β-Naphthoflavone-Induced Mitochondrial Respiratory Damage in Cyp1 Knockout Mouse and in Cell Culture Systems: Attenuation by Resveratrol Treatment

Figure 6

Effects of BNF on ROS production and mitochondrial respiratory complexes. ROS formation in C6 glioma cells treated with BNF, resveratrol, proadefin, or CG223191 was measured as described in Materials and Methods (a). Complex I assay was carried out using Cary 1E UV-visible spectrophotometer by incubating 10 μg of freeze-thawed mitochondrial extract in 1 ml of assay medium. The assay medium consisted of 25 mM potassium phosphate, pH 7.4, 5 mM MgCl₂, 2 mM NaCN, 2.5 mg/ml bovine serum albumin, 13 mM NADH, 65 μM ubiquinone, and 2 μg/ml antimycin A. The decrease in absorbance at 340 nm because of NADH oxidation was measured (b). CcO activity was measured by incubating 10 μg of freeze-thawed mitochondrial extract from control and BNF-treated cells in 1 ml of assay medium (25 mM potassium phosphate, pH 7.4, 0.45 mM dodecylmaltoside, and 15 μM reduced cytochrome c) (c). Results represent mean ± SEM of at least 4 independent experiments. denotes .

(a)

(b)

(c)