Correlação entre mutações pontuais em LTR e níveis de carga proviral em portadores assintomáticos do vírus linfotrópico de células T humanas tipo 1 (HTLV-1)
Arquivos
Data
2014-06-30
Autores
Kleine Neto, Walter 
Orientadores
Sanabani, Sabri Saeed
Tipo
Tese de doutorado
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Resumo
Introdução: Estudos in vitro demonstram que deleções ou mutações pontuais inseridas em cada repetição imperfeita de 21 pb do Elemento Responsivo a Tax (TRE) nos promotores virais do Vírus Linfotrópico de células T humanas Tipo 1 (HTLV-1) selvagens aboliram a indução de Tax. A partir deste dado, supomos que mutações similares poderiam afetar a proliferação de células infectadas por HTLV-1 e alterar a carga proviral (CPv). Para testar esta hipótese, conduzimos uma análise genética de corte transversal para comparar seqüências nucleotídicas completas de LTR que cobrem a região TRE-1 em uma amostragem de portadores assintomáticos de HTLV-1 com diferentes níveis de carga proviral. Métodos: Foi avaliado um total de 94 portadores assintomáticos de HTLV-1 com as seqüências de longa repetição terminal (LTR) extremidade 5` determinadas e carga proviral de DNA de Tax definida usando-se uma sensível técnica de PCR em Tempo Real com SYBR Green. Os 94 indivíduos foram divididos em três grupos baseados na medida da carga proviral: 31 com baixa, 29 com intermediária e 34 com alta CPv. Além disso, cada grupo foi comparado levando-se em conta sexo, idade e genótipo viral. Em outra análise, foi comparada a mediana da carga proviral entre indivíduos infectados com o vírus mutante e vírus o tipo selvagem. Resultados: Usando-se uma análise categórica, a substituição G232A, localizada no domínio A do motivo TRE-1, foi detectada em 38,7% (12/31), 27.5% (8/29), e 61.8% (21/34) dos indivíduos com baixa, intermediária e alta CPv, respectivamente. Uma diferença significativa na detecção desta mutação foi encontrada entre sujeitos com alta e baixa CPv e entre aqueles com alta e intermediária CPv (ambos p < 0,05), mas não entre aqueles com baixa e intermediária CPv (p > 0,05). Estes resultados foram confirmados por uma análise nãoparamétrica que mostrou forte evidência de aumento da CPv dentre indivíduos HTLV-1 positivos com a mutação G232A em relação àqueles sem esta mutação (p < 0,03). Nenhuma diferença significante foi observada entre os grupos em relação à idade, sexo ou subtipo viral (p > 0,05). Conclusão: Os dados descritos aqui mostram que alterações no domínio A do motivo TRE-1 do HTLV-1, em especial a mutação G232A pode aumentar a replicação do HTLV-1 na maioria dos pacientes infectados.
Background: In vitro studies have demonstrated that deletions and point mutations introduced into each 21 bp imperfect repeat of Tax-responsive element (TRE) of the genuine human T-cell leukemia virus type I (HTLV-1) viral promoter abolishes Tax induction. Given these data, we hypothesized that similar mutations may affect the proliferation of HTLV-1 infected cells and alter the proviral load (PvL). To test this hypothesis, we conducted a crosssectional genetic analysis to compare the near-complete LTR nucleotide sequences that cover the TRE1 region in a sample of HTLV-1 asymptomatic carriers with different PvL burden. Methods: A total of 94 asymptomatic HTLV-1 carriers with both sequence from the 5’ long terminal repeat (LTR) and a PvL for Tax DNA measured using a sensitive SYBR Green realtime PCR were studied. The 94 subjects were divided into three groups based on PvL measurement: 31 low, 29 intermediate, and 34 high. In addition, each group was compared based on sex, age, and viral genotypes. In another analysis, the median PvLs between individuals infected with mutant and wild-type viruses were compared. Results: Using a categorical analysis, a G232A substitution, located in domain A of the TRE1 motif, was detected in 38.7% (12/31), 27.5% (8/29), and 61.8% (21/34) of subjects with low, intermediate, or high PvLs, respectively. A significant difference in the detection of this mutation was found between subjects with a high or low PvL and between those with a high or intermediate PvL (both p < 0.05), but not between subjects with a low or intermediate PvL (p > 0.05). This result was confirmed by a non-parametric analysis that showed strong evidence for higher PvLs among HTLV-1 positive individuals with the G232A mutation than those without this mutation (p < 0.03). No significant difference was found between the groups in relation to age, sex or viral subtypes (p > 0.05). Conclusion: The data described here show that changes in domain A of the HTLV-1 TRE-1 motif resulting in the G232A mutation may increase HTLV-1 replication in a majority of infected subjects.
Background: In vitro studies have demonstrated that deletions and point mutations introduced into each 21 bp imperfect repeat of Tax-responsive element (TRE) of the genuine human T-cell leukemia virus type I (HTLV-1) viral promoter abolishes Tax induction. Given these data, we hypothesized that similar mutations may affect the proliferation of HTLV-1 infected cells and alter the proviral load (PvL). To test this hypothesis, we conducted a crosssectional genetic analysis to compare the near-complete LTR nucleotide sequences that cover the TRE1 region in a sample of HTLV-1 asymptomatic carriers with different PvL burden. Methods: A total of 94 asymptomatic HTLV-1 carriers with both sequence from the 5’ long terminal repeat (LTR) and a PvL for Tax DNA measured using a sensitive SYBR Green realtime PCR were studied. The 94 subjects were divided into three groups based on PvL measurement: 31 low, 29 intermediate, and 34 high. In addition, each group was compared based on sex, age, and viral genotypes. In another analysis, the median PvLs between individuals infected with mutant and wild-type viruses were compared. Results: Using a categorical analysis, a G232A substitution, located in domain A of the TRE1 motif, was detected in 38.7% (12/31), 27.5% (8/29), and 61.8% (21/34) of subjects with low, intermediate, or high PvLs, respectively. A significant difference in the detection of this mutation was found between subjects with a high or low PvL and between those with a high or intermediate PvL (both p < 0.05), but not between subjects with a low or intermediate PvL (p > 0.05). This result was confirmed by a non-parametric analysis that showed strong evidence for higher PvLs among HTLV-1 positive individuals with the G232A mutation than those without this mutation (p < 0.03). No significant difference was found between the groups in relation to age, sex or viral subtypes (p > 0.05). Conclusion: The data described here show that changes in domain A of the HTLV-1 TRE-1 motif resulting in the G232A mutation may increase HTLV-1 replication in a majority of infected subjects.
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Citação
KLEINE NETO, Walter. Correlação entre mutações pontuais em LTR e níveis de carga proviral em portadores assintomáticos do vírus linfotrópico de células T humanas tipo 1 (HTLV-1). Tese (Doutorado em Ciências) - Escola Paulista de Medicina, Universidade Federal de São Paulo (UNIFESP), São Paulo, 2014.