miR-150 regula a carga parasitária de Leishmania infantum e os níveis de GZMB nas PBMCs de cães com Leishmaniose Visceral Canina.
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Data
2022-10-27
Autores
Soares, Matheus Fujimura
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Universidade Estadual Paulista (Unesp)
Resumo
A Leishmania infantum causa a leishmaniose visceral, uma doença tropical negligenciada que pode modular a resposta imune do hospedeiro por meio de pequenos RNAs não codificantes chamados microRNAs (miRNAs). Alguns miRNAs são expressos diferencialmente em células mononucleares do sangue periférico (PBMCs) de cães com leishmaniose visceral canina (LCan), incluindo o miR-150, cuja expressão está diminuída. Embora o miR-150 esteja negativamente correlacionado com a carga parasitária de L. infantum, não está claro se o miR-150 afeta diretamente a carga parasitária de L. infantum e (em caso afirmativo) como esse miRNA contribuiria para a infecção. Aqui, isolamos as PBMCs de 14 cães naturalmente infectados (grupo LCan) e seis cães saudáveis (grupo controle) e as tratamos in vitro com o mimetizador ou o inibidor do miR-150. Medimos a carga parasitária de L. infantum usando o qPCR e comparamos os tratamentos. Também medimos os níveis de proteína-alvo do miR-150 preditas in silico (STAT1, TNF-α, HDAC8 e GZMB) usando citometria de fluxo ou ensaios imunoenzimáticos. O mimetizador do miR-150 diminuiu a carga parasitária de L. infantum nas PBMCs de cães do grupo LCan. Também descobrimos que a inibição do miR-150 reduziu os níveis de GZMB. Esses achados demosntram que o miR-150 exerce uma função importante na infecção por L. infantum nas PBMCs caninas, o que poderia ser usado para mais estudos visando o desenvolvimento de drogas.
Leishmania infantum causes visceral leishmaniasis, a neglected tropical disease that can modulate the host immune response by altering the expression of small non-coding RNAs called microRNAs (miRNAs). Some miRNAs are differentially expressed in peripheral blood mononuclear cells (PBMCs) of dogs with visceral canine leishmaniasis (CanL), like the down-regulated miR-150. Even though miR-150 is negatively correlated with L. infantum parasitic load, it is unclear if miR-150 directly affects L. infantum parasitic load and (if so) how this miRNA would contribute to infection. Here, we isolated PBMCs from 14 naturally infected dogs (CanL group) and six healthy dogs (Control group) and treated them in vitro with miR-150 mimic or inhibitor. We measured L. infantum parasitic load using qPCR and compared treatments. We also measured miR-150 in silico predicted target protein levels (STAT1, TNF-α, HDAC8, and GZMB) using flow cytometry or enzyme-linked immunosorbent assays. Increasing miR-150 activity diminished L. infantum parasitic load in CanL PBMCs. We also found that inhibition of miR-150 reduced GZMB levels. These findings demonstrate that miR-150 plays an important role in L. infantum infection in canine PBMCs, and these findings merit further studies aiming at drug development.
Leishmania infantum causes visceral leishmaniasis, a neglected tropical disease that can modulate the host immune response by altering the expression of small non-coding RNAs called microRNAs (miRNAs). Some miRNAs are differentially expressed in peripheral blood mononuclear cells (PBMCs) of dogs with visceral canine leishmaniasis (CanL), like the down-regulated miR-150. Even though miR-150 is negatively correlated with L. infantum parasitic load, it is unclear if miR-150 directly affects L. infantum parasitic load and (if so) how this miRNA would contribute to infection. Here, we isolated PBMCs from 14 naturally infected dogs (CanL group) and six healthy dogs (Control group) and treated them in vitro with miR-150 mimic or inhibitor. We measured L. infantum parasitic load using qPCR and compared treatments. We also measured miR-150 in silico predicted target protein levels (STAT1, TNF-α, HDAC8, and GZMB) using flow cytometry or enzyme-linked immunosorbent assays. Increasing miR-150 activity diminished L. infantum parasitic load in CanL PBMCs. We also found that inhibition of miR-150 reduced GZMB levels. These findings demonstrate that miR-150 plays an important role in L. infantum infection in canine PBMCs, and these findings merit further studies aiming at drug development.
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Palavras-chave
LCan., Leishmania infantum., miRNA., miR-150., GZMB, CanL.