immunology of infectious and parasitic diseases - XXXVII Congress ...

STUDIES ON A SECRETED MYCOBACTERIUM TUBERCULOSIS RICIN-
LIKE LECTIN (SMTL-13) DURING INITIAL INTERACTIONS WITH
MACROPHAGES
STEFANNY LUCÍA VILOCHE MORALES(PG)(1), NICOLE MENEZES DE
SOUZA, (PG)(1), LUCAS NOGUEIRA(2), VERÔNICA HOREWICZ(PG)(1),
DANIEL MANSUR (1), ANDRÉ BÁFICA(1).
(1) Laboratório de Imunobiologia, Departamento de Microbiologia, Imunologia e
Parasitologia, Universidade Federal de Santa Catarina
(2) Núcleo de Estudos e Pesquisa em Saúde, Centro de Ciências da Saúde,
Universidade Estadual Vale do Acaraú
Introduction: Infection by Mycobacterium tuberculosis (Mtb) can lead to a
latent state in which the host is able to control the pathogen growth. While
effective cellular immune responses are suggested as critical to control Mtb
growth inside macrophages, it has been demonstrated that mycobacteriaassociated
factors play an important role in the outcome of infection. For that
matter, mycobacterial proteins are an incredible source for potential adjuvant
molecules. We have previously described a novel secreted 13-kDa lectin in
pathogenic Mtb (sMTL-13). Experimental data suggest that this protein is
recognized by B and T cells in TB patients and acts as a PAMP inducing APC
activation. Thus, sMTL-13 can be considered as a potential adjuvant model to
stimulate both cellular and humoral immune responses.
Methods and Results: To confirm the presence of sMTL-13 in Mtb cell wall,
bacteria were stained with mAb anti-sMTL-13 and analyzed by flow cytometry.
This approach demonstrated that sMTL-13, in addition to being secreted, is
anchored within the Mtb cell wall, probably by the signal peptide sequence. To
gain insights on the mechanisms by which sMTL-13 modulates cellular
infection, murine bone marrow-derived macrophages (BMM) were exposed to
mycobacteria pre-incubated with or without D-Gal, an sMTL-13 cognate
carbohydrate. BMM infected with bacteria pre-treated with D-Gal, but not L-Gal,
were found to present decreased numbers of CFU, suggesting that galactosebinding
proteins, perhaps sMTL-13, are involved in macrophage-Mtb
interactions. Accordingly, compared to untreated or L-Gal pretreated groups, D-
Gal-exposed Mtb-infected macrophages displayed diminished TNF production.
We next evaluated whether sMTL-13 is secreted during macrophage infection.
Confocal microscopy analysis revealed that following 48h of infection, sMTL-13
was found to colocalize with intracellular mycobacteria in macrophages. In
addition, the lectin appeared to be present in the cell cytoplasm.
Conclusions: These findings suggest that anchored sMTL-13 probably
functions during the initial interactions of Mtb and macrophages. Moreover, this
lectin seems to be actively secreted by the bacterium and gains the cytosolic
pathway. To elucidate the pathways involved in sMTL-13-macrophage