Fig. 1

Rac1 haploinsufficiency rescues arthritis and inflammatory signaling in Pggt1b^Δ/Δ mice. a Left, Western blots showing steady-state levels of GTP-bound and total Rac1 in BM macrophages isolated from Pggt1b^Δ/+, Pggt1b^Δ/Δ, and littermate Rac1^Δ/+Pggt1b^Δ/Δ mice. Actin was used as a loading control. Right, Bar graphs showing mean Rac1-GTP levels determined by densitometry (n = 2 per genotype). b Synovitis and erosion score in joints of 12-week-old Pggt1b^+/+ (n = 4), Pggt1b^Δ/Δ (n = 12), and Rac1^Δ/+Pggt1b^Δ/Δ (n = 9) mice. c Cytokine concentrations, 8 h after LPS (10 ng/ml) stimulation, in medium of primary bone marrow (BM) macrophages isolated from Pggt1b^+/+ (n = 3), Pggt1b^Δ/Δ (n = 4), and Rac1^Δ/+Pggt1b^Δ/Δ (n = 3) mice. d Western blots showing levels of mature Il-1β and caspase-1 in supernatants (Sup), and pro-Il-1β and pro-caspase-1 in lysates (Lys) of LPS (200 ng/ml) stimulated BM macrophages; tubulin in lysates was used as a loading control. The antibiotic nigericin (28 mM) was used as a positive control for inflammasome-mediated caspase-1 activation and Il-1β production. e Western blot showing levels of Mmp13 in medium of LPS-stimulated BM macrophages; Actin in lysates was used as a loading control. f Western blots showing phosphorylated (p) and total levels of intracellular signaling mediators in lysates of BM macrophages isolated 0, 15, and 30 min after LPS stimulation. g Concentration of Il-1β in medium of LPS-stimulated BM macrophages (n = 3/genotype) that had been pre-incubated for 1 h with inhibitors of p38 (SB203580; 1 and 5 µM) and ROS (DPI; 500 nM and 5 µM). For c–e, g, similar results were observed in two to three independent experiments. Error bars presented as s.e.m. when n is equal to or more than three. Significance between groups were calculated with two tailed Student’s t test (c, g) and one-way ANOVA with Tukey’s post hoc test (b). n.s. not significant, *P < 0.05, **P < 0.01, ***P < 0.001