Nuclear translocation of thioredoxin following exposure of proliferating endothelial cells to HKa. (A) Cells were cultured in 10 cm² fibronectin-coated plates at a concentration of 3 × 10⁴ cells/mL and stimulated with 10 ng/mL bFGF and 50 nM HKa. At increasing intervals, cell extracts were prepared and separated using 15% SDS-PAGE, as described in “Materials and methods.” After transfer to PVDF, membranes were incubated with peroxidase-conjugated goat anti–mouse thioredoxin antibodies and developed using enhanced chemiluminescence. No thioredoxin translocation occurred in cells exposed to bFGF alone in the absence of HKa (not shown). (B) Experiments were conducted as described in panel A but HKa-exposed cells were incubated concurrently with 150 μg/mL GSH. No nuclear translocation of thioredoxin occurred under these conditions. (C) Experiments were performed as described in panel A but using cells cultured on type I collagen.