Objetivos: Determinar a taxa de prevalência de colonização materna por Estreptococo do grupo B na população de gestantes com idade gestacional de 35 semanas ou mais frequentadoras do pré-natal em dois hospitais (Hospital Universitário Júlio Muller - HUJM e Hospital Beneficente Santa Helena) na cidade de Cuiabá, Mato Grosso. Materiais e Métodos: Entre outubro de 2011 e 2013 foram avaliadas 258 gestantes no HUJM e do Hospital Santa Helena. Após concordarem e assinarem o TCLE, as gestantes de 35 semanas ou mais que não tinham sido submetidas ao exame ginecológico e não estavam em uso de antibióticos e atendiam aos critérios de inclusão, foram submetidas a coleta de swab vaginal e retal conforme protocolo estabelecido. Acondicionado em meio de transporte Stuart e no laboratório cultivado em caldo Granada bifásico IGBL. Após 24 horas, amostras com coloração laranja ou avermelhada foram consideradas positivas, caso contrário, nova leitura com 48 horas de cultivo. Os dados foram submetidos a análise estatística utilizando o EPI-Info da OMS. Resultados: Das 258 amostras 13,95% foram positivas para o EGB com IC (95%) de 9.70% a 18.21%. A avaliação estratificada pela idade gestacional predominou nas gestantes de 36 semanas com 35% de positividade, 10.87% para 37 semanas. E 5.88% para 35 semanas. No trabalho de parto prematuro 33.33% e na amniorrexe prematura 28,57% dos casos eram positivos para o EGB. Os demais parâmetros analisados não mostraram significância estatística. Conclusões: A taxa de prevalência da colonização pelo EGB de uma forma global foi estimada em 13.95%, sendo mais elevada na idade gestacional de 36 semanas com taxa de 35%. Este trabalho deve mudar as políticas públicas de saúde na cidade de Cuiabá / Purpose: To determine the prevalence rate of maternal colonization by Group B Streptococcus in the population of pregnant women in the gestational age of 35 or more weeks, attending prenatal care in two hospitals (HUJM - Hospital Universitário Júlio Muller and Hospital Santa Helena), in the city of Cuiabá, Mato Grosso. Materials and methods: Between October 2011 and October 2013, 258 pregnant women were assessed in HUJM and Hospital Santa Helena. After agreeing and signing the FCCT (Free and clarified Consent Term), those pregnant women of 35 weeks or more, who had not undergone gynecological examination, who were not on antibiotics and who also met the inclusion criteria, were subjected to vaginal and rectal swab collection, according to the established protocol. Stowed in Stuart transport medium and cultivated in IGBL biphasic Granada Broth in the laboratory. After 24 hours, samples with orange or reddish colors were considered positive, otherwise, new evaluation with a 48-hour culture was done. Data were submitted to statistical analysis, using OMS\' EPI-Info. The results: From the 258 given samples, 13.21% were positive for EGB, CI (95%) from 9.70% to 18.21%. Evaluation stratified by gestational age was predominant in pregnant women of 36 weeks with 35% positivity rate, 10.87% of pregnant women of 37 weeks and 5.88% of women of 35 weeks. During preterm labor 33.33% and in premature rupture of membranes, 28.57% cases were positive for GBS. Other analyzed parameters showed no significant statistically. Conclusion: The overall prevalence rate of GBS colonization was estimated at 13.95%, being higher in the gestational age of 36 weeks, with a rate of 35%. The present work should change public health policies in the city of Cuiabá

Base teórica: Devido à diminuição do desenvolvimento de novos antimicrobianos nas últimas décadas, terapias combinadas têm sido empregadas contra bactérias multirresistentes como opção à monoterapia no tratamento de infecções graves. Para tratar infecções causadas por Peusdomonas aeruginosa resistente a carbapenêmicos, amicacina e polimixina B têm sido utilizadas em associações com imipenem, pois possuem diferentes mecanismos de ação, o que, teoricamente, indicaria a possibilidade de efeito sinérgico. Objetivo: O objetivo deste trabalho foi verificar a interação, in vitro, de amicacina e polimixina B em associação com imipenem frente a diferentes isolados de P. aeruginosa resistentes a carbapenêmico. Métodos: Foram selecionados isolados de P. aeruginosa resistentes a carbapenêmico oriundos do Hospital de Clínicas de Porto Alegre, coletados no período de janeiro a março de 2015. Foi realizada eletroforese em gel de campo pulsado e detecção do gene blaSPM-1 para selecionar diferentes clones. Seis isolados (três SPM-1 positivos e três negativos para a carbapenemase) foram selecionados para realização da técnica de time-kill, a fim de avaliar a atividade antimicrobiana das combinações de imipenem com amicacina e imipenem com polimixina B. Resultados: Sinergismo ocorreu para combinações de imipenem com amicacina em 3 isolados, dos quais um era SPM-1 positivo e dois eram SPM-1 negativos e todos apresentaram CIMs relativamente baixas a intermediárias. Quanto às combinações de imipenem com polimixina B, houve sinergismo somente para dois isolados, um SPM-1 positivo e um SPM-1 negativo, contudo houve antagonismo em 5 isolados, dois SPM-1 positivos e três SPM-1 negativos. Para 4 isolados, as combinações de imipenem com amicacina tiveram atividade bactericida, enquanto, para todos os isolados, as combinações de imipenem com polimixina B, bem como polimixina B isoladamente, 1x e 2x a CIM apresentaram atividade bactericida. Conclusão: Sinergismo pode ocorrer, para combinações de imipenem mais amicacina, quando os isolados apresentam CIMs relativamente baixas ou intermediárias para imipenem (≤16 μg/mL a 128 μg/mL) e amicacina (≤32 μg/mL). Entretanto, antagonismo aconteceu independentemente de valores altos ou baixos de concentrações inibitórias mínimas para imipenem e polimixina B. Além disso, a presença ou ausência do gene blaSPM-1 não pareceu influenciar nos resultados. / Background: Due to a decrease on new antibiotic development over the last decades, combined therapies have been employed against multigrug-resistant bacteria as an option to monotherapy in severe infection treatment. In order to treat infections caused by carbapenem resistant Pseudomonas aeruginosa, amikacin and polymyxin B have been used in associations with imipenem, because they possess different mechanisms of action, which could, theoretically, indicate the possibility of synergistic effect. Objective: The aim of this study was to verify the interaction, in vitro, of amikacin and polymyxin B in association with imipenem against various carbapenem resistant P. aeruginosa isolates. Methods: Carbapenem resistant P. aeruginosa isolates have been selected from Hospital de Clínicas de Porto Alegre, collected from January to March 2015. Pulsed field gel electrophoresis and detection of blaSPM-1 gene has been performed to select different clones. Six isolates (three SPM-1 positive and three negative for the carbapenemase) were selected for time-kill assay, in order to assess antimicrobial activity of imipenem plus amikacin and imipenem plus polymyxin B combinations. Results: Synergism occurred for combinations of imipenem plus amikacin in three isolates, from which one was SPM-1 positive and two were SPM-1 negative, and all presented relatively low to intermediate minimum inhibitory concentrations. About imipenem plus polymyxin B combinations, synergism occurred in only two isolates, one SPM-1 positive and one SPM-1 negative, however antagonism occurred in five isolates, two SPM-1 positive and three SPM-1 negative. For 4 isolates, imipenem plus amikacin combinations had bactericidal effect, while, for all isolates, combinations of imipenem plus 1x and 2x the MIC of polymyxin B presented bactericidal activity. Conclusions: Synergism can occur, for imipenem plus amikacin combinations, when isolates present relatively low or intermediate MIC of imipenem (≤16 μg/mL to 128 μg/mL) and amikacin (≤32 μg/mL). However, antagonism happened regardless high or low minimum inhibitory concentrations for imipenem and polymyxin B. Also, the presence or absence of blaSPM-1 gene did not seem to influence the results.

Les anticorps monoclonaux représentent aujourd’hui un outil de choix en thérapeutique et en diagnostic. Les anticorps thérapeutiques sont des biomédicaments en plein essor depuis les années 1970 et représentent 10% du marché des produits pharmaceutiques. Les anticorps monoclonaux sont utilisés dans divers domaines : en cancérologie, pour lutter contre les maladies auto-immunes ou en infectiologie. Le nombre des anticorps monoclonaux en développement ne cesse d’augmenter. Les premiers anticorps monoclonaux utilisés en thérapie étaient d’origine murine et leur administration à l’Homme est susceptible de déclencher des effets secondaires. De nouveaux anticorps visant à limiter voir faire disparaitre ces effets indésirables tels que d’abord les anticorps chimériques, puis les anticorps humanisés et enfin les anticorps totalement humains ont été développés. 9 anticorps totalement humains sont actuellement sur le marché et d’autres sont en cours de développement. Le phage display, les souris transgéniques et l’utilisation de lymphocytes B humains sont les trois stratégies mises en œuvre pour produire des anticorps totalement humains. L’utilisation des lymphocytes B humains, peu étudiée à cause d’un faible rendement et de problèmes de stabilité, a connu ces dernières années un regain d’intérêt grâce à l’immortalisation virale par le virus Epstein-Barr et à la découverte de myélomes humains. Dans ce contexte, l’objectif de mon projet de thèse a été la production d’anticorps monoclonaux humains à partir de lymphocytes B humains. Pour ce faire, deux approches basées sur l’immortalisation virale par le virus Epstein-Barr couplée ou non à une immortalisation cellulaire par des myélomes ont été mises en œuvre. La première approche utilise des lymphocytes B mémoires isolés de sang périphérique de donneurs infectés ou vaccinés. L’entérotoxine B de Staphylococcus aureus (SEB) a été utilisée comme modèle.La deuxième approche implique une immunisation in vitro de lymphocytes B naïfs extraits de sang périphérique. Cette stratégie pourrait permettre la production d’anticorps humains contre des antigènes pour lesquels il n’existe pas de donneurs infectés ou vaccinés. Deux modèles, le peptide N-terminal de la neurotoxine A de Clostridium Botulinium A (BoNT/A) et la protéine de fusion ZZTat101, comportant le domaine ZZ de Staphylococcus aureus lié covalemment à la protéine transactivatrice Tat du virus de l’immunodéficience humaine VIH-1, ont été employés. Nous avons réussi à obtenir des IgMs dirigés contre la neurotoxine Clostridium Botulinium A, ainsi que des IgMs (et peut-être des IgGs) dirigés contre la protéine Tat. L’immortalisation par Epstein-Barr, nous a permis d’isoler 7 lignées de lymphocytes immortalisés sécrétant des anticorps IgMs anti-TBA-Nter humains. L’immunisation in vitro produisant essentiellement des IgMs, la possible production d’IgGs après stimulation par la protéine ZZTat101 se révèle un résultat très intéressant. Nous avons montré que la production d’anticorps par ZZTat101 impliquait les 7 cystéines, la région 22-57 et la liaison aux héparanes sulfates de Tat. / The number of monoclonal antibodies used as drug or under clinical investigation increases rapidly. The first murine monoclonal antibodies (mAbs) used in therapy induces human anti-mouse antibodies (HAMAs) when administered to patients. Such HAMAs hamper the therapeutic efficacy of mAbs and induce side effects. To limit these effects, new antibodies were developed during the, last 30 years. Chimeric, humanized and fully human antibodies were engineered. The use of human monoclonal antibodies (hAbs) appears ideal to solve the problem of HAMAs. Nowadays 9 fully human antibodies are available and others are evaluated in clinical trials or currently investigated in research labs. Three methods exist to produce fully human antibodies: the phage display, the transgenic mice and the use of human B lymphocytes. The majority of fully human antibodies resulted from the phage display and the transgenic mice methods. The use of human B lymphocytes is less investigated due to a poor yield and stability problems. These last years, the immortalization process, thanks to the involvement of the Epstein-Barr virus and human myeloma, induced a rise of interest for human B lymphocytes. In this context we decided to develop fully human monoclonal antibodies using human B lymphocytes through immortalization using the Epstein-Barr virus followed or not by an immortalization with a human/mouse heteromyeloma HM. The first approach is based on hAbs production from peripheral blood memory B lymphocytes isolated from infected or vaccinated donors. The Staphylococcus aureus enterotoxine B (SEB) was used as a model. Memory B lymphocytes were purified and cultured in the presence of Epstein-Barr virus (EBV). The transformation of memory B lymphocytes by EBV allowed the generation of immortalized B lymphocytes lines producing IgGs antibodies directed against SEB. We succeeded in isolating 6 EBV-immortalized memory B lymphocytes lines secreting anti-SEB IgGs antibodies. After many attempts to immortalize EBV immortalized memory B lymphocytes lines secreting anti-SEB antibodies with myeloma, the fusion of a EBV immortalized memory B lymphocytes with the human/mouse heteromyeloma HM led to an hybridoma. Unfortunately this hybridoma has rapidly lost its capacity to secrete d’IgGs anti-SEB. In the second approach the hAbs production implies the in vitro immunization of peripheral blood naïve lymphocytes. This strategy could allow the hAbs production against antigens for which no infected or vaccinated donors may be available. The Clostridium Botulinum neurotoxin A (BoNT/A), the most powerful toxin, and its N-terminal peptide (TBA-Nter) or the fusion protein ZZTat101 were used as models. ZZTat101 is a fusion between the ZZ domain of Staphylococcus aureus and the Tat protein of the human immunodeficiency virus HIV-1. Monocytes, B lymphocytes and T lymphocytes were isolated from human PBMC depleted of Natural killer. These cells were tools to develop efficient in vitro immunization protocols. IgMs directed against TBA-Nter and also IgMs (and possibly IgGs) directed against Tat were obtained. The use of the Epstein-Barr virus induced 7 EBV immortalized lines secreting anti-TBA-Nter IgMs antibodies. Unfortunately, after fusion with the heteromyeloma HM no hybridoma was isolated against TBA-Nter and Tat. The ZZTat101 mechanism involved on humoral response was studied, showing that the 7 cysteines, the region 22-57 and the ability of Tat to bind heparane sulfate are necessary to trigger the humoral response.

Plus de 80% des cancers colorectaux sont initiés par la perte de fonction du gène Apc. Afin d’identifier de nouvelles cibles thérapeutiques, nous avons utilisé des modèles murins présentant des mutations du gène Apc et recherché par des analyses de puces à ADN de nouveaux événements moléculaires impliqués au cours de la carcinogenèse intestinale.Cette approche nous a permis d’identifier une activation de la signalisation Notch tout au long du processus tumoral. Toutefois, cette activation n’est pas un élément clé de la progression tumorale puisque son inhibition n’empêche pas le phénotype tumoral induit par la perte du gène Apc. En parallèle, nos travaux ont permis d’identifier une induction de l’autophagie tout au long de la carcinogenèse intestinale. L’activation de ce processus biologique ouvre, quant à lui, de nouvelles perspectives thérapeutiques dans le traitement du CCR. / Over 80% of colorectal cancers are linked to an Apc mutation. To identify new therapeutic targets, we used mouse models with Apc mutations and performed microarray experiments to identify key molecular events involved in intestinal carcinogenesis. This approach allowed usto identify an activation of the Notch signaling all along tumor progression. However, this induction is dispensable for tumor development since its inhibition did not prevent the Apc phenotype. In addition, we have identified an induction of autophagy throughout intestinal carcinogenesis which appears to be an attractive therapeutic target in the treatment of CRC patients.

Les résultats insuffisants de la radiochimiothérapie conventionnelle ont motivé l’évaluation de nouvelles cibles afin de moduler la radiosensibilité tumorale: voies intrinsèques impliquées dans la réponse aux rayonnements ionisants, vascularisation tumorale, stroma non vasculaire. A travers cette thèse, nous avons évalué trois nouvelles stratégies de radiosensibilisation pharmacologique. Nous avons d’abord étudié en association à la radiothérapie l’intérêt de la modulation de l’axe p53/Mdm2 par le JNJ26854165, un inhibiteur de la dégradation de p53 par le protéasome. Les résultats in vitro et in vivo dans des xénogreffes sous-cutanées de cancers bronchiques non à petites cellules (CBNPC) montrent que cette stratégie permet d’améliorer significativement l’efficacité de la radiothérapie. Nous avons également rapporté des résultats encourageants in vitro dans plusieurs lignées cellulaires tumorales avec un nouvel agent antivasculaire ciblant la tubuline, l’EHT 6706. Cette stratégie augmentait l’efficacité de l’irradiation et potentialisait l’effet antiprolifératif de certains agents de chimiothérapie conventionnelle. Enfin, le développement le plus abouti a consisté en l’évaluation de l’association d’un triple inhibiteur de MET/AXL/FGFR en association à l’irradiation in vitro et dans des modèles de CBNPC implantés en xénogreffes sous-cutanées, mais également sous forme de tumeurs pulmonaires orthotopiques. Cet agent pharmacologique potentialisait l’efficacité de la radiothérapie dans des lignées ne surexprimant pas MET. Il est apparu que l’activité de la drogue faisait intervenir, au moins partiellement, l’inhibition de l’activité d’acteurs de la cytocinèse. Ces trois évaluations, qui s’inscrivent dans la recherche translationnelle, montrent l’importance de la recherche préclinique pour les études d’association aux rayonnements ionisants. Seul un développement préclinique rationnel permettra de faire émerger de nouveaux standards dans le domaine de la biomodulation pharmacologique de la radiosensibilité tumorale. / Insufficient results of conventional chemoradiation have encouraged assessment of new targets for radiosensitization: intrinsic cellular pathways involved in radiation response, tumor angiogenesis, and nonvascular stroma. We have investigated these three strategies for pharmacological radiosensitization. First, we examined the usefulness of targeting p53/Mdm2 pathway in combination with irradiation. In vitro and in vivo results obtained in non-small cell lung carcinoma (NCSLC) showed that this strategy was promising for enhancing radiation efficacy. We also found encouraging results within several cell lines with a novel vascular disrupting agent targeting tubulin. This strategy enhanced radiation effects and also increased the antiproliferative effects of various chemotherapeutics. Finally, the most advanced preclinical development was obtained with a novel MET/AXL/FGFR inhibitor, which improved effectiveness of radiation therapy in vitro and in subcutaneous and orthotopic models of non MET-dependent cell cancer lines. This effect was not only related to an inhibition of stroma/cancer cell interactions, as it probably involved activity toward actors of cytocinesis. These studies, which are part of translational research, highlight the importance of preclinical investigations in the area of radiation research. Only rationale preclinical development will allow new standards to emerge for pharmacological modulation of tumor radiosensitivity.

The nuclear factor-kB (NF-kB) signaling cascade is comprised of two branches, the canonical and alternative NF-kB pathways. Signaling through the alternative NF-kB pathway culminates in the activation of the downstream transcription factor subunits, RELB and NF-kB2. The biological roles of RELB and NF-kB2 within the B cell lineage have been obscured in constitutional knockout mice by the diverse functions of these subunits in non-B cell types. To overcome these limitations, conditional alleles were generated to investigate the roles of RELB and NF-kB2 in B cell development. These alleles allowed the identification of complex functional requirements for RELB and/or NF-kB2 in naïve B cells, germinal center (GC) B cells and plasma cells (PCs). These functional requirements may have implications for B cell malignancies that display mutations that constitutively activate the alternative NF-kB pathway. A large body of work has demonstrated that B cell activating factor (BAFF) signaling is critical for the maintenance of mature B cells. However, the contribution of the alternative NF-kB subunits that are activated downstream of BAFF remained unclear, especially in regards to their specific target genes. We have identified critical, B cell-intrinsic roles for RELB and NF-kB2 in the maintenance of mature B cells. In response to BAFF, these subunits were found to control the expression of anti-apoptotic genes, genes that ensure correct positioning within the B cell niche, and genes involved in promoting B–T cell interactions that allow effective antigen-mediated activation. During the GC B cell reaction, light zone (LZ) B cells undergo affinity-based selection mediated by T follicular helper (Tfh) cells. A subset of LZ B cells show activation of the NF-kB signaling cascade, suggesting a critical role for NF-kB in the selection of high-affinity GC B cells. We here report that GC B cell development occurred normally in mice with conditional deletion of either relb (RELB) or nfkb2 (NF-kB2) in GC B cells. In contrast, the simultaneous ablation of both subunits caused rapid involution of established GCs, similar to what has been observed for ablation of the canonical NF-kB transcription factor subunit c-REL. Intriguingly, RNA-sequencing analysis of relb/nfkb2-deleted GC B cells revealed no overlap between the genes controlled by RELB/p52 and c-REL within GC B cells. This suggests that signaling through the separate NF-kB pathways in GC B cells results in the expression of different biological programs that are independently required for the maintenance of the GC reaction. In addition, we observed that human PCs and PC precursors within the LZ showed high protein levels of NF-kB2 compared to surrounding lymphocytes, suggesting a biological role for this subunit in PCs. Indeed, ablation of nfkb2 alone in GC B cells led to a dramatic decrease in antigen-specific serum IgG1 and antigen-specific IgG1-secreting cells. Interestingly however, the mice developed normal frequencies of PCs, suggesting a role for NF-kB2 in PC physiology rather than differentiation.

Introdução: O número de idosos representa uma parcela crescente da população devido ao aumento da expectativa de vida. Na epidemiologia, há diferenças importantes, em relação aos casos de hepatites virais que acometem os idosos quando comparados à população mais jovem. A Hepatite B é um problema de saúde pública mundial e uma das principais causas de doença hepática avançada, como cirrose hepática e hepatocarcinoma. O risco de hepatocarcinoma entre pessoas infectadas pelo Vírus da Hepatite B é maior em homens com mais de 60 anos, expostos cronicamente ao vírus. Alguns estudos apontam que a probabilidade de cronificação e complicações são maiores nos idosos, sendo uma preocupação para a saúde pública. Nota-se, todavia, escassez de estudos sobre Hepatite B para este grupo etário, sendo o intuito deste trabalho acrescentar informações nesta direção. Objetivo: Verificar a prevalência e fatores associados à infecção pelo vírus da Hepatite B entre idosos. Métodos: Estudo transversal de base populacional, realizado por meio de entrevista domiciliar e exames laboratoriais. Trata-se de amostra representativa de idosos (60 anos ou mais) residentes no Município de São Paulo, Brasil, participantes do Estudo Saúde, Bem-Estar e Envelhecimento, no ano de 2010. Resultados: No município de São Paulo, 14,7% (IC95%: 12,5-17,3%) dos idosos apresentaram diagnóstico positivo para infecção por Hepatite B. Na análise univariada, observou-se maior prevalência para Hepatite B no sexo masculino (19,8%), faixa etária entre 80 anos e mais (19,4%), sem companheiro (16,5%), residindo só (20,8%), com atividade sexual no último ano (19,8%). Na análise múltipla, observou-se que homens (OR= 1,60; IC95%: 1,01-2,52), indivíduos que indicaram morarem só (OR= 1,95; IC95%: 1,07-3,57) e pessoas idosas com relato de atividade sexual no último ano (OR= 1,76; IC95%: 1,05-2,94) apresentaram maior probabilidade de ter infecção pelo vírus da Hepatite B, em modelo controlado por idade e estado civil. Conclusão: A constatação desses fatores pode servir de base para a organização de ações educativas e a implementação de estratégias de prevenção da Hepatite B direcionadas para essa população. / Introduction: The elderly represent a growing portion of the population due to the increase in life expectancy. Epidemiologically, there are important differences between the cases of viral hepatitis in the elderly and those of the younger population. Hepatitis B is a public health concern in the whole world, and one of the major causes of severe liver disease, such as cirrhosis of the liver and hepatocellular carcinoma. The risk of hepatocellular carcinoma in the population infected by Hepatitis B virus is higher among men with more than 60 years of age, with chronic exposure to the virus. Some studies indicate that chronification and complications are more likely to occur in the elderly, making this a health care concern. However, there is a shortage of studies on Hepatitis B for this age group, and the purpose of this study is to add information in this direction. Objective: To verify the prevalence and associated factors of the Hepatitis B virus infection among the elderly. Methods: A cross-sectional population-based study, accomplished through home interviews and laboratorial blood tests. This is a representative sample of older adults residing in the city of São Paulo, Brazil, participants of the \"Health, Well-Being and Aging\" Survey, in the year of 2010. Results: In São Paulo, 14,7% (CI95%: 12,5-17,3%) of older adults showed positive diagnostic for Hepatitis B virus infection. In the univariate analysis, the highest prevalence of Hepatitis B was shown in the male population (19.8%), aged 80 or more years (19.4%), with no partner (16.5%), living alone (20.8%), reporting sexual intercourse in the last year (19.8%). In the multiple analysis, it was observed that men (OR = 1.60, 95% CI: 1.01-2.52), individuals living alone (OR = 1.95, 95% CI: 1.07-3.57) and elderly people that reported sexual activity in the last year (OR = 1.76, 95% CI: 1.05-2.94) were more likely to have Hepatitis B virus infection in a model controlled by age and marital status. Conclusion: The realization of these associations can ground the organization of educational practices and the implementation of strategies to prevent Hepatitis B virus -infection in this age group.

Estima-se que aproximadamente 400 milhões de indivíduos sejam portadores crônicos do vírus da hepatite B no mundo. Quando incide em adultos, a doença apresenta elevada proporção de evolução para a cura, ao passo que na ocorrência de transmissão vertical, o risco de cronificação chega a 90%, aumentando muito a possibilidade de graves conseqüências para a criança, entre as quais cirrose e hepatocarcinoma primário. A possibilidade de triagem para a identificação das gestantes portadoras do vírus da hepatite B e a conseqüente adoção de medidas profiláticas – imunização ativa e passiva – permite a prevenção segura da transmissão vertical. Em 1999, o Programa do Pré-Natal da Secretaria Municipal da Saúde de Ribeirão Preto introduziu em sua rotina o screening para o HBsAg, com a finalidade de identificar as gestantes portadoras do vírus. Este trabalho objetiva estudar alguns aspectos referentes à hepatite B entre as gestantes atendidas pela Rede Municipal de Saúde, bem como realizar uma avaliação sistematizada do Programa do Pré-Natal. Para este trabalho foram criados dois grupos de gestantes. O primeiro formado pelas pacientes com primeira avaliação sorológica do Pré-Natal para aquela gestação, realizada no período de 01 de novembro de 2001 a 31 de outubro de 2002, com a finalidade de estimar a prevalência do HBsAg e também verificar a proporção de recém-nascidos, filhos de mães portadoras deste marcador, que receberam os cuidados preconizados para tal situação. Já o segundo grupo foi formado pelas pacientes, na mesma condição, avaliadas no período de 01 de julho de 2002 a 30 de junho de 2003, com a finalidade de também estimar a prevalência do HBsAg, bem como a proporção de portadoras do HbeAg e anti-Hbe. Os valores de prevalência do HBsAg encontrados foram os seguintes: para o primeiro grupo 0,5 (IC 95% : 0,3 – 0,7) e, 0,4 (IC 95%: 0,2 – 0,6) para o segundo. Verificou-se que em 25,0% dos 24 partos realizados no município não foram realizados os procedimentos de profilaxia preconizados como ideais, no que diz respeito à rapidez da solicitação de vacina e imunoglobulina. Isso foi devido, parcialmente, a deficiências na qualidade de registro das informações em diferentes instâncias. As proporções de portadoras do HBeAg e anti-HBe foram respectivamente 5,9% (IC 95%: 0 – 17,1) e 90,5% (IC 95%: 77,9 – 100). Este trabalho procura apresentar informações que sirvam de base para reflexões a respeito do fluxo de procedimentos do Programa do Pré-Natal, visando elevar sua efetividade e superar os obstáculos encontrados. / It is estimated that about 400 million persons are chronic carriers of the hepatitis B virus worldwide. When it occurs in adults, the disease shows high proportions of benign evolution, meanwhile in vertical transmission the risk of becoming a chronic carrier approaches 90%, elevating the possibilities of serious consequences to the child, including cirrhosis and primary hepatocellular carcinoma. The possibility of performing screening tests for the identification of pregnant women carriers of the hepatitis B virus and the consequent adoption of prophylactic measures – active and passive immunization – allow safe prevention of the vertical transmission. In 1999, the Pre-Natal Program of the Municipal Secretary of Health of Ribeirão Preto, SP, Brasil, included in its routine services the screening for the HBsAg, towards the identification of pregnant carriers of the virus. This study aims to look into some aspects referring to hepatitis B among the pregnants being attended by the Municipal Health Network, as well as perform a careful systematic evaluation of the Pre-Natal Program. In order to perform this study, two groups of pregnants were selected. The first one formed by those patients having had their first pre-natal serological evaluation done for that pregnancy during the period of November 1, 2001 until October 31, 2002, with the purpose of estimating the HBsAg prevalence and also to verify the proportion of the new-borns of HBsAg carriers that received the recommended care for that situation. The second group was formed by those patients on the same condition, evaluated during the period of July 1, 2002 until June 30, 2003, with the purpose of also estimating the HBsAg prevalence, as well as estimating the proportion of HBeAg and anti-HBe carriers. The HBsAg prevalences were the following: for the first group of pregnants 0,5 (CI 95% : 0,3 – 0,7) and 0,4 (CI 95%: 0,2 – 0,6) for the second one. It was verified that in 25,0% of the 24 births that happened in the city, the recommended care were not taken, when it comes to the prompt request of the specific vaccine and immunoglobulin. This fact was, partially, due to deficiency in the data files quality of the different institutions. The proportions of HBeAg and anti-HBe carriers found were, respectively, 5,9% (CI 95%: 0 – 17,1) and 90,5% (CI 95%: 77,9 – 100). This study intends to present data that can be the starting point towards reflections on the established procedures of the Pre-Natal Program, in order to increase its effectiveness and surpass the found obstacles.

Haemophilus influenzae type b (Hib) é uma importante bactéria Gram-negativa causadora de pneumonia, meningite e septicemia em crianças abaixo dos 5 anos de idade. A prevenção contra a doença pode ser alcançada pela imunização da população com vacina conjugada polissacarídeo-proteína, uma vez que a vacina de polissacarídeo não é eficiente. As vacinas conjugadas disponíveis comercialmente custam para o governo brasileiro cerca de US2,7 a dose, sendo necessárias no mínimo 3 doses para imunização completa. O presente estudo desenvolveu um novo método de conjugação de polissacarídeo capsular de Hib (PRP) com toxóide tetânico (TT). O método hidrazona baseia-se em 3 etapas simples: oxidação e derivatização de PRP com espaçador molecular e conjugação com TT na presença de uma carbodiimida e de um éster amino reativo. Após um estudo detalhado de cada etapa do método hidrazona, o novo processo mostrou excelentes resultados de rendimento mesmo após escalonamento. A imunogenicidade e o índice de avidez do conjugado hidrazona foram avaliados e os resultados encontrados foram comparáveis a vacina comercial Hiberix®. A técnica de HPSEC mostrou-se eficaz e o perfil cromatográfico do conjugado hidrazona foi muito similar ao da vacina Hiberix. Finalmente, o novo processo de conjugação de vacina permitiu o desenvolvimento de uma poderosa tecnologia constituindo uma excelente opção para o governo brasileiro. / Haemophilus influenzae type b (Hib) is an important encapsulated bacteria, which causes pneumonia, meningitis and septicaemia in infants. Prevention against infection is achieved by the currently available polysaccharide protein conjugate vaccine. However, due to its high production costs (around U$ 2,7 per dose) this formulation cannot be used in mass immunization programs in Brazil. In the present study, we developed a new method for the conjugation of Hib polysaccharide (PRP) and tetanus toxoid (TT). The hydrazone method is based on 3 singles steps: PRP oxidation, PRP derivatization with linker spacer and conjugation with TT in the presence of carbodiimide and an amino reactive ester. After detailed study of each step of method, the new process showed very good yield of conjugation even when it was scaled-up. The immunogenicity and the avidity index of hydrazone conjugate were evaluated and the results were comparable with those obtained with the commercial vaccine Hiberix®. The HPLC hydrazone profile was very similar to HPLC Hiberix profile. Finally, the new conjugation process allows the development of a powerful vaccine technology, constituting an excellent choice for the brazilian government.

Introdução: As hepatites virais causadas pelo VHB e VHC são um importante problema de saúde pública, representando, juntas, mais de 530 milhões de indivíduos no mundo inteiro, que desenvolveram hepatites crônicas ao serem expostos a esses vírus. No entanto, grande parte da população infectada desconhece sua condição, pois frequentemente encontra-se assintomática. O diagnóstico dessas viroses muitas vezes se dá de forma ocasional, geralmente após procedimentos clínicos rotineiros ou após a realização de doações voluntárias de sangue. Entretanto, já foi demonstrado que mesmo assintomáticos, ou com a presença de sintomas mínimos, os portadores do VHB ou do VHC podem apresentar distúrbios de ordem emocional, social ou mesmo físicos, que repercutem na queda de sua QVRS. Objetivos: O principal objetivo desse estudo foi o de avaliar o impacto que a informação do diagnóstico da presença do VHB ou do VHC poderia causar na QVRS de doadores voluntários de sangue. Método: Participaram do estudo 105 doadores com sorologias alteradas ou para o VHB ou para o VHC nos exames de triagem das doações de sangue, sendo que 65 (62%) eram do sexo masculino e 40 (38%) eram do sexo feminino. Deste total, 32 (30,5%) apresentaram confirmação do diagnóstico para o VHB e 35 (33,3%) apresentaram confirmação do diagnóstico para o VHC. Entretanto, 38 doadores de sangue (36,2%), não tiveram a confirmação dessas viroses após a realização de exames confirmatórios, sendo que seus exames de triagem alterados na doação de sangue foram considerados falso-positivos. Os doadores com diagnóstico sorológico confirmado foram divididos em 2 grupos: um grupo formado por 32 doadores com diagnóstico de hepatite B e um grupo de 35 doadores com diagnóstico de hepatite C. Foram estabelecidos 2 grupos controles para as comparações. O primeiro, um grupo controle negativo, composto de doadores com sorologias negativas em doações de sangue sucessivas, pareados por sexo, cor e idade, e em igual número para cada grupo de doadores infectados. O segundo, um grupo controle falso-positivo, composto por 38 doadores que apresentaram sorologias falso-positivas na doação de sangue. As avaliações da QVRS nos indivíduos infectados foram realizadas em 3 etapas: a) 1ª Etapa: quando compareciam no banco de sangue para a repetição dos exames alterados na doação, contudo sem saberem qual alteração sorológica estava sendo investigada. b) 2ª Etapa: Quando o médico do banco de sangue informava ao doador qual era o seu diagnóstico. c) 3ª Etapa: No momento em que o doador iniciava o acompanhamento clínico ambulatorial. As comparações foram feitas seguindo-se quatro abordagens: a) Comparando-se o nível de percepção das diferenças na QVRS, conforme a evolução das etapas do estudo, através de uma análise longitudinal de cada grupo infectado. b) Comparando-se as diferenças na QVRS entre os grupos infectados com o VHB ou com o VHC e doadores do grupo controle negativo. c) Comparando-se as diferenças na QVRS entre os grupos infectados com o VHB ou com o VHC e doadores do grupo controle falsopositivo. d) Comparando-se a QVRS do grupo de doadores com diagnóstico de hepatite B com a QVRS do grupo de doadores com diagnóstico de hepatite C, no momento em que iniciaram o acompanhamento ambulatorial. As avaliações foram realizadas utilizando-se o questionário SF-36, genérico, de aplicação na população geral, e o questionário LDQOL, de aplicação específica nas doenças hepáticas. Resultados: Nas análises longitudinais, utilizando-se o questionário SF-36, somente o grupo de doadores com diagnóstico de hepatite C demonstrou diferenças estatisticamente significantes da QVRS, nos domínios Dor (p = 0,011), Estado Geral da Saúde (p < 0,001), Aspectos Sociais (p = 0,019), Saúde Mental (p = 0,033) e no Componente Físico da escala sumarizada do SF-36, à custa do aumento nos escores dos questionários. Diferentemente, o grupo de doadores com diagnóstico de hepatite B não apresentou diferenças estatisticamente significantes na análise longitudinal. Nas comparações do grupo de doadores com diagnóstico de hepatite B com seu grupo-controle negativo, utilizando-se o questionário SF-36, foram encontradas diferenças estatisticamente significantes: a) Fase 1 do estudo: nos domínios Dor (p = 0,036), Estado Geral de Saúde (p = 0,007) e no Componente Físico (p = 0,004) da escala sumarizada do SF-36. b) Fase 2 do estudo: nos domínios Capacidade Funcional (p = 0,034), Limitação por Aspectos Físicos (p = 0,048), Dor (p = 0,035), Estado Geral de Saúde (p = 0,002), Saúde Mental (p = 0,047) e no Componente Físico (p = 0,007) da escala sumarizada do SF-36. Fase 3 do estudo: nos domínios Capacidade Funcional (p = 0,028), Dor (p = 0,002), Estado Geral de Saúde (p = 0,006), Vitalidade (p = 0,046) e no Componente Físico (p = 0,006). Nas comparações do grupo de doadores com diagnóstico de hepatite B com o grupo controle falso-positivo, utilizando-se o questionário SF-36, foram encontradas diferenças estatisticamente significantes: a) Fase 1 do estudo: nos domínios Limitação por Aspectos Físicos (p = 0,032). b) Fase 2 do estudo: nos domínios Capacidade Funcional (p = 0,006), Limitação por Aspectos Físicos (p = 0,006), Dor (p = 0,006), Vitalidade (p = 0,017), Limitação por Aspectos Emocionais (p = 0,038), Saúde Mental (p = 0,030), no Componente Físico (p = 0,019) e no Componente Mental (p = 0,022) da escala sumarizada do SF-36.Nas comparações do grupo de doadores com diagnóstico de hepatite C com o seu grupo controle negativo, utilizando-se o questionário SF-36, foram encontradas diferenças estatisticamente significantes: a) Fase 1 do estudo: nos domínios Capacidade Funcional (p = 0,038), Dor (p = 0,003), Estado Geral de Saúde (p<0,011), Vitalidade (p = 0,034), Aspectos Sociais (p = 0,006), Limitação por Aspectos Emocionais (p = 0,024), Saúde Mental (p = 0,007) , no Componente Físico (p = 0,002) e no Componente Mental da escala sumarizada do SF-36. b) Fase 2 do estudo: nos domínios Limitação por Aspectos Físicos (p = 0,040), Dor (p = 0,017), Estado Geral de Saúde (p < 0,001), Vitalidade (p = 0,019), Aspectos Sociais (p = 0,005), Limitação por Aspectos Emocionais (p = 0,025), Saúde Mental (p = 0,034) e no Componente Físico (p = 0,008) da escala sumarizada do SF-36. c) Fase 3 do estudo: nos domínios Dor (p = 0,041), Estado Geral de Saúde (p = 0,003), Vitalidade (p = 0,030) e Limitação por Aspectos Emocionais (p = 0,027). Nas comparações entre o grupo de doadores com diagnóstico de hepatite C e o grupo controle falso-positivo, utilizando-se o questionário SF-36, foram encontradas diferenças estatisticamente significantes : a) Fase 1 do estudo: nos domínios Limitação por Aspectos Físicos (p = 0,040) e Limitação por Aspectos Emocionais (p = 0,042). b) Fase 2 do estudo: Limitação por Aspectos Físicos (p = 0,010), Vitalidade (p = 0,037), Aspectos Sociais (p = 0,010) e Limitação por Aspectos Emocionais (p = 0,005). Nas comparações da QVRS entre o grupo dos doadores com diagnóstico de hepatite B e o grupo dos doadores com diagnóstico de hepatite C no momento em que iniciavam o acompanhamento clínico, utilizando-se o questionário LDQOL, não foram observadas diferenças estatisticamente significantes. Conclusões: A análise longitudinal dos doadores com diagnóstico de hepatite C, as comparações com o seu grupo-controle negativo e com o grupo controle falso-positivo, demonstraram alteração na QVRS. Segundo a análise longitudinal, os doadores com o VHC, ao evoluírem nas etapas do estudo, apresentaram um aumento em seus escores, sugerindo que as informações prestadas pela equipe médica sobre a doença e a perspectiva de um atendimento clínico adequado causaram uma tranquilização que melhorou o impacto inicial do diagnóstico da doença, promovendo melhora relativa da QVRS. Ainda que houvesse melhora dos escores na análise longitudinal, os doadores com o diagnóstico de hepatite C tiveram queda de sua QVRS, demonstrada quando comparados ao seu grupo-controle negativo e com o grupo-controle falso-positivo. Nos doadores do grupo com o diagnóstico de hepatite B, ao contrário, não houve alteração da QVRS que pôde ser demonstrada na análise longitudinal. Este achado sugere a baixa QVRS nestes indivíduos mesmo antes do diagnóstico, e que não houve alteração perceptível de sua queda neste grupo de infectados durante a evolução do estudo. Entretanto, quando comparados ao grupocontrole de doadores negativos e ao grupo-controle falso-positivo, foi demonstrada queda na sua QVRS, que pôde ser associada ao impacto do diagnóstico da infecção pelo VHB / Introduction: Viral hepatitis caused by HBV and HCV are an important public health problem. Together they represent around 530 million individuals who have been exposed to these viruses all over the world. Much of that infected population is unaware of their condition, because they are often asymptomatic. Diagnosing these viral infections often comes only occasionally, usually after clinical routine procedures or after volunteer blood donations. However, it has been shown that even asymptomatic, or even in the presence of minimum symptoms, carriers living with HBV or HCV virus can show emotional, social or even physical disturbs, which echoes in the fall of their Health Related Quality of Life (HRQOL). Aims: The main goal of this study was to evaluate the impact of the diagnosis information of the HBV or HCV presence could cause on HRQOL of voluntary blood donors. Method: The study included 105 donors with altered serology to HBV or HCV tests in blood donations screening. 65 (62%) blood donors were male and 40 (38%) were female. From this total, 32 (30.5%) blood donors were submitted to HBV diagnosis confirmation and 35 (33.3%) were submitted to HCV diagnosis confirmation. However 38 (36.2%) blood donors had no confirmation of these infections after conducting confirmatory tests, and were considered as false positive result tests. Donors with a confirmed serological diagnosis were divided into 2 groups: a group of 32 blood donors with hepatitis B diagnosis and 35 blood donors with hepatitis C diagnosis. Two control groups have been established for comparisons. The first, a negative control group, composed of donors with negative serology, matched by sex, race and age, and in equal numbers for each group of infected donors. The second, a false-positive control group composed by 38 blood donors who had false-positive serology in their blood donation. Assessments of HRQOL in individuals infected were performed in 3 phases: a) Phase 1: When they went to the blood bank for collecting blood samples to new confirmatory tests, however, they werent informed about which serological test was specifically being investigated. B) Phase 2: When the blood banks doctor informed what the donors diagnosis was. C) Phase 3: At the moment the donor started his clinical follow-up. Comparisons were performed according to 4 approaches: A) Comparing the level of perceived HRQOL differences according to the evolution of the studys phases, through a longitudinal analysis of each group infected. B) Comparing the HRQOL differences between the infected donor groups with HBV or HCV and their negative control group. C) Comparing the HRQOL differences between the infected donor groups with HBV or HCV and the false-positive control group. C) Comparing the HRQOL differences between the infected groups with HBV or HCV virus with the false-positive control group. D) Comparing the HRQOL in the group of donors with diagnosis of hepatitis B and the HRQOL in the group of donors with diagnosis of hepatitis C at the time they started the ambulatory monitoring. Evaluations were performed using the SF-36, the generic form applied for the general population, and the LDQOL, specific form applied in liver diseases. Results: In the longitudinal analysis in patients with HBV or HCV using the SF-36, only the group of donors diagnosed with hepatitis C showed significant differences in HRQOL, which occurred in the domains: Bodily Pain (p = 0.011), General Health (p <0.001), Social Function (p = 0.019), Mental Health (p = 0.033) and in the Physical Component of the summarized SF-36 scale, at the costs of increasing in the questionnaire scores. On different way, the hepatitis B group did not show statistically significant differences in longitudinal analysis. Comparing the hepatitis B diagnosis donors group with their negative control group using the SF-36, were found the following statistically significant differences, in the domains: A) Phase 1 Study: Bodily Pain (p = 0.036), General Health (p = 0.007) and in the Physical Component (p = 0.004) of the summarized SF-36 scale. B) Phase 2 Study: Physical Function (p = 0.034), Role Physical (p = 0.048), Bodily Pain (p = 0.035), General Health (p = 0.002), Mental Health (p = 0.047) And in the Physical Component (p = 0.007) of the summarized SF-36 scale. C) Phase 3 Study = Physical Function (p = 0.028), Bodily Pain (p = 0.002), General Health (p = 0.006), Vitality (p = 0.046), and in the Physical Component (p = 0.006) of the summarized SF-36 scale. Comparisons between the hepatitis B diagnosis donors group and their false positive control using the SF-36, were found statistically significant differences in the domains: A) Phase 1 Study : Role Physical (p = 0.032). B) Phase 2 Study = Physical Function (p = 0.006), Role Physical (p = 0.006), Bodily Pain (p = 0.006), Vitality (p = 0.017), Role Emotional (p = 0.038), Mental Health (p = 0.030), and in the Physical Component (p = 0.019) and Mental Component (p = 0,022) of the SF-36 summarized scale. In comparisons of the group of donors with a diagnosis of hepatitis C with their negative control group, using the SF-36 were found statistically significant differences in the domains: A) Phase 1 Study: Physical Function (p = 0.038), Bodily Pain (p = 0.003), General Health (p <0.011), Vitality (p = 0.034), Social Function (p = 0.006), Role Emotional (p = 0.024), Mental Health (p = 0.007), Physical Component (P = 0.002) and Mental Component of the SF-36 summarized scale. B) Phase 2 study: Role Physical (p = 0.040), Bodily Pain (p = 0.017), General Health (p <0.001), Vitality (p = 0.019), Social Function (p = 0.005), Role Emotional (p = 0.025), Mental Health (p = 0.034) and Physical Component (p = 0.008) of the SF-36 summarized scale. C) Phase 3 Study: Bodily Pain (p = 0.041), General Health (p = 0.003), Vitality (p = 0.030) and Role Emotional (p = 0.027). Comparisons between the group of donors diagnosed with hepatitis C and the control group false-positive, using the SF-36 were statistically significant differences: A) Phase 1 Study: Role Physical (p = 0.040) and Role Emotional (p = 0.042). b) Phase 2 Study: Role Physical (p = 0.010), Vitality (p = 0.037), Social Function (p = 0.010) and Role Emotional (p = 0.005). In comparisons of HRQOL between the group of donors with a diagnosis of hepatitis B and the group of donors diagnosed with hepatitis C at the time of beginning the clinical follow-up, using the questionnaire LDQOL, there were no statistically significant differences. Conclusion: The longitudinal analysis of donors diagnosed with hepatitis C, the comparisons with their negative control group and the control group false-positive, showed changes in HRQOL. According to the longitudinal analysis, donors with HCV, when evoluting in the phases of the study showed an increase in their scores, suggesting that the information provided by the medical team about the disease and the prospect of an appropriate clinical care caused a tranquilization that has improved the initial impact diagnose of the disease, promoting a relative improvement on HRQOL. Although there were improvement in scores on the longitudinal analysis, the donors with the diagnosis of hepatitis C had a reduction of their HRQOL, as demonstrated when compared to their negative control group and the control group false-positive. In the donor group with the diagnosis of hepatitis B, in contrast, there was no change in HRQOL that could be demonstrated in longitudinal analysis. But when compared to their control groups negative and false-positive, statistically significant differences were identified. This finding suggests a poor HRQOL in these individuals even before the diagnosis, and that there was no noticeable change to its fall in this group infected during the course of the study. However, when compared to the negative control group of donors and the false-positive control group it was demonstrated decrease in HRQOL of these donors, which might be associated with the impact of the diagnosis of HBV infection