções e neoplasias. Toda a família PON parece estar implicada no desenvolvimento de linfomas. O polimorfismo L55M de PON1 foi relacionado a um maior risco para linfomas em indivíduos da população geral, enquanto PON3 e PON2 foram relacionadas à sobrevida de células tumorais. A Imunodeficiencia comum variável (ICV) é uma doença heterogênea caracterizada pela redução dos niveis de IgG, IgA e/ou IgM e da função de anticorpo. As manifestações clínicas incluem a presença de infecções recorrentes ou crônicas, doenças inflamatórias/autoimunes e incidência aumentada de malignidades como linfomas não-Hodgkin (LNH) e câncer gástrico. OBJETIVO: estudar os polimorfismos de PON1 e PON2 bem como a atividade arilesterase de PON1 e sua relação com o perfil lipídico, morbidade, mortalidade e presença de fatores de risco para linfoma LNH em pacientes com ICV. MÉTODOS/RESULTADOS: Foram avaliadas as frequências alélicas dos polimorfismos de PON1 e PON2, o perfil lipídico e a atividade arilesterase da PON1 em 63 pacientes com ICV e 130 controles saudáveis. No grupo de pacientes foi analisada a presença de fatores de risco para LNH e parâmetros de morbidade e gravidade da doença. O polimorfismo Q192R da PON1 e os polimorfismos de PON2 (S311C e A148G) não diferiram entre os grupos e não apresentaram relação com os parâmetros analisados. O genótipo 55MM e o alelo 55M foram mais frequentes no grupo ICV em relação ao grupo controle. A atividade arilesterase foi similar em pacientes e controles apresentando correlação positiva com os níveis de HDL. Pacientes com o genótipo 55MM apresentaram menor atividade de PON1 associada a maior morbidade da doença representada pela maior frequência de infecções de vias aéreas e maior taxa de internações. O genótipo 55MM também apresentou relação com a presença de fatores de risco para LNH como hiperplasia nodular linfoide (HNL) e linfonodomegalias. Por outro lado, a análise dos alelos demonstrou que a menor morbidade da doença foi associada à presença do alelo 55L, que apresentou relação com menor frequência de HNL e linfonodomegalia e menor ocorrência de óbitos. O alelo 55M apresentou relação com história familiar de imunodeficiências e neoplasias hematológicas. CONCLUSÃO: Este constitui o primeiro relato demonstrando maior frequência do genótipo 55MM e do alelo 55M em pacientes com ICV. Nossos resultados são sugestivos de que a presença do alelo 55L possa estar associado a um melhor prognóstico da doença. Inversamente, sugerem que pacientes com o genótipo 55MM apresentem maior morbidade e, possivelmente, maior risco para LNH / INTRO: The paraoxonase gene family (PON1, PON2 and PON3) has great structural homology. PON1 is associated with the HDL molecule and possess many physiological roles, the major one being of a lactonase. PON1 also protects LDL molecules against oxidative modifications. Although the best known biological role of PONs is the prevention of atherosclerosis, they also act on the oxidative stress involved in the pathogenesis of different conditions such as inflammatory diseases, infections and malignancies. The whole PON family appears to be implicated in the development of lymphomas. The L55M polymorphism of PON1 was related with a higher risk for lymphoma in the general population while PON3 and PON2 were related to survival of tumor cells. The Common Variable Immunodeficiency (ICV) is a heterogeneous disease characterized by reduced levels of IgG, IgA and/or IgM and antibody function. Clinical manifestations include the presence of chronic or recurrent infections, inflammatory/autoimmune diseases and increased incidence of malignancies such as non-Hodgkin lymphoma (NHL) and gastric cancer. OBJECTIVE: to study the PON1 and PON2 polymorphisms and the arylesterase activity of PON1 and its correlation with the lipid profile, morbidity, mortality and the presence of risk factors for NHL in CVID patients. METHODS/RESULTS: We evaluated the allele frequencies of polymorphisms of PON1 and PON2, lipid profile and arylesterase activity of PON1 in 63 patients with CVID and 130 healthy controls. In the group of patients we analyzed the presence of risk factors for NHL and parameters of morbidity and disease severity. The Q192R polymorphism of the PON1 and PON2 polymorphisms (A148G and S311C) did not differ between groups and did not correlate with the parameters analyzed. The 55MM genotype and the 55M allele were more frequent in the CVID group than in control group. The arylesterase activity was similar in patients and controls showing a positive correlation with HDL levels. Patients with genotype 55MM had lower PON1 activity, associated with increased morbidity of the disease represented by the higher frequency of respiratory infections and a higher rate of hospitalization. The 55MM genotype also was correlated with the presence of risk factors for NHL, such as lymphoid nodular hyperplasia (HNL) and lymphadenopathy. Moreover, analysis of the alleles showed that less morbidity of the disease was associated with the presence of the allele 55L, which was correlated with a lower frequency of HNL and lymphadenopathies and fewer deaths. The 55M allele was correlated with a family history of immunodeficiency and hematological malignancies. CONCLUSION: This is the first report showing a greater frequency of 55MM genotype and 55M allele in patients with CVID. Our results suggest that the presence of 55L allele may be associated with a better prognosis. Conversely, these results suggest that patients with the 55MM genotype show higher morbidity and, possibly, higher risk for NHL

Os membros da família paraoxonase (PON1, PON2 e PON3) tem sido objeto de grande interesse por prevenir o estresse oxidativo e o processo inflamatório, condições que estão em evidência em pacientes que possuem anemia falciforme. A anemia falciforme é causada por uma mutação pontual no gene ? globina que resulta em uma alteração na estrutura da molécula, gerando a HbS. Sua fisiopatologia envolve múltiplas alterações nos eritrócitos falcêmicos, hemólise, ativação de mediadores inflamatórios, disfunção das células endoteliais, episódios vaso-oclusivos e estresse oxidativo. Deste modo, o objetivo do presente estudo foi avaliar a atividade da enzima PON1, relacionando-as com os polimorfismos PON1 192 e 55, PON2 311 e 148 e PON3 10340, 2115, 45486 e 55146, bem como avaliar marcadores de inflamação e o perfil lipídico em pacientes portadores de anemia falciforme. A casuística foi composta por 43 indivíduos com anemia falciforme e 43 indivíduos saudáveis. O sangue foi coletado, para as determinações bioquímicas e determinação das atividades arilesterase e paraoxonase da PON1. O DNA foi extraído de leucócitos do sangue periférico pelo método de extração salina. A análise dos polimorfismos foi realizada por PCR/RFLP e por PCR em tempo real. A pesquisa de autoanticorpos anti-LDL oxidada foi feira por ELISA. Em relação ao polimorfismo do gene da PON1 192QR foi encontrada uma diferença significativa entre o genótipo 192RR, que apresentou maior frequencia no grupo caso (32,6%) do que no grupo controle (13,9%) (p=0,0064). A atividade arilesterase apresentou valores significativamente menores nos pacientes com anemia falciforme (p<0,001). Houve correlação positiva entre atividade arilesterase e as variáveis apolipoproteína A1(Apo A1) (p=0,0042), colesterol total (CT) (p=0,0066) e lipoproteína de alta densidade (HDL) (p=0,0283) e correlação negativa com leucócitos (p=0,04). Em relação à atividade paraoxonase foi encontrada uma correlação positiva e significativa entre a atividade da enzima e a transferrina (p=0,0094). Os títulos de anticorpos anti-oxLDL diferiram significativamente entre os grupos, grupo caso apresentando valores maiores quando comparados ao grupo controle (p<0,001). O mesmo aconteceu para a dosagem dos níveis séricos de Proteína C Reativa (p<0,001). Concentrações significantemente diminuídas de CT, lipoproteína de baixa densidade (LDL), HDL e Apo A1 e B foram observadas nos pacientes com anemia falciforme em relação ao grupo controle (p< 0,01). Conclusões: Os pacientes falciformes estão apresentaram mais estresse oxidativo que os indivíduos saudáveis, o que poderia influenciar na gravidade da doença / The members of paraoxonase family (PON1, PON2 and PON3) have been the subject of great interest by preventing oxidative stress and inflammation, conditions that are evident in patients who have sickle cell anemia. Sickle cell anemia is caused by a point mutation in the ? globin gene that results in a change in structure of the molecule, generating the HbS. Its pathophysiology involves multiple changes in sickle cell erythrocytes, hemolysis, activation of inflammatory mediators, endothelial cell dysfunction, vaso-occlusive episodes and oxidative stress. Thus, the objective of this study was to evaluate the activity of PON1 enzyme, associating them to the PON1 192 e 55, PON2 311 e 148 e PON3 10340, 2115, 45486 e 55146 polymorphisms, as well as evaluating inflammation markers and lipid profile in patients with sickle cell anemia. The casuistry has consisted of 43 individuals with SCD and 43 healthy. Blood was collected for biochemical studies and determination of arylesterase and paraoxonase activities of PON1. DNA was extracted from peripheral blood leukocytes by extraction saline. The analysis of the polymorphisms was performed by PCR / RFLP and real time PCR. The determination of autoantibodies anti-oxidized LDL was performed by ELISA. Regarding the PON1 192QR gene polymorphism was found a significant difference between genotype 192RR, with the highest frequency in the case group (32.6%) than in the control group (13.9%) (p = 0.0064). The arylesterase activity values were significantly lower in patients with sickle cell anemia (p <0.001). A positive correlation between arylesterase activity and variables apolipoprotein A1 (Apo A1) (p = 0.0042), total cholesterol (TC) (p = 0.0066) and high density lipoprotein (HDL) (p = 0.0283) and negative correlation with leukocytes (p = 0.04). Regarding paraoxonase activity was found a positive and significant correlation between the enzyme activity and transferrin (p = 0.0094). The titers of anti-oxLDL differed significantly between groups, with higher values in the case group compared to the control group (p <0.001). The same happened to the determination of serum C-reactive protein (p <0.001). Significantly decreased concentrations of TC, low density lipoprotein (LDL), HDL and Apo A1 and B were observed in patients with sickle cell disease in the control group (p <0.01). Conclusions: Patients with sickle cell disease presented more oxidative stress than healthy ones, which could influence in the severity of the disease

Submitted by Ana Maria Fiscina Sampaio (fiscina@bahia.fiocruz.br) on 2012-07-20T20:46:00Z No. of bitstreams: 1 Joelma Figueiredo Menezes Estudo de polimorfismos....pdf: 11425878 bytes, checksum: 175372593b635c3bd50b3a10af52fc30 (MD5) / Made available in DSpace on 2012-07-20T20:46:00Z (GMT). No. of bitstreams: 1 Joelma Figueiredo Menezes Estudo de polimorfismos....pdf: 11425878 bytes, checksum: 175372593b635c3bd50b3a10af52fc30 (MD5) Previous issue date: 2010 / Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, Bahia, Brasil / Indivíduos com doença falciforme (DF) apresentam estresse oxidativo (EO) constante, que é decorrente da deficiência nos sistemas antioxidantes, caracterizados pela produção de espécies reativas do oxigênio (EROS) de origem multifatorial, principalmente geradas pelo processo hemolítico. Objetivo. Investigar polimorfismos relacionados ao estresse oxidativo nos genes HFE da hemocromatose hereditária (c.282C>Y, c.63H>D e c.65S>C); haptoglobina (HP), glutationa S-transferase (GST) (GSTT1 e GSTM1) e paraoxonase (PON1) (c.55L>M e c.192Q>R), associando-os aos dados hematológicos e bioquímicos, aos níveis séricos de vitamina C, paraoxonase 1 e receptor de transferrina e histórico clínico de pacientes com doença falciforme. Casuística e Métodos. A casuística foi composta por 153 indivíduos com DF (HbSS e HbSC) e 196 crianças saudáveis. O perfil de hemoglobinas foi determinado por HPLC. Os polimorfismos nos genes HFE, HP, GST, PON1, a presença da talassemia alfa 3,7Kb e 4,2Kb e dos haplótipos ligados ao grupo de genes da globina beta foram investigados por PCR e PCR-RFLP. A avaliação do perfil lipídico, hepático, função renal, inflamatório e metabolismo do ferro foram realizadas por técnicas espectrofotométricas. Os níveis séricos de vitamina C, receptor de transferrina solúvel e hemeoxigenase 1 total foram detectados por ELISA. Resultados. O estudo de polimorfismos no gene da GST demostrou que o genótipo Mu nulo foi o mais frequente em ambos os grupos estudados; a análise hematológica , nos indivíduos HbSS revelou diferença significante para os parâmetros de leucócitos (p=0,0452), de segmentados neutrófilos (p=0,0224) e o evento de STA (p=0,0423); nos indivíduos com DF foram encontradas diferenças para a uréia (p=0,0288) e ferritina (p=0,0316). No estudo do gene da Haptoglobina nos indivíduos com DF encontrou-se diferença significativa para contagem de reticulócitos (p=0,0167), ferritina sérica (p=0,0493); contagem de linfócitos típicos (p=0.0422) e LDH (p=0,0181); nos HbSS encontrou-se diferença para creatinina (p=0,0178) e ALT (p=0,0127). Para o gene HFE foi encontrada diferença significativa entre os alelos selvagem e mutante para o polimorfismo c.63H>D e contagem de plaquetas (p=0,0311) em indivíduos com DF; para o polimorfismo c.65S>C observou-se diferenças estatisticamente significativas para hemoglobina (p=0,0044) e hematócrito (p=0,0078). Nos HbSC para o polimorfismo c.63H>D observou-se entre os alelos selvagem e mutante diferenças significativas com o VCM (p=0,0032), HCM (p=0,0010) e linfócitos típicos (p=0,0242). Entre os indivíduos HbSS observou-se diferença significativa entre os alelos estudados e plaquetas (p=0,0078). Para a mutação c.65S>C observou-se diferenças significativas para os parâmetros de hemoglobina (p=0,0265) e hematócrito (p=0,0407) nos indivíduos HbSS e hemoglobina (p=0,0051) e hematócrito (p=0,0060) nos indivíduos HbSC. A análise do histórico clínico dos indivíduos com DF e o polimorfismo c.63H>D demonstrou diferença significativa para esplenomegalia, OR=3,38 (0,93 -12,22) e p=0,0402. Nos indivíduos HbSS foi encontrada diferença significativa entre os alelos c.63H>D e infecção, com OR=0,29 (0.07 -1,18) e p=0,0455. Para o gene PON1, a avaliação da atividade da PON1 nos indivíduos DF entre os alelos selvagem e mutante revelou diferença estatisticamente significativa, tendo o alelo selvagem maior atividade da PON1 que alelo mutante; para o polimorfismo PON1c.192Q>R observou-se entre os alelos selvagem e mutante diferenças significativas para as variáveis bioquímicas VLDL-C (p=0,0267) e triglicérides (p=0,0127). Nos HbSC verificou-se diferença estatística significativa para o PON1c.192Q>R e concentração de hemoglobina (p=0,0459), hematócrito (p=0,0225) e contagem de linfócitos típicos (p=0,0364). Para o PON1c.55L>M observou-se diferença significativa com a idade (p=0,0139), plaquetas (p=0,0109), creatinina (p=0,0329) e PCR (p=0,0141). Observou-se associação entre atividade da PON1 e esplenectomia (p=0,001); para hemeoxigenase e ferro sérico (p=0,023); e para vitamina C observamos correlação com colesterol HDL (p=0,037). Há correlação entre glutationa e vitamina C (p=0,035).Conclusões: Os polimorfismos estudados podem estar agindo sinergicamente com a hemoglobina variante S e assim influenciar na gravidade da doença, necessitando de outros estudos para validar estes resultados, uma vez que algumas destas investigações foram realizadas pela primeira vez neste grupo de indivíduos. / Patients with sickle cell disease (SCD) has continued oxidative stress (OS), which is resulting from ineffective antioxidant systems, characterized by production of reactive oxygen species (ROS) of multifactorial origin, mainly generated by the hemolytic process. Objective: To investigate polymorphism related to oxidative stress in hereditary hemochromatosis HFE gene (c.282C>Y, c.63H>D c.65S and D>C), haptoglobin (HP), glutathione S-transferase (GST) (GSTT1 and GSTM1) and paraoxonase (PON1) (c.55L>M and c.192Q>R), in association with the hematological and biochemical data, serum levels of vitamin C, paraoxonase 1, transferrin receptor and clinical manifestations of SCD patients. Methods. The sample included 153 individuals with SCD (HbSS and HbSC) and 196 healthy children. The profile of hemoglobin was determined by HPLC. The HFE gene polymorphisms, HP, GST, PON1, the presence of alpha thalassemia 3.7 Kb, 4.2 Kb and haplotypes of the beta gene cluster were investigated by PCR and PCR-RFLP. The lipid profile, liver, renal function, inflammation and iron metabolism was performed by spectrophotometric techniques. Serum levels of vitamin C, soluble transferrin receptor and total hemeoxigenase were investigated by ELISA. Results: The GST Mu null genotype was more frequent in both groups and both polymorphisms were in Hardy-Weinberg equilibrium. In the HbSS observed significant differences in the parameters of leukocytes (p = 0.0452) and segmented neutrophils (p = 0.0224) and event STA (p = 0.0423). Individuals with SCD had differences for urea (p = 0.0288) and ferritin (p = 0.0316). Evaluating the Haptoglobin gene in individuals with SCD found a significant difference for reticulocyte count (p = 0.0167), serum ferritin (p = 0.0493); typical lymphocyte count (p = 0.0422) and LDH (p = 0.0181); in the HbSS were found differences for creatinine (p = 0.0178) and ALT (p = 0.0127). For the HFE gene was found significant differences were observed between wild and mutant alleles for the polymorphism c.63H> D and platelet count (p = 0.0311) in individuals with SCD; for polymorphism c.65S> C were differences were statistically significant for hemoglobin (p = 0.0044) and hematocrit (p = 0.0078). In the HBSC observed significant differences in MCV (p = 0.0032), MCH (p= 0.0010) and typical lymphocytes (p = 0.0242). Among subjects HbSS were observed significant differences between alleles and platelets (p = 0.0078). For mutation c.65S> C showed significant differences in the parameters of hemoglobin (p=0.0265) and hematocrit (p=0.0407) in HbSS individuals, and hemoglobin (p=0.0051) and hematocrit (p=0.0060) in subjects HbSC. The analysis of the clinical history of individuals with SCD and polymorphism c.63H> D there was a significant difference for splenomegaly, OR = 3.38 (0.93 -12.22) and p = 0.0402. In HbSS individuals significant difference was found between alleles c.63H> D and infection, with OR = 0.29 (0.07 -1.18) p = 0.0455. For the PON1 gene, The activity assessment of PON1 in the individuals DF with wild and mutant genotypes showed statistically significant differences, the wild-type allele showed higher PON1 activity than the mutant allele; for the biochemical parameters to polymorphism PON1c.192Q> R and the values of VLDL-C (p = 0.0267) and triglycerides (p = 0.0127). In HbSC there was a statistically significant difference for PON1c.192Q> R and hemoglobin (p = 0.0459), hematocrit (p = 0.0225) and typical lymphocyte count (p = 0.0364). For PON1c.55L> M there was significant difference with age (p = 0.0139), platelets (p = 0.0109), creatinine (p = 0.0329) and CRP (p= 0.0141). There was an association between PON1 activity and splenectomy (p = 0.001) for hemeoxigenase and serum iron (p = 0.023), and vitamin C showed a correlation with HDL cholesterol (p = 0.037). There is a correlation between glutathione and vitamin C (p=0.035) Conclusions: The polymorphisms could be acting synergistically with this hemoglobin variant S, and influence the severity of disease, have been necessary further studies to validate these results, since some of these investigations were performed for the first time in this group of individuals.