Publicação
Mycobacteria manipulation of host proteases and inflammatory pathways during infection within human macrophages and dendritic cells
| Resumo: | Macrophages and dendritic cells (DCs) play an essential role during mycobacterial infection, acting as key effector cells in bacterial killing and antigen presentation. Cathepsins are host proteases involved in pathogen destruction and antigen presentation. This makes these proteases, as well as both cell types, perfect targets for mycobacterial manipulation. In the first part of this thesis the role of mycobacteria during macrophage activation and DC maturation was addressed. By flow cytometry the surface expression of activation and maturation markers of mycobacteria infected human macrophages and DCs, was analyzed. It was demonstrated that: (1) M. tuberculosis H37Ra prevents the induction of the classical activated macrophage phenotype; (2) induces the maturation process of immature DCs; (3) the outcome of infection with M. tuberculosis H37Ra differs from that with M. smegmatis. These results indicate that M. tuberculosis infection has a differential role during macrophage activation and DC maturation and that M. tuberculosis H37Ra interferes with these processes differently of the non-pathogenic M. smegmatis. In the second part, we aimed to decipher the role of cathepsins during mycobateria infection of macrophages and DCs. Cathepsins protein levels were analyzed by western blot after infection with pathogenic or non-pathogenic mycobacteria. It was shown that cathepsin B and S levels during M. tuberculosis infection are distinct from those of host cells that internalized M. smegmatis and this is dependent on the host cell species tested. In addition, the role of these proteases in M. tuberculosis intracellular survival was evaluated by silencing cathepsins expression, using shRNA lentiviral vectors. It was observed, that cathepsin B and S knockdowns led to an increase in M. tuberculosis H37Ra intracellular survival. Altogether, these evidences point for a role of both cathepsins in the control of M. tuberculosis intracellular growth and demonstrate that M. tuberculosis modulates these cathepsins differently of the non-pathogenic M. smegmatis. |
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| Autores principais: | Marques, Joana Pereira, 1989- |
| Assunto: | Mycobacterium tuberculosis Macrófagos Células dendríticas Teses de mestrado - 2013 |
| Ano: | 2013 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade de Lisboa |
| Idioma: | inglês |
| Origem: | Repositório da Universidade de Lisboa |
| Resumo: | Macrophages and dendritic cells (DCs) play an essential role during mycobacterial infection, acting as key effector cells in bacterial killing and antigen presentation. Cathepsins are host proteases involved in pathogen destruction and antigen presentation. This makes these proteases, as well as both cell types, perfect targets for mycobacterial manipulation. In the first part of this thesis the role of mycobacteria during macrophage activation and DC maturation was addressed. By flow cytometry the surface expression of activation and maturation markers of mycobacteria infected human macrophages and DCs, was analyzed. It was demonstrated that: (1) M. tuberculosis H37Ra prevents the induction of the classical activated macrophage phenotype; (2) induces the maturation process of immature DCs; (3) the outcome of infection with M. tuberculosis H37Ra differs from that with M. smegmatis. These results indicate that M. tuberculosis infection has a differential role during macrophage activation and DC maturation and that M. tuberculosis H37Ra interferes with these processes differently of the non-pathogenic M. smegmatis. In the second part, we aimed to decipher the role of cathepsins during mycobateria infection of macrophages and DCs. Cathepsins protein levels were analyzed by western blot after infection with pathogenic or non-pathogenic mycobacteria. It was shown that cathepsin B and S levels during M. tuberculosis infection are distinct from those of host cells that internalized M. smegmatis and this is dependent on the host cell species tested. In addition, the role of these proteases in M. tuberculosis intracellular survival was evaluated by silencing cathepsins expression, using shRNA lentiviral vectors. It was observed, that cathepsin B and S knockdowns led to an increase in M. tuberculosis H37Ra intracellular survival. Altogether, these evidences point for a role of both cathepsins in the control of M. tuberculosis intracellular growth and demonstrate that M. tuberculosis modulates these cathepsins differently of the non-pathogenic M. smegmatis. |
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