Attachment and spreading of ECs on NG2. (A) NG2 binding to the EC surface. NG2 immunoreactivity is seen on the surfaces of MAEC incubated with soluble NG2/EC+, but not on untreated cells (control). DAPI was used as a nuclear counterstain. Scale bar, 20 μm. (B) EC attachment to NG2-coated surfaces. MAEC were tested for their ability to adhere to wells coated with Matrigel (30 μg/ml), type I collagen (30 μg/ml), and various concentrations of NG2/EC+ (1–100 μg/ml). Three high-power microscope fields were counted in each replicate well, and results were expressed as cells per field. Each bar represents the mean ± SD (n = 3). Statistically significant differences compared with the control are indicated (*p < 0.005; **p < 0.05). Insets show the stained cells in control (left) and NG2-coated (100 μg/ml) wells (right). Adherent cells in the right-hand panel represent roughly 90% of the input cells. Scale bar, 50 μm. (C) Spreading of ECs on NG2-coated surfaces. Staining with rhodamine-phalloidin was used to visualize F-actin organization in MAEC after adherence for 1 h to wells coated with BSA (control), type I collagen (30 μg/ml), NG2/EC+ (10 μg/ml), or NG2/EC- (10 μg/ml). Scale bar, 20 μm.