Fig. 5

HDCA does not significantly inhibit the DCA-induced reduction of ΔΨ_m and increased production of ROS in isolated rat liver mitochondria. Isolated mitochondria were incubated with 100 µM DCA, 500 µM HDCA, 100 µM DCA + 500 µM HDCA, or no bile acid addition (control) for 5 min. In the coincubation experiments, mitochondria were pretreated with 500 µM HDCA alone for 5 min prior to addition of DCA. Isolated mitochondria (1 mg protein/ml) were suspended in respiration buffer and incubated for 15 min at 37°C with 50 nM DiOC₆(3), 2 µM HE, or 5 µM H₂DCFDA and analyzed by cytofluorometry. The percentages reflect (A) the disruption in ΔΨ_m; (B) the increased production of superoxides; and (C) the increased production of peroxides during treatment with DCA, and the absence of significant protection by HDCA. The data shown are representative of at least three different experiments and the treatment groups are indicated at left.