Differential protection of lipid and protein oxidation by bilirubin and glutathione. (A) Bilirubin blocks oxidant-induced accumulation of 4-hydroxynonenal (4-HNE) adducts. HEK293 cells were treated with combinations of 500 μM tert-butyl hydroperoxide (TBH) and 100 nM bilirubin (BR) for 16 h and stained for 4-HNE. Data represent means ± SEM from 4 samples. *, P < 0.001 from each other treatment (ANOVA followed by Tukey-Kramer post hoc test). Representative of 3 experiments. (B) Greater potency of water-soluble antioxidants for protein oxidation than bilirubin. HEK293 cells were radiolabeled with [35S]methionine, and protein was oxidized with combinations of H2O2 (10 μM), bilirubin (5 μM), and the water-soluble antioxidants, glutathione (5 μM) and ascorbate (5 μM). Data represent means ± SEM of triplicate determinations, representative of 3 experiments. *, P < 0.001 (ANOVA followed by Tukey-Kramer post hoc test).