Figure 1.
Figure 1. Intravital imaging reveals a marginated pool of Ly6Chigh and Ly6Clow monocytes in BM vasculature. (A) Representative 3-dimensional (3D) 2-photon laser scanning microscopy (3D-TPLSM) pictures showing medullar monocytes in the parenchymal and vascular BM compartments of the skull in MacBlue×Cx3cr1gfp/+ mice (original magnification ×573). (B) Representative tracks of vascular monocytes displaying arrested (blue; original magnification ×620), crawling (pink; original magnification ×546), and rolling (green; original magnification ×300) behaviors. Vasculature is labeled with 2-MDa Rhodamin-Dextran (red), and bone matrix is identified by SHG (blue). (C) Pie graphs represent the relative proportion of the different monocyte behaviors in each mouse strain (percent ± standard deviation [SD] from 3 to 5 independent experiments are indicated). (D) Relative frequency distribution of vascular monocyte mean velocity (data are pooled from at least 3 independent experiments; percent indicates the proportion of cells faster than 6 µm/min for each mouse strain, and Kruskal-Wallis with Dunn’s multicomparisons test is performed. Statistical significance between the 2 mutants and WT mice is indicated). (E) Scheme illustrates cell behavior and intravascular staining in the BM sinusoids. Representative dot plot showing CD45 intravascular staining (red) overlaid with noninjected control mouse (black) in the blood and the BM. (F) Quantification of the blood/tissue partitioning of monocyte subsets in the different mouse strains. (G) Graphs represent the ratio of intravascular medullar monocytes in 1 thighbone to the total number of circulating monocytes. Black bars represent mean ± standard error of the mean (SEM). (Mice are pooled from at least 3 independent experiments; median is indicated in black, and Kruskal-Wallis with Dunn’s multicomparison test is performed.) (H) Bar graphs show the respective marker expression (as mean fluorescence intensity [MFI]) on intravascular BM monocytes (CD45+ cells after in vivo staining), parenchymal BM monocytes (CD45− after in vivo staining), and blood monocytes (CD45+ cells from the blood) in WT mice. (Bars represent mean ± SEM. n = 12-14 mice out of 2 to 3 independent experiments; Kruskal-Wallis with Dunn’s multicomparisons test is performed.) *P < .05, **P < .01, ***P < .001, ****P < .0001.

Intravital imaging reveals a marginated pool of Ly6Chighand Ly6Clowmonocytes in BM vasculature. (A) Representative 3-dimensional (3D) 2-photon laser scanning microscopy (3D-TPLSM) pictures showing medullar monocytes in the parenchymal and vascular BM compartments of the skull in MacBlue×Cx3cr1gfp/+ mice (original magnification ×573). (B) Representative tracks of vascular monocytes displaying arrested (blue; original magnification ×620), crawling (pink; original magnification ×546), and rolling (green; original magnification ×300) behaviors. Vasculature is labeled with 2-MDa Rhodamin-Dextran (red), and bone matrix is identified by SHG (blue). (C) Pie graphs represent the relative proportion of the different monocyte behaviors in each mouse strain (percent ± standard deviation [SD] from 3 to 5 independent experiments are indicated). (D) Relative frequency distribution of vascular monocyte mean velocity (data are pooled from at least 3 independent experiments; percent indicates the proportion of cells faster than 6 µm/min for each mouse strain, and Kruskal-Wallis with Dunn’s multicomparisons test is performed. Statistical significance between the 2 mutants and WT mice is indicated). (E) Scheme illustrates cell behavior and intravascular staining in the BM sinusoids. Representative dot plot showing CD45 intravascular staining (red) overlaid with noninjected control mouse (black) in the blood and the BM. (F) Quantification of the blood/tissue partitioning of monocyte subsets in the different mouse strains. (G) Graphs represent the ratio of intravascular medullar monocytes in 1 thighbone to the total number of circulating monocytes. Black bars represent mean ± standard error of the mean (SEM). (Mice are pooled from at least 3 independent experiments; median is indicated in black, and Kruskal-Wallis with Dunn’s multicomparison test is performed.) (H) Bar graphs show the respective marker expression (as mean fluorescence intensity [MFI]) on intravascular BM monocytes (CD45+ cells after in vivo staining), parenchymal BM monocytes (CD45− after in vivo staining), and blood monocytes (CD45+ cells from the blood) in WT mice. (Bars represent mean ± SEM. n = 12-14 mice out of 2 to 3 independent experiments; Kruskal-Wallis with Dunn’s multicomparisons test is performed.) *P < .05, **P < .01, ***P < .001, ****P < .0001.

Close Modal
This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal