Effect of hypoxia on the expression and/or phosphorylation of components of the canonical NF-κB signaling pathway in MDMs: regulation by HIF-1α and -2α. (A) Fold induction (hypoxia, 0.1% O2/normoxia, 20.9% O2) of mRNA levels for individual NF-κB signaling proteins in primary human MDMs. The contribution of both HIFs-1 and -2 to the regulation of many of these genes was also assessed using siRNA to knock down the expression of each α subunit in MDMs. (B) Effect of normoxia (N) or hypoxia (0.1% O2; H) for 18 hours on the expression of IKK-β and p65 mRNA by murine BMDMs from wild-type or HIF-1–deficient mice. *P < .05 compared with corresponding normoxic group. +P < .05 compared with macrophages from wild-type mice exposed to hypoxia. (C) Immunoblots showing the effect of exposure to normoxia or hypoxia (0.1% O2) for 18 hours on the levels of total and phosphorylated IKK-β and p65/RelA in primary human MDMs. Vertical lines have been inserted to indicate repositioned lanes from the same gel. (D) Effects of normoxic (N) or hypoxic (0.5% O2; H) culture on the level of total or phosphorylated p65 protein in control (+/−) or HIF-2α–deficient murine BMDMs. Similar results were obtained using wild-type or HIF-1α–deficient murine BMDMs (data not shown).
