Figure 3.

Activation of HIF and NFκB in uremia is blunted by inflammatory cytokine inhibition. NIH/3T3 cells expressing NFκB luciferase reporter were incubated with different concentrations of normal and uremic serum and 5 µM of the TNFα antagonist SPD304 (A) or 10 nM of echinomycin which is a HIF-1α inhibitor (B). Luciferase activity as reflection of NFκB activation in reporter cells was measured after serum and drug exposure for 24 h. NIH/3T3 cells expressing HIF luciferase reporter were incubated with different concentrations of normal and uremic serum and 2 µg/ml of neutralizing anti-IL6 monoclonal antibody (C) or 50 nM of QNZ which is an NFκB inhibitor (D). Luciferase activity as reflection of HIF activation in reporter cells was measured after serum exposure for 24 h in normoxia (21% O₂). Unpaired t test, n = 3. Experiments were done in triplicate, error bars represent SEM *p ≤ 0.05, **p < 0.01, ***p < 0.001. CS, normal serum; US, uremic serum.