AUTHOR=Avila-Calderón Eric Daniel , Medina-Chávez Olín , Flores-Romo Leopoldo , Hernández-Hernández José Manuel , Donis-Maturano Luis , López-Merino Ahidé , Arellano-Reynoso Beatriz , Aguilera-Arreola Ma. Guadalupe , Ruiz Enrico A. , Gomez-Lunar Zulema , Witonsky Sharon , Contreras-Rodríguez Araceli 

TITLE=Outer Membrane Vesicles From Brucella melitensis Modulate Immune Response and Induce Cytoskeleton Rearrangement in Peripheral Blood Mononuclear Cells

JOURNAL=Frontiers in Microbiology

VOLUME=11

YEAR=2020

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2020.556795

DOI=10.3389/fmicb.2020.556795

ISSN=1664-302X

ABSTRACT=<p>Similar to what has been described in other Gram-negative bacteria, <italic>Brucella melitensis</italic> releases outer membrane vesicles (OMVs). OMVs from <italic>B. melitensis</italic> 16M and the rough-mutant <italic>B. melitensis</italic> VTRM1 were able to induce a protective immune response against virulent <italic>B. melitensis</italic> in mice models. The presence of some proteins which had previously been reported to induce protection against <italic>Brucella</italic> were found in the proteome of OMVs from <italic>B. melitensis</italic> 16M. However, the proteome of OMVs from <italic>B. melitensis</italic> VTRM1 had not previously been determined. In order to be better understand the role of OMVs in host-cell interactions, the aim of this work was to compare the proteomes of OMVs from <italic>B. melitensis</italic> 16M and the derived rough-mutant <italic>B. melitensis</italic> VTRM1, as well as to characterize the immune response induced by vesicles on host cells. Additionally, the effect of SDS and proteinase K on the stability of OMVs was analyzed. OMVs from <italic>B. melitensis</italic> 16M (smooth strain) and the <italic>B. melitensis</italic> VTRM1 rough mutant (lacking the <italic>O</italic>-polysaccharide side chain) were analyzed through liquid chromatography-mass spectrometry (LC-MS/MS). OMVs were treated with proteinase K, sodium deoxycholate, and SDS, and then their protein profile was determined using SDS-PAGE. Furthermore, PBMCs were treated with OMVs in order to measure their effect on cytoskeleton, surface molecules, apoptosis, DNA damage, proliferation, and cytokine-induction. A total of 131 proteins were identified in OMVs from <italic>B. melitensis</italic>16M, and 43 in OMVs from <italic>B. melitensis</italic> VTRM1. Proteome comparison showed that 22 orthologous proteins were common in vesicles from both strains, and their core proteome contained Omp31, Omp25, GroL, and Omp16. After a subsequent detergent and enzyme treatment, OMVs from <italic>B. melitensis</italic> VTRM1 exhibited higher sensitive compared to OMVs from the <italic>B. melitensis</italic> 16M strain. Neither OMVs induced IL-17, proliferation, apoptosis or DNA damage. Nonetheless, OMVs from the smooth and rough strains induced overproduction of TNFα and IL-6, as well as actin and tubulin rearrangements in the cytoskeleton. Moreover, OMVs from both strains inhibited PD-L1 expression in T-cells. These data revealed significant differences in OMVs derived from the rough and smooth <italic>Brucella</italic> strains, among which, the presence or absence of complete LPS appeared to be crucial to protect proteins contained within vesicles and to drive the immune response.</p>