Extended Data Fig. 6: Establishment of CNS, peripheral and BBB signatures in vascular ECs from the developing fetal to the adult brain vasculature.

a, UMAP plots of the ECs from fetal brains in ascending fetal age ranging from gestational week 9 (444 ECs), gestational week 14.4 and 16.4 (3762 ECs), gestational week 15.5 (18,314 ECs) to gestational week 18 (3601 ECs); plots are colour-coded for fetal CNS signature (green) and fetal peripheral signature (yellow). b, Dotplot heatmap of fetal CNS, fetal peripheral, and BBB signatures’ expression in fetal brains of indicated fetal age and adult/control brains (temporal lobes). c, Dotplot heatmap of fetal CNS signature genes’ expression in fetal brains of indicated fetal age and adult/control brains (temporal lobes). d, Dotplot heatmaps of fetal CNS and BBB signatures expression at the level of AV specification for the indicated entities. Colour scale: red, high expression; blue, low expression, whereas the dot size represents the percentage expression within the indicated entity. e-j, Violin plots showing the expression of selected CNS specific and BBB marker genes in fetal brains of indicated fetal age and adult/control brains (TL). Markers shown were further validated by IF and/or IMC. k-t’₄ Mass cytometry (IMC) imaging of fetal brains of fetal age GW9 (k-l’₄), GW14 (m-n’₄), GW18 (o-p’₄), GW21 (q-r’₄) and adult/control brains (temporal lobe) (s-t’₄) tissue samples using metal-conjugated antibodies. Representative pseudocolor images of combined and individual channels of the different markers (white, overlap; yellow, CLDN5; cyan, GLUT1; grey, BSG; red, CD31; blue, DNA intercalator), Scale bars: 250 μm in overviews, 50 μm in zooms. Shown are 5 antibodies stained for (Supplementary Table 24). Arrowheads identify blood vessels ECs expressing the indicated markers in the different tissues. Arrows identify blood vessel ECs not expressing the indicated markers in the different tissues.