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Doctoral Thesis
DOI
https://doi.org/10.11606/T.42.2020.tde-16122019-104755
Document
Author
Full name
Edione Cristina dos Reis
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2019
Supervisor
Committee
Pontillo, Alessandra (President)
Aguiar, Renato Santana de
Braga, Tárcio Teodoro
Sumida, Telma Miyuki Oshiro
Title in Portuguese
Caracterização genética e funcional de células dendríticas de indivíduos HIV+ estimuladas com flagelina
Keywords in Portuguese
Células dendríticas
Flagelina
HIV-1
Inflamassoma
Abstract in Portuguese
O inflamassoma NLRP3 desempenha um papel fundamental na ativação de células dendríticas (DC) em resposta a adjuvantes vacinais, no entanto, anteriormente foi demostrado que não é adequadamente ativado em DC de indivíduos infectados pelo HIV (HIV-DC), explicando, pelo menos em parte, a fraca resposta à imunização desses indivíduos. Levando em conta que vários receptores citoplasmáticos são capazes de ativar o inflamassoma, e que os componentes bacterianos são considerados eficientes adjuvantes, postulamos que a flagelina bacteriana (FLG), um ligante natural do inflamassoma NAIP/NLRC4, poderia resgatar a ativação do complexo em HIV-DC. O objetivo deste trabalho foi avaliar a capacidade da FLG em ativar adequadamente DC, principalmente HIV-DC, através da estimulação do inflamassoma. Para isso, doadores sadios (HD) e indivíduos infectados pelo HIV foram recrutados para isolamento de monócitos do sangue periférico e obtenção de DC. A ativação de DC pela FLG foi avaliada pelo perfil fenotípico, secreção de citocinas e capacidade de estimular a proliferação de linfócitos T CD4+ e a produção de IFN-γ. A clivagem da caspase-1, a expressão gênica, a formação de specks NLRP3 e NLRC4 foram analisadas para verificar a ativação do inflamassoma. A FLG ativou adequadamente HD-DC e HIV-DC, e foi capaz de estimular a ativação do inflamassoma em células de ambos os grupos. De fato, FLG induziu uma maior produção de IL-1ß em HIV-DC em comparação com LPS. Curiosamente, enquanto que em HD-DC a FLG ativou specks de NLRC4 e NLRP3, em HIV-DC specks de NLRP3 resultaram significativamente reduzidos. Além disso, NLRC4 e NLRP3 parecem estar colocalizados no complexo. Em conclusão, a FLG ultrapasssa o defeito NLRP3-inflamassoma em HIV-DC, através da ativação de NAIP / NLRC4, indicando um possível uso futuro do componente bacteriano como um eficiente adjuvante para indivíduos imunocomprometidos.
Title in English
Functional and genetic characterization of dendritic cells from HIV-1 patients stimulated with flagellin
Keywords in English
Dendritic cells
Flagellin
HIV-1
Inflammasome
Abstract in English
NLRP3 inflammasome plays a key role in dendritic cells (DC) activation in response to vaccine adjuvants, however we previously showed that it is not properly activated in DC of HIV+ patients (HIV-DC), explaining, at least in part, the poor response to immunization of these patients. Taking in account that several cytoplasmic receptors are able to activate inflammasome, and that bacterial components are considered as a novel and efficient adjuvant, we postulated that bacterial flagellin (FLG), a natural ligand of NAIP/NLRC4 inflammasome, could rescue the activation of the complex in HIV-DC. The aim of this work was evaluate the capacity of FLG to properly activate DC, especially HIV-DC, through the stimulation of inflammasome. For this, healthy donors (HD) and HIV+ patients were recruited for peripheral blood monocytes isolation and DC obtainment. DC activation by FLG was evaluated by the meaning of phenotypic profile, cytokines secretion and ability to stimulate CD4+T lymphocytes proliferation and IFN-γ production. Caspase-1 cleavage, genes expression, NLRP3 and NLRC4 specks formation were analysed to verify inflammasome activation. FLG properly activated HD-DC and HIV-DC, and it was able to stimulate inflammasome activation in both types of cells. Of note FLG induced a higher IL-1ß production in HIV-DC compared to LPS. Interestingly, while in HD-DC FLG activated both NLRC4 and NLRP3 specks, in HIV-DC NLRP3 specks resulted significantly diminished reduced. Moreover, NLRC4 and NLRP3 appeared to co-localized in the complex. In conclusion, FLG by-passes NLRP3-inflammasome defect in HIV-DC, through the activation of NAIP/NLRC4, indicating a possible future use of the bacterial component as an efficient adjuvant in immunocompromised individuals.
 
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Release Date
2021-12-15
Publishing Date
2020-02-14
 
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