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Adriana Antônia Da Cruz Furini Malária Vivax No Estado Faculdade de Medicina de São José do Rio Preto Programa de Pós-graduação em Ciências da Saúde Adriana Antônia da Cruz Furini Malária vivax no Estado do Pará: influência de polimorfismos nos genes TNFA, IFNG e IL10 associados à resposta imune humoral e ancestralidade genômica. São José do Rio Preto 2016 Adriana Antônia da Cruz Furini Malária vivax no Estado do Pará: influência de polimorfismos nos genes TNFA, IFNG e IL10 associados à resposta imune humoral e ancestralidade genômica. Tese apresentada à Faculdade de Medicina de São José do Rio Preto para obtenção do Título de Doutor no Programa de Pós Graduação em Ciências da Saúde, Eixo Temático: Medicina e Ciências Correlatas. Orientador: Prof. Dr. Ricardo Luiz Dantas Machado São José do Rio Preto 2016 Furini, Adriana Antônia da Cruz Malária vivax no Estado do Pará: influência de polimorfismos nos genes TNFA, IFNG e IL10 associados à resposta imune humoral e ancestralidade genômica./ Adriana Antônia da Cruz Furini São José do Rio Preto, 2016. 124p. Tese (Doutorado) – Faculdade de Medicina de São José do Rio Preto – FAMERP Eixo Temático: Medicina e Ciências Correlatas Orientador: Prof. Dr. Ricardo Luiz Dantas Machado 1.Ancestralidade; 2. Anticorpos; 3. Citocinas; 4. Malária; 5.Plasmodium vivax. ADRIANA ANTÔNIA DA CRUZ FURINI Malária vivax no Estado do Pará: influência de polimorfismos nos genes TNFA, IFNG e IL10 associados à resposta imune humoral e ancestralidade genômica. BANCA EXAMINADORA TESE PARA OBTENÇÃO DO GRAU DE DOUTOR Presidente/Orientador: Prof. Dr. Ricardo Luiz Dantas Machado 2º Examinador:Prof. Dr. Carlos Eugênio Cavasini 3º Examinador: Profa. Dra. Heloísa da Silveira Paro Pedro 4º Examinador: Profa. Dra. Eny Maria Goloni Bertollo 5º Examinador: Profa. Dra. Maristela Sanches Bertasso Borges Suplentes: Prof. Dra. Érika Cristina Pavarino Prof. Dra. Sônia Maria Oliani São José do Rio Preto, 05/08/2016. SUMÁRIO Dedicatória........................................................................................................... i Agradecimentos...................................................................................................ii Epígrafe................................................................................................................v Lista de Figuras...................................................................................................vi Lista de Tabelas.................................................................................................vii Lista de Abreviaturas e Símbolos......................................................................viii Resumo..............................................................................................................xii Abstract.............................................................................................................xiv Introdução ...........................................................................................................1 Objetivos..............................................................................................................9 Artigos Científicos..............................................................................................11 Artigo 1...............................................................................................................12 Artigo 2...............................................................................................................40 Conclusões........................................................................................................49 Referências........................................................................................................51 Apêndices..........................................................................................................69 Anexos.............................................................................................................109 i DEDICATÓRIA Às minhas filhas Marina e Fernanda, por toda felicidade, de me fazer descobrir tesouros nunca antes sonhados. O melhor de todos os sonhos, ser mãe. Vocês são únicas, arteiras, amorosas, geniosas e felizes! Amo vocês até o infinito. Ao meu eterno namorado, meu marido, meu amigo, meu grande amor: Marcos Amorielle Furini. Amo você. Aos meus pais: “Eu teria tanto para falar, que com palavras nem sei dizer, como foi, é, e sempre será infinito meu amor por vocês”. Tenho certeza que me assistiram do céu..... À minha irmã Edna, querida companheira, incansável na busca da felicidade de todos. Aos meus irmãos, sobrinhos, sobrinhas e família. ii AGRADECIMENTOS A Deus por me dar coragem, força, persistência e oportunidades. Ao meu orientador, Prof. Dr. Ricardo Luiz Dantas Machado, por toda sua dedicação ao meu trabalho nesses quatro anos, e por eu ter compartilhado da sua sabedoria e conhecimento. Obrigada, por tudo que me ensinou meu professor, meu mestre, meu amigo. À aluna Michele Encinas do Curso de graduação em Farmácia e ao aluno Diego Longo Madi do Curso de graduação em Biomedicina (Centro Universitário de Rio Preto), pela disponibilidade, empenho, dedicação, prontidão, enfim por tudo o que fizeram por mim para que meu projeto se concretizasse. Meu eterno obrigada. À pesquisadora Dra. Joseli Ferreira de Oliveira da Fundação Oswaldo Cruz (FioCruz) – Rio de Janeiro pela disponibilidade, prontidão e dedicação para utilização do laboratório e análises de citocinas. Ao Dr. Gustavo Capatti Cassiano. Meu amigo foi um prazer realizar um projeto de Doutorado no mesmo laboratório que você, por sua dedicação a pesquisa, por acreditar que as coisas tem que dar certo, e que isso requer persistência. Obrigada pelos ensinamentos, aconselhamentos. Você é um exemplo de pesquisador a ser seguido. Às minhas companheiras de laboratório, Maíra, Marcela, Pamella, Simone pelos momentos que passamos juntas, pela amizade. iii Aos amigos Juan Camilo Sanchéz Arcilla, Virginia Pereira da FioCruz Rio de Janeiro pela disponibilidade, empenho, dedicação, prontidão. Pelo acolhimento impecável nos momentos que passamos juntos. Obrigada ao Prof. Dr. Carlos Eugênio Cavasini, e as minhas amigas do Laboratório CIM, Valeria Fraga e Luciana Moran, pelo comprometimento, dedicação, ajuda nas análises laboratoriais, e fluxo das amostras. Sem vocês esse trabalho não seria possível. Aos meus amigos e amigas: Bárbara Cruz, Nayara Cruz e Gabriela Cruz, Alessandra Cruz, Franciele Maira, Karina Santana, Maristela Bertasso, Greici Gomes, Regiane Rocha, Neide Blaz, Patricia Perez, Ricardo Fochi, Gisele, Tabata, Cleide Silveira, Edna Matins, Paula Guimarães, Daniela Goés, Daniela Hubner, Glacy Claro, Angela Schempp, Gisele Assunção, Gisele Bueno, Cintia, Raquel Arid, Ivone Fontes, Heloisa. Amizade verdadeira é aquela que o tempo não apaga, a distancia não destrói e acima de tudo o coração não esquece.” Alguns não vejo há anos, mas não poderia deixar de lembrá-los em um momento tão especial da minha vida. Ao meu sogro Décio, minha sogra Sandra e a Tia Darci que me auxiliaram inúmeras vezes ao cuidar das minhas filhas para as viagens a congressos durante o Doutorado. À Faculdade de Medicina de São José do Rio Preto – FAMERP e ao Programa de Pós Graduação em Ciências da Saúde pela excelência em qualidade nos cursos Strictu sensu. iv À Capes – Coordenação de Aperfeiçoamento de Pessoal de Nível Superior pela Bolsa de Estudos. Ao Centro Universitário de Rio Preto, pelo incentivo para a realização desse trabalho. Aos pacientes, muito obrigada. Às instituições de fomento e pesquisa, em especial aos auxílios CNPq, pelos recursos financeiros que foram imprescindíveis para a execução desse trabalho. Aos organizadores do seminário Laveran/Deane sobre malária, Dr. Cláudio Tadeu Daniel Ribeiro e Dra. Maria de Fátima Ferreira da Cruz, pela oportunidade de participar deste evento. Enfim, obrigada a todos, que diretamente ou não, contrubíram para a realização desse trabalho. v EPÍGRAFE “Tenho a impressão de ter sido uma criança brincando à beira-mar, divertindo-me em descobrir uma pedrinha mais lisa ou uma concha mais bonita que as outras, enquanto o imenso oceano da verdade continua misterioso diante de meus olhos”. (Isaac Newton) vi LISTA DE FIGURAS Artigo 1: Figura 1. Binary logistic regression model used to evaluate the frequency of individuals carrying the mutant allele of all analyzed SNPs relative to the individual proportions of genetic ancestry. The shading around the lines represents the 95% confidence interval. The graph was constructed using the ggplot2 package in the R program ....................................................................... 24 vii LISTA DE TABELAS Artigo 1: Tabela1.Characteristics of the study population ................................................... 21 Tabela 2. Distribution of the genotypes between vivax malaria-infected and non-infected individuals ........................................................................................ 22 Tabela 3. Haplotype frequencies in the TNFA and IL10 genes in vivax malaria-infected and non-infected individuals ...................................................... 22 Tabela 4. Haplotype frequency and its association with the proportions of African, European and Native American ancestry ................................................ 23 Artigo 2: Tabela 1. List of polymorphisms, methods, restriction enzymes, primers used for investigation of SNPs in genes TNFA, IFNG, IL10 .......................................... 55 Tabela 2. Levels of IgG antibodies against P. vivax blood stage proteins according to the studied genotypes .....................................................................
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