FIGURE 2.

Regulation of TMPRSS6 by hypoxia, BIP, and DMOG is dependent on HIF-1α and HIF-2α. Hep3B cells were transfected with control siRNA, HIF-1α siRNA, HIF-2α siRNA or combined HIF-1α & HIF-2α siRNA followed by incubation either in hypoxia (0.5% O₂ for 48 h), or in the presence of 25 μm BIP or 0.5 mm DMOG for 16 h. A and B, quantitative real-time PCR showing TMPRSS6 and CA9 mRNA expression respectively (*, p < 0.0001 by two-way ANOVA with p < 0.01 for each siRNA using Bonferroni's post-test analysis, †, p < 0.01 relative to single siRNA treatments). mRNA expression levels are normalized to cyclophilin, and data points are averages of three independent biological replicates. C, immunoblot analysis of TMPRSS6, CA9, HIF-1α, and HIF-2α levels in Hep3B cells. β-Actin levels were measured to control for loading differences.