Submitted by Erika Demachki (erikademachki@gmail.com) on 2015-02-03T15:39:00Z No. of bitstreams: 2 Dissertação - Tamiris Augusto Marinho - 2013.pdf: 1742779 bytes, checksum: bdad0a86938bf149e3128fa15a41e2c9 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Approved for entry into archive by Erika Demachki (erikademachki@gmail.com) on 2015-02-03T15:39:48Z (GMT) No. of bitstreams: 2 Dissertação - Tamiris Augusto Marinho - 2013.pdf: 1742779 bytes, checksum: bdad0a86938bf149e3128fa15a41e2c9 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Made available in DSpace on 2015-02-03T15:39:48Z (GMT). No. of bitstreams: 2 Dissertação - Tamiris Augusto Marinho - 2013.pdf: 1742779 bytes, checksum: bdad0a86938bf149e3128fa15a41e2c9 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2013-01-11 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Hepatitis B virus (HBV) remains a major cause of liver disease worldwide despite vaccination programs implemented over the last decade. Worldwide, it is estimated that 2 billion people are infected with HBV and that more than 240 million are chronically infected. Patients with chronic hepatitis B are at risk for developing liver cirrhosis and hepatocellular carcinoma. Recyclable waste collectors have a lifestyle that is characterized by unfavorable social and environmental factors. There is currently very little data on HBV infection in this population. Therefore, the aim of the present study was to investigate the epidemiological profile of HBV infection in a population of recyclable waste collectors in Goiânia-GO. A cross-sectional survey was carried out with 431 individuals who were recruited in all 15 recycling cooperatives in Goiânia-GO. All individuals were interviewed, and their serum samples were tested for the presence of HBV serological markers. HBsAg and anti-HBc-positive samples were tested for HBV-DNA by polymerase chain reaction, and were genotyped by sequencing of the S region. The overall HBV prevalence infection was 12.8% (95% CI: 9.8-16.2). A multivariate analysis of risk factors showed that age >40 years and illicit drug use were independently associated with HBV infection. HBV-DNA was detected in 2/3 HBsAg-positive samples, and in 1/52 anti-HBc-reactive samples, resulting in an occult HBV infection rate of 1.9%. HBV genotypes A (subgenotype A1), D (subgenotype D3) and F (subgenotype F2) were identified. Only 12.3% of this population showed serological evidence of previous hepatitis B vaccination. These findings highlight the need of public health programs to recyclable waste collectors, including the hepatitis B vaccination. / O vírus da hepatite B (HBV) continua a ser uma das principais causas de doença hepática em todo o mundo, apesar dos programas de vacinação implementados ao longo da última década. Estima-se que 2 bilhões de indivíduos já foram expostos ao HBV e mais de 240 milhões estão cronicamente infectados em todo mundo. Os indivíduos com hepatite B crônica apresentam risco elevado para o desenvolvimento de cirrose hepática e carcinoma hepatocelular. Catadores de materiais recicláveis vivem em condições sociais e ambientais precárias. Atualmente, existem poucos dados sobre a infecção pelo HBV nesta população. Portanto, o objetivo geral do presente estudo foi investigar o perfil epidemiológico da infecção pelo HBV em uma população de catadores de materiais recicláveis em GoiâniaGO. Estudo transversal realizado com 431 indivíduos recrutados nas 15 cooperativas de reciclagem em Goiânia-GO. Todos os participantes foram entrevistados e suas amostras de soro testadas para os marcadores sorológicos do HBV. As amostras HBsAg e anti-HBc positivas foram submetidas à detecção do HBV-DNA pela reação em cadeia da polimerase, e genotipadas por sequenciamento da região S. A prevalência global da infecção por HBV foi de 12,8% (IC 95%: 9,8-16,2). A análise multivariada dos fatores de risco mostrou que idade superior a 40 anos e uso de drogas ilícitas foram independentemente associados à infecção pelo HBV. O DNA viral foi detectado em 2/3 amostras HBsAg positivas e em 1/52 amostras anti-HBc reagentes, resultando, assim, em um índice de infecção oculta pelo HBV de 1,9%. Os genótipos A (subgenótipo A1), D (subgenótipo D3) e F (subgenótipo F2) foram identificados. Apenas 12,3% da população mostrou evidência sorológica de vacinação prévia contra hepatite B. Estes achados evidenciam a necessidade de programas de saúde pública voltados para os catadores de materiais recicláveis, incluindo a vacinação contra hepatite B.

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nÃo hà / A coinfecÃÃo entre o VÃrus da ImunodeficiÃncia Humana (HIV) e o VÃrus da Hepatite B (HBV) possui os mesmos fatores de transmissÃo e como consequÃncia os fatores de risco associados, explicam a alta prevalÃncia destes agentes infecciosos no nosso meio. O presente estudo estimou a prevalÃncia da coinfecÃÃo HIV e HBV e descreveu as caracterÃsticas individuais que agem como fatores de risco para aquisiÃÃo desta coinfecÃÃo, com o intuito de utilizar esta informaÃÃo para o aconselhamento. A amostra utilizada foi composta pelos 805 pacientes infectados com o HIV no estado do Piauà que buscaram o LACEN-PI para monitoramento da carga viral e dos linfÃcitos T CD4+. A prevalÃncia da hepatite B (HB), utilizando o marcador anti-HBc total, foi de 29,3% e, para o HBsAg este valor ficou em 2,5%. A prevalÃncia do Anti-HBc total foi 38,3% na faixa acima dos 40 anos, 38,6% para o sexo masculino, 31,9% entre os solteiros, 47,7% entre os aposentados, 50,7% entre os que relataram antecedente de icterÃcia, 54% entre os que tiveram hepatite com diagnÃstico mÃdico, 40,7% entre os com passagem por reformatÃrio ou prisÃo, 38,1% entre usuÃrios de droga nÃo endovenosa, 35,7% entre os com contato sexual com usuÃrio de droga ilÃcita, 48,8% entre os com preferÃncia homossexual/bissexual, 44,9% entre os que disseram ter contato sexual raro com prostituta, 37,1% entre os que tiveram DST e 31,4% para os com carga viral abaixo de 10.000 cÃpias/mL de sangue. Foram observadas significÃncias estatÃsticas entre as variÃveis e a frequÃncia de positividade do anti-HBc total. As informaÃÃes deste trabalho poderÃo ser utilizadas no combate, aconselhamento e prevenÃÃo do avanÃo, do nÃmero de casos HB em pacientes HIV positivos. / The coinfection between the Human Immunodeficiency Virus (HIV) and Hepatitis B Virus (HBV) has the same transmission factors and consequently the associated risk factors explain the high prevalence of these infectious agents in our midst. This study estimated the prevalence of HIV and HBV coinfection and described the individual characteristics that act as risk factors for acquisition of coinfection, in order to use this information for advice. The sample was composed of 805 patients infected with HIV in the state of Piauà who sought the LACEN-PI for monitoring viral load and CD4 + T lymphocytes. The prevalence of Hepatitis B (HB), using the marker anti-HBc, and was 29.3% for HBsAg this value was 2.5%. The prevalence of Anti-HBc was 38.3% aged over 40 years, 38.6% for males, 31.9% among unmarried, 47.7% among retirees, 50.7% among who reported a history of jaundice, 54% among those who were diagnosed with hepatitis, 40.7% among those passing through reformatory or prison, 38.1% among non-intravenous drug users, 35.7% with sexual contact with illicit drug users, 48.8% with a preference among homosexual / bisexual, 44.9% among those who reported having sexual contact with a prostitute rare, 37.1% among those who had STD and 31.4% for those with viral load below 10,000 copies / mL of blood. We observed statistical significance between variables and the frequency of positive anti-HBc. The information in this work could be used in combat, counseling and prevention of advancement, the number of HB cases in HIV positive patients.

This thesis describes the study of the three-dimensional structure and dynamics of the hepatitis B virus (HBV) encapsidation signal, epsilon, by means of nuclear magnetic resonance (NMR) and mutational data. HBV replicates by reverse transcription of an RNA pregenome into the viral DNA genome, which becomes enclosed in viral particles (encapsidation). Epsilon is a stem-loop structure within the RNA pregenome and both the primary sequence and secondary structure of epsilon are strongly conserved, in agreement with its essential function of propagating HBV. Epsilon is therefore a potential target for drug design. Studying the structure of epsilon requires development of new methods in the field of structural biology, as it is such a large RNA. Knowing the structure of epsilon will help to better understand the encapsidation mechanism and priming step of reverse transcription. This will help us in the search for antiviral drugs that block epsilon and prevent the viral reverse transcriptase from binding. NMR spectroscopy is a method that provides detailed structural and dynamical data in solution under natural conditions. However, the size of the molecules that can be studied with NMR is limited. NMR spectra become more and more difficult to interpret as the size of the molecule increases. To circumvent this problem, large RNA molecules can be divided into smaller parts and only the parts essential for NMR studies are selected. The information obtained from these smaller fragments can then be used to determine the structure of the larger molecule. Furthermore, a new method of enzymatically synthesizing nucleoside triphosphates with isotopes suitable for NMR has made it possible to specifically label the RNA molecules. Using this method it is possible to derive highly detailed molecular structures of RNA up to a size of 150 nucleotides. The method of selective isotope labelling was applied to different parts of HBV epsilon. Three RNA fragments of 27 (apical loop), 36 (internal bulge) and 61 (whole epsilon) nucleotides (nt) were synthesized in the unlabelled form. The 27-nt and 36-nt RNAs were also synthesized with (13C, 15N, 1', 3', 4', 5', 5"-2H5)-labelled uridines. The 61-nt sequence was (13C, 15N)-guanidine labelled. This labelling allowed unambiguous assignment of otherwise inaccessible parameters. The unlabelled and labelled RNA sequences provided the necessary data for structure derivation of the whole epsilon. The apical loop of epsilon forms a pseudo-triloop motif. There is only one conformation of the loop that fulfils all the restraints, including experimental chemical shifts. However, the loop adopts several structures that fulfil the experimental distance, torsion angle and residual dipolar coupling restraints. This may reflect true flexibility. Indeed, relaxation studies on the unlabelled and labelled 27-nt sequences show that the residues that show multiple conformations are flexible. This can be an important feature for the recognition and subsequent binding of epsilon to the viral polymerase. The information gained on the HBV encapsidation signal is useful in our understanding of the initiation of replication of the virus. This can in turn contribute to the search for drugs against HBV.

bstract Cancer has been the first lethal factor in Taiwan, and hepatitis B is also a serious problem in our country. We have identified that the extracts of Taiwanese Schisandraceous plants, including Schizandra arisanensis, Kadsura matsudai and K. japonica, have inhibitory effect on type B hepatitis surface antigen (HBsAg) and e-antigen (HBeAg). Besides, we also find that the extracts of Echinops grijsii can inhibit the growth of human cancer cell line, KB and Hela. To purify active compounds by using column chromatography and high-performance liquid chromatography, we have furnished fifty-eight compounds from above Schisandraceous plants, including thirty-seven C18 lignans composed of [6.8.6]-dibenzocyclooctadiene skeleton, nine C19 homolignans, four triterpenes, three steroids and five cyclic-aromatic compounds. In the case of Echinops grijsii, we got thirteen major compounds containing eight thiophenes, four triterpenes, and a sterol. Structural elucidation of the novel homolignan and ten lignans are based on the spectral and chemical analyses, mainly by using two-dimensional nuclear magnetic resonance (NMR) of 1H and 13C nucleus. From the anti-HBsAg and anti-HBeAg assay, we found that four of new C18 lignans and one known (+)-gomisin K3 (33) lignan exhibited the significant inhibitory effects on surface antigen of hepatitis B virus. Moreover, to modify the known lignans, kadsurarin with different halogens and functional group having nitrogen atom, as well as (+)-gomisin K3 with several kinds of phenyl compounds and sulfur functional groups are processed. The preliminary structures and bioactivity relationships (SAR) studies demonstrated that (+)-gomisin K3 with sulfuric functional group could decrease cytotoxicity and increase inhibitory effect on surface antigen and e-antigen of hepatitis B virus. In the part of studying synthesis by using inactive 5¡¦-(but-3-en-1-ynyl)-2,2¡¦ -bithiophene (1) from Echinops grijsii, the yne-ene moiety of its structure was hydrolyzed into carbonyl group and then was attached a serious of various carbon number of hydroxy groups. Besides the above experiment, £\-trithienyl (1) was served as a substrate and made it into unsaturated carbonyl group, which then was linked with a series of hydroxy groups. Cytotoxicity datum showed that the derivatives of bi-thiophene have better anticancer activity than derivatives of tri-thiophene. And the assay results also exhibited that the bi-thiophene derivatives with hydroxy group with less than three carbon numbers have better inhibitory activity against cancer cells.

Submitted by Hiolanda Rêgo (hiolandar@gmail.com) on 2013-06-10T18:28:37Z No. of bitstreams: 1 Dissertação_ICS_Iury Souza.pdf: 1047267 bytes, checksum: 06bc8acdb629b7e8ae63feab201995ad (MD5) / Made available in DSpace on 2013-06-10T18:28:37Z (GMT). No. of bitstreams: 1 Dissertação_ICS_Iury Souza.pdf: 1047267 bytes, checksum: 06bc8acdb629b7e8ae63feab201995ad (MD5) / CAPES / Cerca de 33,3 milhões de pessoas apresentam infecção pelo Human Immunodeficiency Virus (HIV) no mundo; 180 milhões estão infectados pelo Hepatitis C Virus HCV e estima-se que 360 milhões apresentem infecção ativa pelo Hepatitis B Virus (HBV). Outra realidade mundial é a co-infecção entre esses vírus. Os dados mostram a importância global dessas viroses e a urgência do desenvolvimento de novos ensaios de diagnóstico sensíveis, específicos, rápidos e de baixo custo, que possam atender à demanda de entidades públicas inseridas em programas para prevenção e diagnostico dessas doenças. O presente trabalho consiste em validação relativa de um novo teste imunocromatográfico desenvolvido pela empresa canadense Medmira para detecção de anticorpos específicos contra HIV, HCV e HBV. Os resultados encontrados foram extremamente favoráveis para a detecção de anticorpos específicos para HIV, apresentando 98,6% de sensibilidade e 100% de especificidade. Para o anti-HBV a sensibilidade e especificidade encontradas foram de 90,0% e 98,6%, e de 86,3% e 100%, para anti-HCV, respectivamente. Nenhuma reatividade cruzada foi encontrada e a reprodutibilidade e repetitividade foram de 100%. O índice kappa e a acurácia global do teste foram de 0,91 (0,88-0,94) e 95,5% (93,5-97,5), respectivamente. Conclui-se que o ensaio imunocromatográfico é clinicamente útil em triagens rápidas para detecção de anticorpos anti-HIV, HCV e HBV. / Salvador

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2017-08-04T14:12:52Z No. of bitstreams: 2 license_rdf: 811 bytes, checksum: e39d27027a6cc9cb039ad269a5db8e34 (MD5) Dissertação - Tássia Brena da Costa.pdf: 2477280 bytes, checksum: c34cd8503224fb8886f2d07b5e20b69a (MD5) / Made available in DSpace on 2017-08-04T14:12:52Z (GMT). No. of bitstreams: 2 license_rdf: 811 bytes, checksum: e39d27027a6cc9cb039ad269a5db8e34 (MD5) Dissertação - Tássia Brena da Costa.pdf: 2477280 bytes, checksum: c34cd8503224fb8886f2d07b5e20b69a (MD5) Previous issue date: 2016-03-29 / CNPQ / FACEPE / A metabonômica pode ser definida como um conjunto de ferramentas, analíticas e de estatística multivariada, utilizadas para identificar mudanças de concentração dos metabólitos em um dado biofluido, associando-as à perturbação sofrida pelo organismo. Sendo assim, ela seria capaz de identificar qualquer doença no organismo, desde que seja empregado o biofluido adequado e as informações sejam corretamente extraídas. Para isso, a ferramenta mais empregada é a Espectroscopia de Ressonância Magnética Nuclear de Hidrogênio-1 (RMN de ¹H), e é necessário o uso de técnicas quimiométricas para extrair as informações do espectro. Neste trabalho, foram construídos modelos metabonômicos para: (1) identificar pacientes portadores de esteatose, e dos vírus da hepatite B (HBV) e da hepatite C (HCV), utilizando amostras de urina; e (2) classificar o grau de fibrose hepática em pacientes com hepatites crônicas por HBV ou HCV, utilizando amostras de soro sanguíneo. O modelo para classificação de pacientes com esteatose, obteve 100% de sensibilidade e de valor preditivo positivo. Para identificar esteatose independentemente de ser um portador de HBV ou HCV, o modelo construído obteve 97,9% de exatidão. Para classificar portadores de HBV e HCV, os modelos apresentaram sensibilidade de 100% e 92,6%, respectivamente. O modelo construído para diferenciar pacientes com diferentes lesões no fígado: esteaose e hepatites virais B ou C, obteve 94% de exatidão. Para classificar pacientes com fibrose significativa; fibrose avançada; e cirrose, alcançamos 98,4; 100; e 96,8% de exatidão, respectivamente. Através da combinação dos resultados dos modelos de fibrose significativa e fibrose avançada, foi possível determinar os pacientes com grau F2, no METAVIR, com percentual de acerto de 96,8%. Nas análises de fibrose, a exatidão observada para os modelos metabonômicos foram superiores aos observados para os métodos não-invasivos normalmente utilizados na prática clínica, APRI (do inglês, Aspartate aminotransferase Platelet Ratio Index) e FIB-4. A estratégia metabonômica demonstrou capacidade de avaliar a presença de diferentes doenças hepáticas em uma única análise, não invasiva, e determinar o grau de fibrose hepática, de forma minimamente invasiva. / The metabonomics can be defined as a set of analytics and multivariate statistics tools, used to identify the metabolite concentration changes in a certain biofluid, associating them to the disturbance suffered. Therefore, it would be able to identify any disease in the body, if employed the appropriate biofluid and correctly extract the information. The most commonly used tool is Nuclear Magnetic Resonance Spectroscopy for hydrogen-1 (¹H NMR), and chemometrics techniques are used to extract the information of the spectrum. In this work we built metabonomics models to: (1) identify patients with steatosis, hepatitis B (HBV) and hepatitis C (HCV), using urine samples; and (2) classify the degree of liver fibrosis in patients with chronic hepatitis, HBV or HCV, using blood serum samples. The classification model for patients with steatosis obtained 100% to sensitivity and positive predictive value. To identify steatosis, without regard the presence of HBV or HCV, the constructed model achieved 97.9% accuracy. To classify carriers of HBV and HCV, the models showed 100 and 92.6% of sensitivity, respectively. The constructed model to differentiate patients with different liver damage: steatosis and viral hepatitis B or C, achieved 94% accuracy. To classify patients with significant fibrosis; advanced fibrosis; and cirrhosis, the models reached 98.4; 100; and 96.8% accuracy, respectively. By combining the results of significant fibrosis models and advanced fibrosis, it determined the patients with F2 in the METAVIR, with 96.8% of accuracy. In fibrosis analysis, the accuracy observed for metabonomics models were higher than those observed for the non-invasive methods commonly used in clinical practice, APRI (Aspartate aminotransferase Platelet Ratio Index) and FIB-4. The metabonomics strategy demonstrated ability to assess the presence of different liver diseases in a single non-invasive analysis and determine the degree of liver fibrosis, in a minimally invasive way.

Indiana University-Purdue University Indianapolis (IUPUI) / The function(s) of the intracellular form of HBeAg, previously reported as the preCore protein intermediate (p22) without the N-terminal signal peptide, remains elusive. Here, we propose to elucidate the translocation of p22 during its formation from endoplasmic reticulum (ER) to cytosol, how it differs from core in its inability to form a capsid and the biological functions of cytoplasmic and nuclear p22. Firstly, we have identified that a portion of p22, after the cleavage of its signal peptide in ER, is released back into the cytosol through an ERAD-independent mechanism, as neither wildtype nor dominant-negative p97 affected the ER-to-cytosol translocation of p22 or ER-Golgi secretion of HBeAg. Secondly, despite sharing the same sequence with core protein except for the extended 10 amino acid precore region at the N-terminus, we observed that p22 wildtype and C-7Q mutant are unable to form a capsid. Thirdly, we report that p22 but not the secreted HBeAg significantly reduced interferon stimulated response element (ISRE) activity and expression of interferon stimulated genes (ISGs) upon interferon-alpha (IFN- α) stimulation. Furthermore, in line with this, RNA-seq analysis of ISG induction profile from IFN-α treated patients showed that HBeAg(+) patients exhibited reduced and weak antiviral ISG upregulations compared to HBeAg(-) patients. Further, mechanistic study indicated that while p22 did not alter the total STAT1 or p-STAT1 levels in IFN-α treated cells, it blocked the nuclear translocation of p-STAT1 by interacting with karyopherin α1, indicating that the cytoplasmic p22 may impede JAK-STAT signaling to help the virus evade host innate immune response and cause resistance to IFN therapy in patients. Additionally, nuclear p22 and nuclear core were found to interact with the promoter regions (ISRE – containing) of ISGs, suggesting a new mechanism of inhibition of ISG expression upon stimulation. Finally, we found that the nuclear p22 can bind to cccDNA minichromosome and affects cccDNA maintenance and/or transcription. Thus, our results indicate that there is a novel ER sorting mechanism for the distribution of the intracellular and secretory HBeAg, and the intracellular HBeAg may contribute to HBV persistence by interfering with IFN-α elicited JAK-STAT signaling and regulating cccDNA metabolism.

Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Bc. Eva Davidovová Supervisor: prof. PharmDr. Petr Pávek, Ph.D. Title of diploma thesis: Viral hepatitis: novel insights and novel therapeutic interventions. Viral hepatitis is a well-known worldwide problem. Hepatitis B virus (HBV) and hepatitis C virus (HCV) are characterized by the development of serious complications, especially with regard to the transition to the chronic stage of the disease, associated with the development of fibrosis, cirrhosis and hepatocellular carcinoma (HCC). Interactions between viruses and host cells are quite complicated and not always fully understood. In general, the infection cycle of viruses is a multi-step process. A closer understanding of the entire life cycle of the virus is a major prerequisite for the invention of effective drugs. Viral hepatitis B and C have long been treated mainly with interferon alfa. Ribavirin was later added to HCV treatment and nucleoside / nucleotide analogs (NA) were introduced for HBV. Interferon was later pegylated to improve its properties. However, these drugs did not provide sufficient efficacy and were additionally associated with a number of side effects. It is precisely because of these disadvantages of the current...