Figure 3
Effect of oxygen normalization on HIF-1α, HIF-2α, Glut-1, BNIP3, PAI-1, iNOS and VEGF protein expression in human endothelial cells. Human microvascular endothelial cells were exposed as described in Figure 1. Western blots were performed against HIF-1α (A), HIF-2α (B), Glut-1 (C), BNIP3 (D), PAI-1 (E), iNOS (F) on cellular lysates and ELISA was performed against VEGF (G) on cellular supernatants. Graphs indicate the densitometric analysis relative to the respective normoxic protein expression (dashed line) with a representative blot for each protein above each graph. Acute hypoxia (2.5%02; 4 hours) ("AH") was used as a positive control for the induction of proteins. Data in graphs represent the mean +/- S.E.M. from three separate experiments with a total sample size of 4–6. Asterisk, P < 0.05; double asterisk, P < 0.01; triple asterisk, P < 0.001 as compared to cells incubated in normoxia for three weeks.