Background: H.
pylori are generally considered as extracellular organisms, with
exclusive colonization of the gastric milieu. Yet, several extra gastric
manifestations are associated with this infection. The aim of the
present study was to investigate the feasibility of toxin transfer by
extracellular vesicles, from bacterial and epithelial origins.
Methods: Tox-positive
H. pylori and its two cagA and vacA mutant strains were used to produce
bacterial vesicles (BVs) and to infect AGS cells. The produced BVs and
the infected cell vesicles (ICVs) were collected by ultracentrifugation
and evaluated by western blotting, DLS and electron microscopy. These
two sets of vesicles were applied to a second set of recipient AGS
cells, in which the acellular transfer of toxins, IL-8 production and
downstream morphologic changes were assessed, by western blotting, ELISA
and light microscopy, respectively.
Results: The
BVs were positive for H. pylori membrane markers (BabA and UreB), VacA
and CagA toxins, except for from the corresponding mutant strains. The
ICVs were larger in size and positive for bacterial markers, as well as
epithelial markers of CD9, LGR5, but negative for nuclear (Ki76) or
cytoplasmic (β-actin) markers. Bacteria-independent transfer of CagA and
VacA into the recipient cells occurred upon treatment of cells with BVs
and ICVs, followed by cellular vacuolation and elongation. IL-8
production was induced in recipient AGS cells, treated with BVs (1279.4 ±
19.79 pg/106 cells), early (8 h, 1171.4 ± 11.31 pg/106 cells) and late (48 h, 965.4 ± 36.77 pg/106 cells) ICVs (P < 0.0001).
Conclusion: Our
data indicates that ICVs, with mixed bacterial and epithelial
constituents, similar to BVs, are capable of transferring bacterial
toxins into the recipient cells, inducing IL-8 production and subsequent
morphologic changes, in an acellular manner.