Immunostaining and ultra-structural analysis of microvessels.

(A–B) Perlecan is expressed in the sub-endothelial BMs of normal brain capillaries and is absent in the mutant tissue. (C–D) Laminin-1 is present in both wild type and perlecan-null BMs. (E–F) SMA is expressed in the microvessels sprouting into the brain parenchyma in wild type E17.5 (E), as well as mutant brains (F). (G–I) Ultrastructural analysis of endothelial cells from E14.5 brain capillaries. Note the tight association between the endothelial cell (ec) and the directly adjacent cells (ac) in the wild type brain capillary (G). The BM is visible on the upper surface of the adjacent cells (arrows in G). In the mutant brains, gaps are evident between the endothelial cells and the adjacent cells and pericytes (p) (arrows in H and I). (J–L) Electron micrographs of E16.5 skin microvessels. Electron dense material (arrows) is deposited at the abluminar side and appears as a BM-like structure in the wild type (J) as well as in the perlecan-null vessels (K,L).Vessel lumen (VL). Bars: (A–F) 250 µm; (G–L) 400 nm.