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

ROLE OF APOPTOSIS IN IL-17 PRODUCTION DURING CANDIDA
ALBICANS INFECTION
GUSTAVO FERNANDO DA SILVA QUIRINO (IC) (1); CARINA ZERBETTO
SEGATO (PG) (1); TACITO GRAMINHA CAMPOIS (PG) (1); RICARDO
SERGIO ALMEIDA (2); IONICE FELIPE(1)
(1) - Lab. de Imunologia Experimental, Depto. de Ciências Patológicas –
CCB/UEL; (2)–Lab. de Micologia Médica, Depto. de Microbiologia – CCB/UEL
Introduction: Citrobacter rodentium is one pathogen that targets mitochondria
and induces apoptosis, and blockade of apoptosis during C. rodentium infection
impairs the characteristic TH17 response (J. Leukoc.Biol.89, 1-12, 2011).
However, there are no evidences that IL-17 production against Candida
albicans is triggered by this pathway. Our group previously demonstrated that
C. albicans CR15 was able to induce apoptosis of macrophages in vitro
whereas C. albicans 577 was not. In order to clarify if apoptosis is involved in
IL-17 production, the profiles of cytokines production during C. albicans
infection were compared.
Methods and Results: Strains 577 and CR15 were cultivated in YPD medium
for 24h at 30°C, harvested, diluted to 1x10 7 ml -1 PBS and inoculated
intraperitoneally in Swiss mice (n=3 per group). After 0.5, 2, 6, 18 and 24 hours,
the peritoneal exsudate of the animals was collected with 2ml RPMI 1640
medium, following centrifugation. The supernatant was stored at -20°C to
analyze cytokine levels through ELISA method and the cell pellet was added to
coverslips in cell culture plates for 1h at 37°C, 5% CO2, to allow the adherence
of phagocytes. Groups infected for 0.5 and 2h had their cells stained with
Annexin-V-FITC, followed by 4%-paraformaldehyde fixation and then analyzed
under fluorescence Leica microscope, to qualitatively assess the induction of
apoptosis of phagocytic cells by C. albicans. In addition, all groups had their
cells fixed in methanol and stained with Giemsa, to assess the cell population in
the exsudate. Our results show that C. albicans CR15 was able to induce
apoptosis of phagocytic cells 30 minutes postinfection, whereas 577 was not.
CR15 induced peak levels of IL-1β (45.06 ± 19.9 pg/mL after 2h), IL-6 (33.6 ±
5.1 ng/mL after 24h), TGF-β (155.6 ± 31.21 pg/mL after 6h) and IL-17 (206.6 ±
61.9 pg/mL after 18h). On the other hand, we show that neither of cytokines that
precede IL-17 production were detected in C. albicans 577 infection, although it
has induced IFN-γ production (252.2 ± 124.4 pg/mL). The neutrophil and
macrophage population in the exsudate reaches a peak after 18h of infection
with both strains.
Conclusions: The results suggest that apoptosis of macrophages induced by
C. albicans CR15 could be involved in IL-17 production since this cytokine was