Bortezomib exerts double-edged effect on HUVECs, through HIF-1α and NF-kB. HUVECs were treated with bortezomib at 10 nM and 100 nM for 24 hours and then assessed for their proliferative activity (A) or for their ability to form capillaries on Matrigel (B) (original magnification ×10). Data are presented as the mean plus or minus standard error of the mean (SEM) from 3 independent experiments in duplicate in panel A; in panel B one experiment representative of 4 is depicted. (C) Quantification of total (dark column) and nuclear (gray column) HIF-1α expression as calculated by fluorescent pixel intensity at laser confocal microscopy. Data are presented as the mean plus or minus SEM from 20 cells derived from 3 different fields. (D) HIF-1α (green) and GLUT-1expression (red) were assessed in HUVECs treated with bortezomib at 10 nM and 100 nM by confocal analysis (original magnification ×60). Bortezomib up-regulated HIF-1α and NF-kB p65 expression as evaluated by Western blot analysis (E) and induced their nuclear accumulation as evaluated by confocal analysis through merge with DAPI (F) (original magnification ×60). **P < .001.