CX3CR1-dependent Ly6C^high-monocyte margination controls monocyte deployment during CLP-induced peritonitis. (A) Quantification by flow cytometry of BM, lung, and spleen-intravascular Ly6C^high, Ly6C^low monocytes, and neutrophils in Sham and CLP-operated after in vivo CD45 intravascular staining (bars represent mean ± SEM from 7 to 11 different mice per time point out of 3 to 4 independent experiments; Mann-Whitney test is performed). *P < .05; **P < .01; ***P < .001; **** P < .0001. (B) 3D-TPLSM images of the skull BM from MacBlue×Cx3cr1^gfp/+ 4 hours after CLP treated or not with the F1 (50 µg injected intraperitoneally) (original magnification ×440). Representative track paths are represented in green. (C) Quantification of monocyte mean velocity and straightness in the vascular lumen of the BM sinusoids from MacBlue×Cx3cr1^gfp/+ mice treated or not with CX3CR1 antagonism (F1) 4 hours after CLP (median is indicated in red. Data are pooled from different movies out of 3 different mice. Mann-Whitney test is performed; ****P < .0001). (D) In vivo 3D-TPLSM image of the peritoneal vasculature showing ECFP⁺ monocyte adherence to endothelium 4 hours after CLP in the presence or not of F1 (original magnification ×230). Representative track paths are represented in green (see supplemental Video 8). (E) Scatter plots show the distribution of the absolute number of Ly6C^high and Ly6C^low monocytes in the lung versus the peritoneal cavity in Sham, CLP, and CLP treated with F1 4 hours after operation. Statistical differences in the distribution have been measured by a 2-way ANOVA with Bonferroni’s multiple comparison tests.