Fig. 2.

Endothelial Gab1 deficiency leads to defective ischemia/VEGF-induced angiogenic responses. (A) Histological staining analyses of H&E, COX, and NADH for ischemic and nonischemic anterior tibial skeletal muscles from EGKO and control mice. Solid arrows in the ischemic muscle section from control mice show inflammatory cell infiltration, and empty arrows indicate the centralized nuclei in regenerating muscle cells. (B) Immunohistochemical analysis of ischemic and nonischemic anterior tibial skeletal muscles from EGKO and control mice, using anti-mouse CD31 antibody. Arrows indicate CD31-positive (brown) capillaries. Microvascular density was quantified as CD31-positive area relative to the entire area. The results represent the mean (± SEM) of six sections from three animals. **P < 0.01 compared with control. (C) Immunoblot analysis of VEGF expression in ischemic and nonischemic anterior tibial skeletal muscles. GAPDH was used as loading control. (D) Representative results of Matrigel implant assay. Matrigel containing VEGF (200 ng/mL) were injected into the abdominal s.c. tissues (300 μL per mouse). After 14 d, Matrigel plugs were harvested from EGKO and control mice 20 min after injection of FITC-dextran into tail veins. Upper: Matrigel plug photographs were taken under natural light. (Insets: Confocal FITC images of microvasculature in implanted Matrigel plugs.) Lower: Sections of Matrigel stained with CD31. (E) Representative results of ring assay of thoracic aortae isolated from EGKO and control mice. Thoracic aortic rings were embedded in Matrigel containing VEGF (20 ng/mL), incubated for 6 d, and photographed every 3 d. (Scale bars: A and B, 50 μm; D, 100 μm.)