The prototypical NLRP3 and NLRC4 inflammasomes.

<p>(A) The NLRP3 inflammasome is activated by both bacterial (e.g., MDP, bacterial RNA, β-glucan), exogenous (e.g., silica, alum), and endogenous (e.g., uric acid cristals, ATP) stimuli. (B) The NLRC4 inflammasome is activated by flagellin in a TLR5-independent fashion.</p

Diagram representing the differential caspase-1/IL-1β activation pathways in monocytes and macrophages.

<p>Caspase-1 is constitutively activated in monocytes, and these cells release mature IL-1β after single stimulation with TLR ligands. IL-1β secretion is induced by endogenously-released ATP. In contrast, macrophages need a double stimulation: one stimulus (TLR-ligands) induces transcription, and a second stimulus (ATP) induces IL-1β secretion.</p

Susceptibility to In Vivo Experimental Models of Infection in Mice Deficient in IL-1β, IL-18, or Inflammasome Components.

<p>Lower (susceptibility): increased survival of the knock-out mice in the experimental model.</p><p>Higher (susceptibility): decreased survival of the knock-out mice in the experimental model.</p><p>ND, not done.</p

Additional file 1: Appendix 1. of An electronic trigger tool to optimise intravenous to oral antibiotic switch: a controlled, interrupted time series study

Electronic trigger tool algorithm to identify patients eligible for iv to oral switch. #high dose of a penicillin or cephalosporin only indicated for severe infection (e.g. endocarditis or meningitis) or an antibiotic class which is given only if no oral formulation is available (carbapenem). (JPEG 63 kb

The influence of 3MA on the ATP-dependent release of IL-1β.

<p>PBMCs were pre-incubated for 1 hour in the presence or absence of 3MA and were stimulated for 4 hours with LPS (10 ng/ml). After the stimulation, supernatants were discarded and refreshed with RPMI or with RPMI containing 1 mM ATP and cells were incubated for another 15 min. Data are shown as mean ± SEM of supernatant IL-1β levels obtained in 8 volunteers, **p<0.01.</p

Modulation of inflammatory cytokine production by autophagy inhibition.

<p>Freshly isolated human PBMCs were pre-incubated for 1 hour at 37°C in culture medium in the presence or absence of 3MA (10 mM) and afterwards stimulated with culture medium, LPS (10 ng/ml) or Pam3Cys (10 µg/ml). After 24 hours incubation, IL-1β (A), TNFα (B) and IL-10 (C) were measured in the supernatant by specific ELISA. Data are presented as means ± SEM of cells harvested from 15 volunteers, **p<0.01, ***p<0.001.</p

Assessment of LC3 I and II levels in PBMCs under starving conditions or pharmacological treatment with 3MA.

<p>Human PBMCs were pre-incubated for 1 hour at 37°C in either RPMI, starvation medium (EBSS) or EBSS with 3MA (10 mM) in which inhibitors of lysosomal fusion have been added: Ammonium chloride 20 mM and Leupeptine 100 µM. This was followed by 3 hours stimulation with culture medium, LPS (10 ng/ml) or Pam3Cys (10 µg/ml) prepared in the corresponding media (RPMI or EBSS). Cells were lysed and western blot of LC3 fractions I and II has been performed.</p

MAPK influence on the 3MA-dependent modulation of IL-1β and TNFα production.

<p>PBMC samples conditioned for 1 hour in either medium or in the presence of MAPK inhibitors: MEK inhibitor (5 µM), JNK inhibitor (20 µM) or p38 inhibitor (1 µM) were subjected to 1 hour treatment with RPMI or 3MA (10 mM), followed by stimulation with RPMI, LPS (10 ng/ml) or Pam3Cys (10 µg/ml). After 24 hours incubation, specific ELISA was performed to determine the level of IL-1β in response to LPS (A) and Pam3Cys (B) stimulations, same as for TNFα (panels C and D, respectively). Data from 6 volunteers are shown as mean ± SEM. (E) Western blot of total and phosphorylated p38 in cells pre-treated for 1 hour with RPMI or 3MA (10 mM) and subjected to 1 hour stimulation with RPMI, LPS (10 ng/ml) or Pam3Cys (10 µg/ml). (F) Quantification of the effect of 3MA on the phosphorylation of p38.</p

The effects of 3MA on transcription and processing of inflammatory cytokines.

<p>Cells pre-treated for 1 hour with culture medium with or without 3MA (10 mM) were stimulated for 4 hours with RPMI, LPS (10 ng/ml) or Pam3Cys (10 µg/ml). RT-PCR was performed and relative levels of IL-1β and TNFα (B) mRNA were determined in 4 volunteers. (C) Western blot of p35 caspase-1 after 1 hour pre-incubation with or without 3MA (10 mM in RPMI), followed by 2 hours stimulation with LPS (10 ng/ml). The picture is representative for results obtained from 6 volunteers.</p

The effects of starvation on mRNA levels of inflammatory cytokines.

<p>Cells pre-incubated for 2 hours in either starvation medium (Earle's Balanced Salt Solution) or RPMI were stimulated for 4 hours with medium, LPS (10 ng/ml) or Pam3Cys (10 µg/ml) prepared respectively in starvation medium or RPMI. Subsequently, RT-PCR was performed and IL-1β mRNA levels are depicted as mean ± SEM of cells harvested from 6 volunteers, *p<0.05 (A). RT-PCR results of IL-1β (B) and TNFα (C) mRNA levels in cells pre-incubated for 2 hours with RPMI, starvation medium or starvation medium and 3MA (10 mM) followed by 4 hours stimulation with RPMI or LPS (10 ng/ml). Results are shown as mean ± SEM of data obtained in 4 volunteers.</p