Antivascular effects of CD34-TRAIL+ cells. (A) NOD/SCID mice bearing subcutaneous tumor nodules 10 mm in diameter were randomly assigned to receive CD34-TRAIL+ cells, mock-transduced CD34+ cells (3 × 106 cells/mouse, intravenously), recombinant sTRAIL (500 μg/mouse, intraperitoneally), and control vehicle. Forty-eight hours after treatment, NOD/SCID mice were intravenously injected with 0.2 mL of sulfo-NHS-LC-biotin (5 mg/mL) to biotinylate tumor vasculature. Tumors were then excised, and biotinylated endothelium was revealed by AlexaFluor 488–streptavidin for confocal microscopy analysis (i-iv; 40×/1.0 NA oil objective) or sequentially incubated with HRP-streptavidin and 3,3′-diaminobenzidine for light microscopy analysis (v-viii; 20×/0.75 NA dry objective). After counterstaining with hematoxylin, sections were analyzed using ImageJ for quantification of vascular parameters. Representative confocal and histologic images of in vivo biotinylated mice receiving the different treatments are shown. (B) Endothelial area was calculated on whole tissue sections as (streptavidin-HRP stained area)/(total tissue area) × 100. *P < .001, compared with controls. #P < .001, compared with sTRAIL. (C) Vessel wall thickness was calculated on transversally oriented vessels. *P < .001, compared with controls. #P < .001, compared with sTRAIL.
