PAI -inhibition induces angiogenesis during HL-ischemic recovery via FGF-2- and VEGF-A–mediated pathways. (A-C) HL-ischemia–induced C57BL/6 mice were treated with the PAI-1 inhibitor or vehicle. Ischemic sections of PAI-1 inhibitor or vehicle-treated mice 3 days after the HL procedure were costained for F4/80 (A), F4/80 and VEGF-A or F4/80 and FGF-2 (B), or Gr-1 and VEGF-A or Gr-1 and FGF-2 (C). Nuclei were counterstained with DAPI (blue). Left panels are representative immunofluorescent images. Arrows indicate VEGF-A⁺, FGF-2⁺, F4/80⁺, or Gr-1⁺ cells. Right panel shows the quantification of the indicated cell populations per high-power field (HPF; n = 5/group for each experiment). (D-G) HL-ischemia–induced C57BL/6 mice were treated with the PAI-1 inhibitor and coinjected with neutralizing doses of anti–FGF-2, anti–VEGF-A, or anti-IgG control Abs (n = 4/group). (D-E) Ischemic muscle tissues from Ab-treated animals 14 days after the HL procedure were immunofluorescently costained for FGF-2/VWF and FGF-R1/VWF. Nuclei were counterstained with DAPI (blue staining). Arrows indicate FGF-2⁺/VWF⁺ and FGF-R1⁺/VWF⁺ cells (scale bars, 200 mm). Right panel shows the indicated cell populations quantified per HPF. (F) Left panel, ischemic muscle tissue sections were stained with H&E (scale bars, 200 mm). Right panel shows the quantification of necrotic areas in ischemic H&E-stained tissue sections. (G) Blood flow was determined at the indicated time points. (H) Ischemic muscle tissue sections stained with Abs against VWF antigen on day 14 were used to determine capillary density. Data represent means ± SEM. *P < .05.