EGFL7 associates with the ECM and affects ECs in vitro and in vivo. (A) Human fetal brain tissue slices containing venous leptomeningeal vessels were stained for EGFL7, collagen IV and counterstained with TO-PRO-3. Bar represents 100 µm. (B) Recombinant His-tagged EGFL7 was added onto a microtiter plate coated with vitronectin, fibronectin, collagen I, collagen IV, laminin-111, or anti-His antibody and the interaction between the purified proteins was measured by ELISA. Values were normalized to BSA. (C) Dishes were coated with EGFL7 or vitronectin and were covered with 50 000 HUVECs, which adhered to both recombinant proteins. Adhesion to the negative control BSA was subtracted from each value. (D) Growth-factor–reduced Matrigel was mixed either with phosphate-buffered saline or recombinant EGFL7 and was injected subcutaneously into mice. ECs that infiltrated into the plugs were detected by staining with biotinylated isolectin B4. The number of isolectin B4⁺ cells was counted in 5 nonoverlapping microscopic fields per plug (data presented as mean ± standard error of the mean [SEM], n = 12, *P = .005). The bar represents 50 µm. (E) Likewise, the average length of the CD31⁺ vessel-like structures (arrows) (ie, cavities surrounded by cells and cells arraying in line but without cavities) was quantified in these plugs (data presented as mean ± SEM, n = 12, *P < .05). The bar represents 50 µm.