HISA big data in biomedicine and healthcare 2013 conference

Additional file 5. Biofilm formation by the S. suis serotype 2 (S2) and serotype 9 (S9) wild-type and agI/II -deficient mutant strains in the absence of porcine fibrinogen. Biofilm formation capacity was quantified after 24 h of incubation at 37 °C in the absence of porcine fibrinogen. Data represent the mean ± SEM from at least three independent experiments

Proposed model of <i>S. suis</i> recognition by DCs.

<p>(A) The release of IL-1β, IL-6, IL-10, IL-23 and TNF-α is TLR2-dependent. TLR2 is also involved in the surface expression of MHC-II and CD86. (B) Other TLRs would also be implicated in the release of IL-1β, IL-6, IL-10, IL-23 and TNF-α. (C) TLR3 might be involved in the MyD88-independent production of CXCL10 and expression of CD86. (D) Collaboration between TLR2 and TLR9 is involved in the production of IL-12p70 and CXCL10 and the expression of CD40. (E) Collaboration among TLR2 and NOD2, with a minor contribution of TLR4, is involved in the release of CXCL1. (F) NOD2 also contributes to the release of IL-23. Recognition of <i>S. suis</i> peptidoglycan (PG) might be involved in NOD2 activation.</p

CD40 expression by DCs in response to <i>S. suis</i> depends on both TLR2 and TLR9.

<p>WT DCs and TLR2^−/− DCs (10⁶ cells/ml) pre-treated or not with an antagonist for TLR9 (ODN2088; 5 µM), were stimulated with <i>S. suis</i> parental strain 31533 (10⁶ CFU/ml) for 16 h. Non-stimulated cells served as negative control (C-). For comparative purposes, MyD88^−/− DCs were also included. Twenty thousand gated events were acquired per sample. Quadrants were drawn based on FITC- and PE-control stains and were plotted on logarithmic scales. Histograms were obtained by gating cells based on positive CD11c staining. * P<0.05 denotes values that are significantly lower than those obtained with WT DCs.</p

Surface expression of co-stimulatory molecules by DCs in response to <i>S. suis</i>.

<p>WT, MyD88^−/−, and TLR2^−/− DCs (10⁶ cells/ml) were stimulated with S. <i>suis</i> (10⁶ CFU/ml) for 16 h. Non-stimulated cells served as negative control (C−). (A) Percentage of CD40 positive cells. (B) Percentage of CD86^high positive cells. (C) Percentage of MHC-II^high positive cells. Twenty thousand gated events were acquired per sample. Quadrants were drawn based on FITC- and PE-control stains and were plotted on logarithmic scales. CD40, CD86 and MHC-II histograms were obtained by gating cells based on positive CD11c staining. * P<0.05 denotes values that are significantly lower than those obtained with WT DCs.</p

Immune Receptors Involved in <em>Streptococcus suis</em> Recognition by Dendritic Cells

<div><p><em>Streptococcus suis</em> is an important swine pathogen and an emerging zoonotic agent of septicemia and meningitis. Knowledge on host immune responses towards <em>S. suis</em>, and strategies used by this pathogen for subversion of these responses is scarce. The objective of this study was to identify the immune receptors involved in <em>S. suis</em> recognition by dendritic cells (DCs). Production of cytokines and expression of co-stimulatory molecules by DCs were shown to strongly rely on MyD88-dependent signaling pathways, suggesting that DCs recognize <em>S. suis</em> and become activated mostly through Toll-like receptor (TLR) signaling. Supporting this fact, TLR2^−/− DCs were severely impaired in the release of several cytokines and the surface expression of CD86 and MHC-II. The release of IL-12p70 and CXC10, and the expression of CD40 were found to depend on signaling by both TLR2 and TLR9. The release of IL-23 and CXCL1 were partially dependent on NOD2. Finally, despite the fact that MyD88 signaling was crucial for DC activation and maturation, MyD88-dependent pathways were not implicated in <em>S. suis</em> internalization by DCs. This first study on receptors involved in DC activation by <em>S. suis</em> suggests a major involvement of MyD88 signaling pathways, mainly (but not exclusively) through TLR2. A multimodal recognition involving a combination of different receptors seems essential for DC effective response to <em>S. suis</em>.</p> </div

Bacterial strains and plasmids used in this study.

<p>LTA; lipoteichoic acid, PG; peptidoglycan.</p

Oligonucleotide primers used in this study for construction of in-frame deletion mutants.

a<p>Oligonucleotide primers were from Invitrogen.</p

Effect of MyD88 deficiency on NF-κB expression by <i>S. suis</i> infected-DCs.

<p>WT DCs or MyD88^−/− DCs were incubated with the parental strain 31533, the non-encapsulated mutant B218 or the cell wall mutant Δ<i>dltA</i>/Δ<i>pgdA</i> strain (10⁶ CFU/ml). After a bacterial-cell contact of 8 h, cells were fixed and labeled with an antibody specific for NF-κB p65 (Alexa-Fluor 488, green) and analyzed by confocal microscopy.</p

Cytokine production by DCs in response to <i>S. suis</i>.

<p>WT, MyD88^−/−, and TLR2^−/− DCs (10⁶ cells/ml) were stimulated by different <i>S. suis</i> strains (10⁶ CFU/ml) for 16 h. Non-stimulated cells served as negative control (C−). Sample dilutions giving optical density readings in the linear portion of the ELISA standard curves were used to quantify cytokine levels. * P<0.05 denotes values that are significantly lower than those obtained with WT DCs.</p

Effect of MyD88 deficiency on the capacity of DCs to internalize <i>S. suis</i>.

<p>Bacteria (10⁶ CFU/ml) were pre-opsonized with 20% complete normal mouse serum for 30 min prior to incubation with DCs (10⁶ cells/ml). Phagocytosis was left to proceed for 2 h before antibiotics were directly added into the wells for 1 h to kill extracellular bacteria. Viable intracellular bacteria were determined by quantitative plating of serial dilutions of the lysates onto THB agar. * P<0.05 denotes values that are significantly different from those obtained with the <i>S. suis</i> parental strain 31533.</p