Chemical inhibition of NF-κB abrogates PEDF antiangiogenic function. (A-B) Endothelial cell (EC) apoptosis shown in HMVECs treated with the indicated combinations of bFGF (10 ng/mL) and PEDF (10nM), with or without BMS-345541. (A) Representative images of apoptosis detected by in situ TUNEL (green), counterstained with propidium iodide (red). (B) Quantitative analysis of the data shown in panel A. Statistical significance was determined by a 1-way analysis of variance. (C) Representative images of mouse corneal angiogenesis induced by bFGF and blocked by PEDF, with or without BMS-345541. (D) Quantitative analysis of the experiment in panel C. Angiogenesis is scored as positive corneas of total implanted or as clockwork vascular area (percentage of bFGF control). P values (determined by Fisher Exact test) are shown where significant. (E-F) Angiogenesis was induced in Matrigel plugs with bFGF (250 ng/mL) and blocked by PEDF with or without BMS-345541. Plugs were sectioned and stained for CD31 (red). Images were viewed with a Nikon Eclipse TE 2000-U microscope with a 10×/0.45 objective and Fluoromount G imaging medium (Southern Biotech), and captured with a Nikon LH-M100 C-1 camera. Metamorph, Adobe Photoshop CS3 Extended Version 10.01, and Corel Photo-Paint 10 software were used. (F) Quantification of the experiment in panel E. P values are shown. (G) DIVAA assay: angioreactors containing Matrigel with indicated compounds were implanted in the flanks of nude mice for 13 days; ECs were visualized by exposure with fluorescein isothiocyanate-lectin and quantified by fluorimetry. P values (determined by analysis of variance) are shown.