Figure 4.
Figure 4. CCR2 and CX3CR1 control monocyte compartmentalization during inflammation. Blood/tissue partitioning of Ly6Chigh and Ly6Clow monocytes in the lungs (A-C) and the spleen (D-F) are determined by in vivo CD45 intravascular staining in the different mouse strains after intraperitoneal injection of 100 ng/kg LPS in WT, CCR2-, and CX3CR1-deficient mice. (A,D) Graphs represent mean ± SEM of the absolute number per milligram of tissue of intravascular (CD45+) monocytes. (B,E) Graphs represent mean ± SEM of the absolute number of parenchymal (CD45−) monocytes. (C,F) Pie graphs represent the relative proportion of parenchymal (dark) and intravascular (clear) monocytes in indicated mouse strains and tissue for all graphs, n = 6-14 mice per group and per time point from at least 2 independent experiments. Kruskal-Wallis with Dunn’s multicomparisons test is performed, and the significance for each time point compared with untreated mice [t0] is indicated in colored *. Significances between mouse strains are indicated by black * for each time point. Mann-Whitney test is performed). (G) Representative fluorescent wide field pictures of the inner side of the parietal sheet of the peritoneal membrane (green autofluorescence) at different time point after LPS injection in MacBlue×Cx3cr1gfp/+ and MacBlue×Cx3cr1gfp/gfp mice show ECFP+ monocytes (cyan) accumulation along the linea alba (dark area; original magnification ×20). (H) Quantification of monocyte accumulation along the linea alba in the different mouse strains (n = 3-8 mice in each group, Kruskal-Wallis with Dunn’s multicomparisons test is performed for each strain). *P < .05; **P < .01; ***P < .001; ****P < .0001. (I) Representative time-lapse in vivo 3D images of the peritoneal membrane vasculature in MacBlue×Cx3cr1gfp/+ shows monocyte accumulation, adhering to the endothelium at different time points after LPS injection (original magnification ×20). Vessels are labeled by Rhodamin-Dextran injection prior to imaging session (red). SHG shows conjunctive tissue of the peritoneal membrane. (J) Comparative 3D-TPLSM images showing monocyte accumulation in the peritoneal vasculature 4 hours after LPS injection in MacBlue×Cx3cr1gfp/+ (WT) and MacBlue×Cx3cr1gfp/gfp mice (Cx3cr1−/−) (original magnification ×106).

CCR2 and CX3CR1 control monocyte compartmentalization during inflammation. Blood/tissue partitioning of Ly6Chigh and Ly6Clow monocytes in the lungs (A-C) and the spleen (D-F) are determined by in vivo CD45 intravascular staining in the different mouse strains after intraperitoneal injection of 100 ng/kg LPS in WT, CCR2-, and CX3CR1-deficient mice. (A,D) Graphs represent mean ± SEM of the absolute number per milligram of tissue of intravascular (CD45+) monocytes. (B,E) Graphs represent mean ± SEM of the absolute number of parenchymal (CD45−) monocytes. (C,F) Pie graphs represent the relative proportion of parenchymal (dark) and intravascular (clear) monocytes in indicated mouse strains and tissue for all graphs, n = 6-14 mice per group and per time point from at least 2 independent experiments. Kruskal-Wallis with Dunn’s multicomparisons test is performed, and the significance for each time point compared with untreated mice [t0] is indicated in colored *. Significances between mouse strains are indicated by black * for each time point. Mann-Whitney test is performed). (G) Representative fluorescent wide field pictures of the inner side of the parietal sheet of the peritoneal membrane (green autofluorescence) at different time point after LPS injection in MacBlue×Cx3cr1gfp/+ and MacBlue×Cx3cr1gfp/gfp mice show ECFP+ monocytes (cyan) accumulation along the linea alba (dark area; original magnification ×20). (H) Quantification of monocyte accumulation along the linea alba in the different mouse strains (n = 3-8 mice in each group, Kruskal-Wallis with Dunn’s multicomparisons test is performed for each strain). *P < .05; **P < .01; ***P < .001; ****P < .0001. (I) Representative time-lapse in vivo 3D images of the peritoneal membrane vasculature in MacBlue×Cx3cr1gfp/+ shows monocyte accumulation, adhering to the endothelium at different time points after LPS injection (original magnification ×20). Vessels are labeled by Rhodamin-Dextran injection prior to imaging session (red). SHG shows conjunctive tissue of the peritoneal membrane. (J) Comparative 3D-TPLSM images showing monocyte accumulation in the peritoneal vasculature 4 hours after LPS injection in MacBlue×Cx3cr1gfp/+ (WT) and MacBlue×Cx3cr1gfp/gfp mice (Cx3cr1−/−) (original magnification ×106).

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