DF increases HMEC-1 proliferation in 3D-collagen I gels. Cells (45 × 104 cells/well in 6-well plates) were seeded in collagen I (0.6 mg/mL) and treated with DF (25 μmol/L, every 3 days). After 6 days, the cells were counted. Each data point is the average of triplicate wells plus or minus SD. Relative HMEC-1 proliferation = 1.6-fol (± 0.08-fold). P less than 10−2 DF (25 μmol/L) versus C (A). (B) Confocal micrograms of HMEC-1 cells in 3D collagen I gels after 6 days in culture. HMEC-1 cells (105 cells/well) were seeded in collagen I (0.6 mg/mL) and treated with DF (5 or 25 μmol/L, every 3 days). At the end of treatment (6 days), the cells were fixed with 4% paraformaldehyde + 0.25% glutaraldehyde, permeabilized with 0.2% Triton X-100, and stained with rhodamine phalloidin (160 nmol/L). (C) Skeletonization by Image J of the images in panel A. (D) Quantitation of skeletonization by Image J of the confocal micrograms of HMEC-1 cells in 3D collagen I gels in (B) after 6 days in culture. Each experiment was performed in triplicate. Fold increase of skeletonized line = 3.5 (± 0.6; 5 μmol/L DF), and 3.5 (± 0.2; 25 μmol/L DF). *P (vs C) less than .02 (5 μmol/L); P (vs C) less than 10−4 (25 μmol/L DF). (E) Quantitation of skeletonization by Image J of the confocal micrograms of HMEC-1 cells in 3D collagen I gels treated with DF at multiple concentrations (0.25-25 μmol/L, every 3 days) after 6 days in culture. Each experiment was performed in triplicate.
