Novel Pycard gene polymorphism impairs Nlpr3 inflammasome-induced IL-1β production in mice selected for low inflammatory response

A SNP based linkage study in mouse lines phenotypically selected for maximal (AIRmax) or minimal (AIRmin) acute inflammatory response (AIR), mapped a major locus, Inflammatory response modulator 1 (Irm1) at 128Mb (3.5Mb interval) in chr 7 controlling AIR, measured by leukocyte and IL-6 levels in exudates and IL-1β production by circulating leukocytes after Nlpr3 inflammasome activation. Sequencing of this region in mice with extreme high or low IL-1β levels revealed 14 SNPs between the two groups, narrowing the locus interval to 420Kb. Candidate genes at Irm1 include Pycard with an undescribed exon 3 (C/T) mutation leading to E19K substitution at the pyrin domain, and Itgam, Rgs10 and BC017158 with intronic SNPs. In Pycard, the C allele was fixed in high responder AIRmax mice whereas the C and T alleles frequencies were 39% and 61%, respectively in AIRmin. We then investigated the effect of this novel Pycard SNP in inflammation phenotypes.Methods AIRmin mice bearing the 3 genotypes at Pycard: CC, CT, TT were produced by genotype-assisted mating. Inflammatory response was measured in AIRmax and in the 3 AIRmin sublines by the number of infiltrating cells and IL-6 concentration in the 24h exudate induced by sc Biogel P-100 bead injection and ex vivo IL-1β production by circulating leukocytes after E coli LPS (1 ug) and ATP (5mM) activation.Results IL-1β levels were similar in AIRmaxCC (4.5-±0.4 ng/ml) and AIRminCC (3.4±2.4 ng/ml) whereas AIRminCT produced 0.3±0.5 and AIRminTT <0.05 ng/ml IL-1β. Leukocyte influx and IL-6 levels in inflammatory exudates were not affected.Conclusion The E19K substitution in Pycard causes a negative effect in inflammasome activation for IL-1β production, without interfering in other inflammation phenotypes.

Slc11a1 (Nramp1) alleles interact with acute inflammation loci to modulate wound-healing traits in mice.

Lines of mice were obtained by selective breeding for maximum (AIRmax) or minimum (AIRmin) acute inflammation. They present distinct neutrophil influx and show frequency disequilibrium of the solute carrier family 11a member 1 (Slc11a1) alleles. This gene is involved in ion transport at the endosomes within macrophages and neutrophils, interfering in their activation. Homozygous AIRmax and AIRmin sublines for the Slc11a1 gene were produced to examine the interaction of this gene with the acute inflammatory loci. The present work investigated wound-healing traits in AIRmax and AIRmin mice, in F(1) and F(2) intercrosses, and in Slc11a1 sublines. Two-millimeter ear punches were made in the mice and hole closure was measured during 40 days. AIRmax mice demonstrated significant tissue repair while AIRmin mice did not. Significant differences between the responses of male and female mice were also observed. Wound-healing traits demonstrated a correlation with neutrophil influx in F(2) populations. AIRmax( SS )showed higher ear-wound closure than AIRmax( RR ) mice, suggesting that the Slc11a1 S allele favored ear tissue repair. QTL analysis has detected two inflammatory loci modulating ear wound healing on chromosomes 1 and 14. These results suggest the involvement of the acute inflammation modifier QTL in the wound-healing phenotype.