Submitted by Hiolanda Rêgo (hiolandar@gmail.com) on 2013-09-19T16:24:56Z No. of bitstreams: 1 Tese_Med_ Anselmo de Santana Souza.pdf: 4782838 bytes, checksum: af3640ed9c747b1632ed0e17ef967c30 (MD5) / Approved for entry into archive by Flávia Ferreira(flaviaccf@yahoo.com.br) on 2013-09-22T23:49:14Z (GMT) No. of bitstreams: 1 Tese_Med_ Anselmo de Santana Souza.pdf: 4782838 bytes, checksum: af3640ed9c747b1632ed0e17ef967c30 (MD5) / Made available in DSpace on 2013-09-22T23:49:14Z (GMT). No. of bitstreams: 1 Tese_Med_ Anselmo de Santana Souza.pdf: 4782838 bytes, checksum: af3640ed9c747b1632ed0e17ef967c30 (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq); National Institute of Health (NIH). / O HTLV-1 é o agente etiológico da leucemia/linfoma de células T do adulto (ATLL) e da mielopatia associada ao HTLV-1 (HAM/TSP). Tem-se documentado que células mononucleares de indivíduos infectados não proliferam quando estimuladas com antígenos não relacionados ao vírus como, por exemplo, derivado proteico purificado de Mycobacterium tuberculosis (PPD) e toxoide tetânico (TT). Alguns fatores que podem estar relacionados a essa falta de resposta são as funções de células T regulatórias e disfunção de células apresentadoras de antígeno. Objetivo: Avaliar a resposta imune de indivíduos infectados pelo HTLV-1 ao toxoide tetânico. Materiais e Métodos: Foram selecionados portadores assintomáticos do HTLV-1 baixo produtor de IFN- e controles sadios. Realizou-se sorologia para TT. Os indivíduos soronegativos para TT foram imunizados. Antes e após imunização, fez-se a sorologia para TT e avaliação da expressão de citocinas (IFN- , TNF e IL-10) por linfócitos T CD4+ e T CD8+ estimulados com TT. Os monócitos dos pacientes e controles, estimulados com TT, foram avaliados para a expressão de HLA-DR, CD80, CD86, TNF, IL-12 e IL-10 antes da imunização. Resultados: Após imunização, os pacientes apresentaram menores títulos de IgG anti-TT quando comparados com os controles (p = 0,007). As células mononucleares dos pacientes, estimuladas com TT, não aumentaram a produção de IFN- , TNF e IL-10 após imunização. A frequência de linfócitos T CD4+ expressando IFN- , TNF e IL-10, após estímulo, foi menor nos pacientes do que nos controles pós-imunização (p = 0,01, p = 0,04 e p = 0,01, respectivamente). Os monócitos dos pacientes não aumentaram a expressão de HLA-DR após estímulo com TT. A expressão de TNF e IL-12 por monócitos de pacientes elevaram-se após estímulo com TT (p = 0,009 e p = 0,006, respectivamente). Conclusões: Os indivíduos infectados pelo HTLV-1, após esquema de vacinação, apresentaram diminuição da resposta imune humoral e celular contra TT. Os monócitos destes pacientes exibiram uma disfunção na apresentação antigênica através do mecanismo de expressão de HLA-DR, porém, o segundo sinal (expressão de CD80 e CD86) e expressão de citocinas não apresentaram anormalidades. Tais resultados sugerem que estes mecanismos imunológicos podem participar no aumento da susceptibilidade dos indivíduos infectados pelo HTLV-1 a adquirir outras doenças infecciosas. / Salvador

Submitted by Fernanda Rodrigues de Lima (fernanda.rlima@ufpe.br) on 2018-08-29T22:24:00Z No. of bitstreams: 2 license_rdf: 811 bytes, checksum: e39d27027a6cc9cb039ad269a5db8e34 (MD5) DISSERTAÇÃO Mirela Lopes Ribeiro.pdf: 2216257 bytes, checksum: 49fcbcbc58366871eb0657347a258122 (MD5) / Approved for entry into archive by Alice Araujo (alice.caraujo@ufpe.br) on 2018-09-10T23:09:58Z (GMT) No. of bitstreams: 2 license_rdf: 811 bytes, checksum: e39d27027a6cc9cb039ad269a5db8e34 (MD5) DISSERTAÇÃO Mirela Lopes Ribeiro.pdf: 2216257 bytes, checksum: 49fcbcbc58366871eb0657347a258122 (MD5) / Made available in DSpace on 2018-09-10T23:09:58Z (GMT). No. of bitstreams: 2 license_rdf: 811 bytes, checksum: e39d27027a6cc9cb039ad269a5db8e34 (MD5) DISSERTAÇÃO Mirela Lopes Ribeiro.pdf: 2216257 bytes, checksum: 49fcbcbc58366871eb0657347a258122 (MD5) Previous issue date: 2017-06-21 / CAPES / O vírus linfotrópico para células T de humanos (HTLV) pode alterar a evolução clínica do vírus da imunodeficiência humana (HIV). Tanto o HTLV como o HIV compartilham as mesmas vias de transmissão e esta coinfecção pode mascarar o diagnóstico da síndrome da imunodeficiência humana (aids). O presente estudo se propôs estimar a prevalência da infecção pelo HTLV e descrever os fatores de risco para coinfecção em pessoas vivendo com HIV/aids (PVHA) acompanhadas no Ambulatório de Doenças Infecciosas e Parasitárias (DIP) do Hospital das Clínicas da Universidade Federal de Pernambuco (HC-UFPE), na cidade do Recife, nordeste do Brasil, no período de 2013 a 2016. Foi realizado um estudo transversal e os pacientes após a assinatura do termo de consentimento livre e esclarecido, foram entrevistados e procedeu-se a coleta sanguínea e a análise dos prontuários. As amostras foram encaminhadas ao Setor de Virologia do Laboratório de Imunopatologia Keizo Asami (LIKA) da UFPE para a realização do ELISA e do teste confirmatório Western Blotting (WB). Foram incluídos no estudo 720 indivíduos vivendo com HIV/aids, dos quais 457 (63,4%) eram do sexo masculino, 347 (48,1%) solteiros, a maioria acima de 40 anos (61,5%) e 374 (51,9%) eram de cor/raça parda. A prevalência da coinfecção HTLV/HIV, determinada pelo ELISA e confirmada pelo WB, foi de 1,5% (11/720). Ressaltando que 10 foram HTLV-1 e um HTLV-2. Dos coinfectados, seis eram homens e cinco mulheres, 73% maior de 40 anos de idade e maioria de raça parda. Não foi encontrada associação entre os fatores de risco para a aquisição da coinfecção. A mediana das contagens de TCD4+ foi maior entre os coinfectados HTLV/HIV, e a mediana da primeira contagem da carga viral do HIV foi maior nos pacientes monoinfectados. A coinfecção HTLV/HIV foi baixa refletindo o perfil da população estudada, que difere de outras regiões do Brasil. / Human T-cell lymphotropic virus (HTLV) can affect the clinical evolution of human immunodeficiency virus (HIV). Both HTLV and HIV share the same transmission routes and this coinfection may mask the diagnosis of human immunodeficiency syndrome (aids). This study aimed to estimate the prevalence of HTLV infection and to describe the risk factors for coinfection in people living with HIV/aids (PLWHA), attended at the outpatient clinic of the Hospital das Clínicas of the Universidade Federal de Pernambuco (HC-UFPE), in Recife, Northeast of Brazil, from 2013 to 2016. A cross-sectional study was performed and the patients were interviewed, after having signed the informed consent forms. The blood samples were collected and medical records were also analyzed. Samples were sent to the Virology Department of the Laboratório de Imunopatologia Keizo Asami (LIKA) at UFPE, for analysis using ELISA and Western Blotting (WB). A total of 720 individuals living with HIV/aids were studied, of which 457 (63.4%) were male, 347 (48.1%) were single, the majority were over 40 years old (61.5%) and 374 (51.9%) were pardo color. The prevalence of HTLV/HIV coinfection, determined by ELISA and confirmed by WB, was 1.5% (11/720). Noticing that 10 were HTLV-1 and one was HTLV-2. Among the coinfected people, six were male and five were female, 73% were over 40 years old and the marjority were pardo color. There was no association between risk factors and the coinfection. The median CD4+ T cells counts were higher among coinfected; however, the median of the first HIV viral load was higher in monoinfected patients. The HTLV/HIV coinfection was low, reflecting the profile of the studied population, which differs from other Brazilian regions.

Doctorat en Sciences médicales / info:eu-repo/semantics/nonPublished

Thesis (Ph. D.)--Ohio State University, 2006. / Title from first page of PDF file. Includes bibliographical references (p. 130-152).

No Brasil vÃrios estudos demonstraram prevalÃncia de coinfecÃÃo pelos vÃrus linfotrÃpico de cÃlulas T humanas â HTLV e vÃrus da imunodeficiÃncia humana â HIV, dentre grupos especÃficos de indivÃduos, que variou de 0,58% a 11,4%. O CearÃ, segundo dados anteriores, representa dentre os Estados do Nordeste, uma Ãrea de baixa endemicidade para a infecÃÃo pelos vÃrus HTLV. Este estudo tem por objetivo estudar aspectos clÃnicos e epidemiolÃgicos da coinfecÃÃo por HTLV e HIV, em Hospital de referÃncia para tratamento de pacientes com HIV do CearÃ. Este estudo à descritivo, do tipo transversal, realizado no perÃodo de maio de 2001 a outubro de 2002. Foram colhidas 420 amostras de sangue de pacientes soropositivos ao HIV, confirmados por Elisa e Western Blot que posteriormente foram testadas para HTLV-I/II, no Centro de Hematologia do Cearà â HEMOCE. Entrevistou-se 337 pacientes e pesquisou-se 165 prontuÃrios mÃdicos para obtenÃÃo de informaÃÃes referentes à dados sÃcio-econÃmicos, fatores de risco para HTLV, prÃticas sexuais e aspectos clÃnicos. Os resultados revelaram valor de soroprevalÃncia geral de 0,95%, distribuÃdos em 0,23% de HIV-HTLV-I e 0,47% de HIV-HTLV-II, seguido de 01 (0,23%) amostra com sorologia indeterminada. NÃo foi evidenciada concomitÃncia de infecÃÃo pelos vÃrus HTLV-I e HTLV-II. A populaÃÃo estudada concentrou maior nÃmero de pacientes na faixa etÃria de 30 a 39 anos, era predominantemente de baixa renda (67,6%), menor grau de escolaridade (44,8%) e constituÃda na sua maioria por heterossexuais (67,8%). Quanto Ãs manifestaÃÃes clÃnicas pesquisadas em 119 indivÃduos, 105 (88,2%) manifestaram doenÃa intercorrente e 14 (11,8%) foram assintomÃticos, sendo 111 (93,27%) com definiÃÃo para diagnÃstico de AIDS. Um percentual elevado dos entrevistados amamentou (38,5%), sendo baixa a exposiÃÃo ao uso de tatuagem (12,2%) e a transfusÃo de sangue (15,9%). Foi notada que a escassez no uso de drogas intravenosas (4,8%), um menor nÃmero de negros (5,6%) e maior nÃmero de preferÃncia heterossexual (67,8%), poderiam ser os principais fatores apontados como responsÃveis pela baixa prevalÃncia encontrada em nosso Estado. / Several studies carried out in Brazil have shown a serum-prevalence rate of HIV / HTLV (Human Immunodeficiency - virus / Human T-Lymphotropic virus) co-infection of 0.58% to 11.4% among specific groups of individuals. Based on previous data, the State of Cearà is considered an area of low HTLV prevalence in the northeastern Brasil. This study evaluated the clinical and epidemiological aspects of the HIV / HTLV co-infection in a reference hospital for the treatment of HIV infected patients in CearÃ. A descriptive, cross sectional study was performed, in the period of May of 2001 to October of 2002. Blood samples were randomly collected from 420 HIV-positive patients, through Elisa and Western Blot tests, that later were serologically tested for HTLV-I/II in the Hematological Center of Cearà - HEMOCE. Interviews were done in 337 patients and 165 files were searched for socio-economic, risk factors for HTLV, sexual practice and clinical aspects. The results confirmed a general seroprevalence value of 0.95%, distributed as 0.23% of HIV-HTLV-I and 0.47% of HIV-HTLV-II, followed by one (0.23%) sample of undetermined serology. Concomitant infection was not evidenced by the viruses HTLV-I and HTLV-II. The population studied was more frequently 30 to 39 years old, had predominantly lower income (67.6%) and educational (44.8%) levels and were heterosexual mainly (67,8%). In 119 patients evaluated, 105 (88.2%) complained of HIV-related diseases, 14 (11.8%) were asymptomatic and 111 (93.3%) were diagnosed with AIDS. An elevated percentage was breast fed (38.5%), few had had tattoos (12.2%), and also did receive blood products (15,9%). The scarce use of intravenous drugs (4.8%), the few numbers of black individuals (5.6%) and higher numbers of heterosexuals (67.8%), were pointed as possible reasons for the low HTLV prevalence found in this research.

Bien que très proches dans leur organisation génomique, le rétrovirus HTLV-1 est impliqué dans le développement de la leucémie à cellule T de l’adulte (ATL) alors que l’infection par HTLV-2 n’a jamais été associée à des désordres hématologiques malins. La transformation des cellules infectées par HTLV-1 a longtemps été attribuée uniquement à la protéine virale transactivatrice Tax (Tax-1). Cependant, son expression est très faible dans les cellules ATL. La protéine HBZ a été découverte en 2002. Elle est traduite à partir d'un ARNm transcrit à partir du LTR 3' d'HTLV-1 et est exprimée par les cellules infectées issues de tous les patients HTLV-1 quel que soit leur statut clinique. HBZ participe au maintien du phénotype tumoral en stimulant la prolifération des cellules leucémiques et intervient dans l'échappement du virus au système immunitaire. Des analyses in silico nous avaient permis de détecter un cadre ouvert de lecture sur le brin complémentaire de l’ARN génomique d’HTLV-2. Mon travail de thèse a consisté à amplifier et caractériser d’une part le transcrit APH-2 et d’autre part la protéine qui en est issue. Nous avons démontré dans un premier temps que la transcription d’APH-2 était initié dans le LTR 3’ et que le transcrit APH-2 était épissé et poly-adénylé. Nous avons ensuite mis en évidence l’expression d’APH-2 dans les lignées infectées par HTLV-2, ainsi que dans des cultures de lymphocytes issus de deux porteurs sains africains. La mise au point d’une technique quantitative de RT-PCR nous a permis de détecter APH-2 ex vivo chez 94% des individus d’une série de 51 porteurs sains américains. Nous avons aussi montré que l'expression de cet ARNm était proportionnelle à la charge provirale. APH-2 code une protéine de 183 acides aminés dont nous avons mis en évidence l’expression dans la lignée MO. Mes travaux ont aussi permis de démontrer le rôle inhibiteur d’APH-2 sur la transcription virale malgré l’absence d’un domaine bZip classique, ainsi que son interaction avec le facteur de transcription CREB. Par immunofluorescence, nous avons établi la localisation nucléaire d’APH-2. La protéine semble associée aux corps PML grâce à une région de six arginines comprise entre les résidus 78 et 92. Cependant, contrairement à HBZ, nous n’avons pas observé d’interactions entre APH-2 et les facteurs cJun, JunD ou le cofacteur de transcription CBP/p300. De plus nous avons observé qu’APH-2 était incapable d’induire la prolifération des lymphocytes in vitro alors qu’une lymphocytose est souvent observée chez les porteurs d’HTLV-2. Grâce à une approche comparative, mes travaux ont ainsi permis d’apporter des éléments nouveaux dans la compréhension de la différence de pathogénicité qu’il existe entre HTLV-1 et HTLV-2. / Although they are very similar in their genomic organization, the HTLV-1 retrovirus is involved in the development of adult T cell leukaemia (ATL) while HTLV-2 has not been associated to any malignant haematological disorders. The tumoral transformation of infected cells was widely associated to the viral transactivactor protein Tax (Tax-1), which modulates many cellular functions. However, its expression is slightly in ATL cells. In 2002, the HBZ protein was discovered, encoded from the 3’ LTR by the complementary strand of HTLV-1 and expressed by all HTLV-1 infected people. HBZ participates in the maintenance of the tumoral phenotype by stimulating leukemic cells proliferation and is involved in the immune system escape. We recently detected a coding region by an in silico analysis in the complementary strand of HTLV-2. My work consisted in the characterization of the APH-2 transcript, and in a second part, of the associated protein. At first, we characterized the APH-2 transcription initiation in the 3’LTR and that transcript was spliced and poly-adenylated and demonstrated that the APH-2 expression in all HTLV-2 cell lines and in short cultured lymphocytes from African healthy carriers. We used a quantitative RT-PCR on uncultured cells from 51 American HTLV-2 healthy carriers and we we detected APH-2 expression in 94% of them. We then showed that APH-2 RNA expression is correlated to the HTLV-2 proviral load. The APH-2 transcript encoded a 183 amino acid protein that was shown to be expressed in the HTLV-2 infected Mo cell line. Our work demonstrated the inhibitory functions of APH-2 in the viral transcription and its interaction with the transcriptional cofactor CREB despite the lack of a bZip domain. By an immunofluorescence approach we established the nuclear localisation of APH-2, which is in particular in the PML nuclear bodies. We demonstrated that six arginines in the 78-92 amino acids region is involved in this PML colocalization. Contrary to HBZ, we didn’t observe any interaction with between APH-2 and cJun or JunD factors nor with the transcriptional cofactor CBP/p300. Furthermore we showed that APH-2 is not involved in lymphocyte proliferation in vitro although a lymphocytosis is often observed in HTLV-2 carriers. According to this comparative approach, my work allowed us to better understand the difference of pathogenicity existing between HTLV-1 and HTLV-2.

No description available.

Chez les rétrovirus, la production des protéines virales dépend de l'expression d'un transcrit sens pleine longueur, initié dans le LTR5' et épissé de façon alternative. Des études ont récemment mis en évidence l'existence d'un nouveau type de transcrit antisens, initié dans le LTR3' de certains rétrovirus humains (VIH-1 et HTLV-1). Chez le virus HTLV-1, plusieurs études ont permis l'identification de deux formes d'épissage du transcrit antisens ainsi que la caractérisation d'une nouvelle protéine virale, nommée HBZ (HTLV-1 bZIP factor). Cette protéine se localise au noyau et régule négativement la transcription virale en interagissant avec des facteurs de transcription contenant un motif leucine zipper. Cette étude a pour objectif de caractériser la transcription antisens des rétrovirus humains HTLV-2, HTLV-3 et HTLV-4. Des études de RT-PCR ont permis de détecter un transcrit antisens chez des cellules 293T transfectées par l'ADN proviral des rétrovirus HTLV-2, -3 et -4. Le séquençage des signaux obtenus a confirmé la présence d'épissage dans chacun des transcrits étudiés. Des sites d'initiation de la transcription ont pu être identifiés par des analyses de 5' RACE sur un clone du virus HTLV-2. Un site fonctionnel de polyadénylation a été identifié chez un clone de ces trois rétrovirus. La comparaison des séquences de la protéine HBZ et des ORF antisens des rétrovirus HTLV-2, -3 et -4 démontre que le motif leucine zipper ne semble pas présent chez ces derniers. Dans le but de permettre l'expression et la détection des protéines antisens, les parties codantes des transcrits antisens des rétrovirus HTLV-2, -3 et -4 ont été clonées dans un vecteur d'expression contenant une étiquette Myc. L'analyse des cellules transfectées par microscopie confocale a permis de détecter la protéine antisens de HTLV-3 (APH-3) à la fois dans le noyau et dans le cytoplasme ainsi que les protéines antisens de HTLV-2 et -4 (APH-2 et APH-4) majoritairement dans le noyau. La co-transfection des vecteurs d'expressions de APH-2, APH-3 et APH-4 en présence des vecteurs exprimant les protéines virales Tax1, Tax2 ou Tax3 en plus du vecteur contenant le LTR de HTLV-1 ou de HTLV-2 en amont du gène de la luciférase dans des cellules 293T, a permis de mettre en évidence une inhibition de l'activation de la transcription virale dépendante de Tax par ces nouvelles protéines antisens. Finalement, afin de caractériser l'activité promotrice antisens de ces rétrovirus, le gène rapporteur de la luciférase a été inséré dans les deuxièmes exons de APH-3 et APH-4, compris dans les vecteurs contenant la portion 3' des génomes proviraux de HTLV-3 et -4. La transfection de ces constructions dans des cellules lymphocytaires humaines (Jurkat E6.1), suivie d'une stimulation par une série d'agents activateurs de ces dernières, a permis d'observer une modulation de la transcription antisens. Ces recherches portant sur la transcription antisens des rétrovirus humains ont permis de mettre en évidence que les transcrits antisens des rétrovirus HTLV-2, -3 et -4 ont la capacité de coder pour des nouvelles protéines virales. Les analyses de séquences effectuées ainsi que les observations en microscopie confocale laissent croire que ces protéines pourraient jouer des rôles différents de ceux précédemment attribués à la protéine HBZ. De plus amples études seront nécessaires afin d'identifier les rôles de ces protéines nouvellement découvertes dans le cycle de réplication viral. ______________________________________________________________________________ MOTS-CLÉS DE L’AUTEUR : APH, Rétrovirus, HTLV, Transcription antisens, HBZ.

Human T-cell lymphotropic virus type I (HTLV-I) is a retrovirus that is the causative agent of the fatal disease adult T-cell leukemia (ATL). HTLV-I silently infects over twenty million people worldwide; up to ten percent of these will develop ATL in their lifetime. There are currently no effective treatments for this disease. HTLV-I expresses its genome as polypeptides that must be processed in order to produce infectious virions. Like other retroviruses, HTLV-I encodes an aspartic acid protease to process these polypeptides into mature form. Because the protease is essential in the virus life cycle, it is an attractive target for the treatment of HTLV-I-induced ATL. The present work examines the structure and function of HTLV-I protease. A theoretical structure of the protease is presented, and the function of the C-terminal extension is considered. In order to determine which residues are involved in binding substrate, two experiments were performed: first, several residues were mutated to the corresponding residues in HIV-1 protease to determine whether HTLV-I protease can be made to process an HIV-1 protease substrate; second, an alanine scan was performed to knock out individual residues to assess their importance in binding substrate. This work builds knowledge of the structure and function of HTLV-I protease. By understanding which residues play a role in binding substrate and by developing a clearer picture of the structure of the protease, it will be possible to develop specific inhibitors for HTLV-I protease.

Thesis (Ph. D.)--Ohio State University, 2009. / Title from first page of PDF file. Includes bibliographical references (p. 146-178).