Figure 4

Effect of oxygen normalization on HIF target mRNA expression in human endothelial cells. Human microvascular endothelial cells were exposed as described in Figure 1. Total RNA was extracted and the amount of HIF-1α (A), VEGF (B), Glut-1 (C), and iNOS (D) transcripts were analyzed by quantitative real-time RT-PCR as described in Materials and Methods. In the graphs (bottom panels), all values are expressed relative to the HPRT gene expression for each condition. Acute hypoxia (2.5% 0₂; 4 hours- "AH") was used as a positive control for the induction of transcripts. Data in graphs represent the mean +/- S.E.M. from three separate experiments with a sample size of 3–5 with representative gels above each graph. Asterisk, P < 0.05; double asterisk, P < 0.01; triple asterisk, P < 0.001 as compared to cells incubated in three weeks of normoxia (21%0₂) (N) (dashed line).