Fig. 4

R848 induces neutrophil infiltration of peripheral and central tissues, but depletion of monocytes and B lymphocytes. Male, CD-1 mice received an intraperitoneal injection of R848 (200 μg, dissolved in DMSO and sterile saline) or control solution (DMSO diluted in sterile saline), together with an intravenous injection of nafamostat (3 mg/kg) or vehicle (sterile saline). Fresh liver was collected 6-h post-challenge and a subset of each group was cut at 12 μm thickness (n = 5/group). Immunostaining for neutrophils (MBS+), macrophages (Iba-1+) and B lymphocytes (B220+) was performed to determine leukocyte density. Liver neutrophils (A) and macrophages (B), lung neutrophils (C), macrophages (D) and B lymphocytes (E), and brain neutrophils (F) and microglia (G) were quantified, blinded, and presented as cells per mm². Naïve animals were included to establish baseline (dotted line). Data presented as mean ± SEM and analysed by two-way ANOVA, ****p < 0.0001, **p < 0.01 main effect, ^#p < 0.05, ^##p < 0.01, ^###p < 0.001, ^####p < 0.0001 control vs. R848 with Sidak’s post hoc test