Extended Data Figure 4: EC numbers, proliferation and type H vessels in Notch mutants. | Nature

Extended Data Figure 4: EC numbers, proliferation and type H vessels in Notch mutants.

From: Endothelial Notch activity promotes angiogenesis and osteogenesis in bone

Extended Data Figure 4

a, b, Representative flow-cytometric graph plots showing the quantification of total and EdU-labelled endothelial cells from RbpjiΔEC (a) or Fbxw7iΔEC (b) mutant bones or corresponding littermate controls, as indicated. CD31+ CD45 Ter119 endothelial cells were substantially reduced in RbpjiΔEC mutants (a). Conversely, endothelial cell number and proliferation were increased after inactivation of Fbxw7 (b). c, Maximum intensity projections of EdU-labelled (green fluorescence) tibial metaphysis showing proliferating cells in 4-week-old Notch gain-of-function (Fbxw7iΔEC) and loss-of-function (RbpjiΔEC) mutants and corresponding littermate controls. Endothelial cells were visualized by Emcn immunostaining (red) and nuclei by DAPI (blue). d, Confocal images of Emcn (red) immunostained endothelial distal columns and arches next to growth plate chondrocytes (ch) highlighting the distribution of filopodia (arrowheads). Note that filopodia were directed towards chondrocytes in controls whereas directionality was compromised in RbpjiΔEC mutants. e, Quantitation of filopodia indicating higher variability in RbpjiΔEC samples (n = 6 mice from 4 independent litters). Error bars, ± s.e.m. P values, two-tailed unpaired t-test. f, Quantitative analysis of type H vessels in 4-week-old Fbxw7iΔEC mutants and littermate controls (n = 6 mice from 4 independent litters). Error bars, ± s.e.m. P values, two-tailed unpaired t-test. g, h, Maximum intensity projections of CD31 (green) and Endomucin (red) immunostained vessels in the metaphysis of 4-week-old RbpjiΔEC (g) and Fbxw7iΔEC mutants (h). Note loss of type H vessels and decreased CD31 (green) staining in the RbpjiΔEC metaphysis, while CD31+ vessels were extended after endothelial-cell-specific inactivation of Fbxw7. Arrows indicate arteries.

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