Fig. 8.

Apoptosis increases in fetal cells transfected with a dominant-negative mutant IκBα and exposed to hyperoxia. Control cells, cells treated with lipofectamine only (L), and cells transfected with empty vector (V), WT IκBα, or Y42F were exposed to room air, H₂O₂ (8 h), or hyperoxia (24 h). Cell lysate or genomic DNA was collected and subjected to caspase-3 activity assay and Western blot analysis or DNA laddering assay, respectively. A: caspase-3 activity increases in fetal cells transfected with Y42F and exposed to hyperoxia. Left: fold change in caspase-3 activity in cells exposed to hyperoxia compared with respective controls. Air, fetal cells exposed to room air; O₂, fetal cells exposed to 24 h of hyperoxia; L, lipofectamine-treated cells exposed to room air; L-O₂, lipofectamine-treated cells exposed to 24 h of hyperoxia; V, empty vector-transfected cells exposed to room air; V- O₂, empty vector-transfected cells exposed to 24 h of hyperoxia; WT, WT IκBα-transfected cells exposed to room air; WT-O₂, WT IκBα-transfected cells exposed to 24 h of hyperoxia; Y42F, Y42F-transfected cells exposed to room air; Y42F-O₂, Y42F-transfected cells exposed to 24 h of hyperoxia; H₂O₂, fetal cells exposed to 1 mM H₂O₂ for 8 h; Y42F-H₂O₂, Y42F-transfected cells exposed to 1 mM H₂O₂. Values are means ± SE of 6 independent experiments for each group. Significantly different from time 0: *P < 0.05; †P < 0.0001. Right: representative Western blot showing levels of procaspase-3 in control fetal cells and cells overexpressing Y42F after exposure to hyperoxia (24 h) or H₂O₂ (8 h), with calnexin as loading control. B: representative agarose-ethidium bromide gel showing increased DNA laddering in fetal cells transfected with Y42F and exposed to hyperoxia. Positive control, sample DNA provided by manufacturer; air, control cells; hyperoxia, cells exposed to 24 h of hyperoxia; vector, empty vector-transfected cells; Y42F, Y42F-transfected cells.