Fig. 5. Effects of exogenous PDGF-BB and anti-PDGFRß neutralizing antibody (APB5) in the Matrigel plug assay in vivo. (A-C) PDGF-BB (1 µg/ml; A) or the anti-PDGFRß neutralizing antibody (APB5, 100 µg/ml; C) was mixed with to VEGF-A (200 ng/ml) and FGF-2 (1 µg/ml) in the Matrigel plug assay. (B) V/F treatment only. (Top row) H & E staining. Scale bars: 30 µm. Red arrows show mature blood vessels containing RBCs; orange arrowheads indicate vessels with extravascular RBCs. (Second and third rows) Immunohistochemistry. SMA staining (red) is shown in the second row, and merged images with PECAM1staining (green) are shown in the third row. White arrowheads indicate detached SMA⁺ cells, and arrows indicate PECAM1⁺ cells not covered by SMA⁺ cells. (Fourth row) Permeability assay using intravenously administrated dextran (green), merged with SMA (red) and PECAM1 (blue) images. Arrows indicate leakage of dextran. Scale bars: 20 µm. (Fifth row) Electron micrographs. Scales are indicated in each photograph. MC, mural cells; EC, endothelial cells; RBC, red blood cells; yellow arrows, tight junctions formed between endothelial cells. (D,E) Induction of new vessels and vessel-like structures in Matrigel plugs was quantified as in Fig. 1 but with a new category (orange bars) representing vessels with extravascular RBCs, which indicate hemorrhage. *P<0.05, **P<0.01 by Student's t-test. Corresponding figures for NG2 and desmin staining are shown in Fig. S3 in supplementary material.