Introduction Deletion of specific NF-κB subunits in mice alters the outcome of Helicobacter felisinfection. Nfkb1^-/- mice developed more severe gastric atrophy than wild-type (WT) mice 6 weeks after H. felisinfection, whereas Nfkb2^-/- mice were protected from this pathology. The mechanisms underlying these outcomes remain unclear, but gastric Il1btranscript abundance was increased in H. felisinfected Nfkb1^-/- mice relative to WT, and polymorphisms at the IL1B locus have also been associated with gastric cancer in humans. Il1btranscription is regulated by classical pathway NF-κB signalling. IL-1β secretion also requires the formation of inflammasome complexes, which form following intracellular pathogen recognition, and lead to the autocatalysis of pro-caspase 1 and subsequent cleavage of IL-1β. We hypothesised that inflammasome signalling was altered in mice with abrogated NF-κB signalling and that this influenced H. felisinduced pathology. We therefore investigated inflammasome mediated signalling in bone marrow derived dendritic cells (BMDCs) from mice lacking specific NF-κB subunits.

Method Cells were harvested from C57BL/6, Nfkb1^-/-,Nfkb2^-/- and c-Rel^-/- bone marrow. Dendritic cells were differentiation using 20 ng/ml GM-CSF for 7 days. BMDCs and WT derived gastric epithelial organoids were primed with 20 ng/ml LPS and exposed to 300 μg/ml silica, 5 mM ATP, H. pylori(ATCC 53726) or H. felis(ATCC 49179) (MOI 1:100) with or without a pan-caspase inhibitor (Z-VAD-fmk), an inhibitor of NADPH oxidase (APDC) or 50 mM KCl. Secreted IL-1β and TNF concentrations were measured by ELISA.

Results LPS alone induced TNF, but not Il-1β, secretion in all genotypes of BMDCs. Both silica and ATP induced IL-1β secretion in WT BMDCs pre-stimulated with LPS (750 ± 65 and 530 ± 77 pg/ml). BMDCs derived from NF-κB deficient mice secreted similar amounts of Il-1β in response to these stimuli. Inflammasome inhibitors returned IL-1β secretion to unstimulated levels. Gastric epithelial organoid cultures treated with LPS and silica did not secrete either IL-1β or TNF. Following exposure to H. felisor H. pylori, Nfkb1^-/- BMDCs, but not other genotypes, exhibited a 2.6 fold increase in IL-1β secretion compared to untreated cells or cells stimulated with LPS. This was abrogated by Z-VAD-fmk.

Conclusion These data identify potent inflammasome activation in NF-κB deficient BMDCs. H. pyloriand H. felisalso weakly stimulated inflammasome activation in NFkb1-/-BMDCs. This mechanism may contribute to the more severe gastric phenotype that is observed in NFkb1^-/- mice in vivofollowing H. felisinfection. Further studies are required to identify how NF-κB1 deletion influences inflammasome formation, and whether altered IL-1β transcription is involved.

Disclosure of interest None Declared.