FIG. 5.

Statin treatment lowers ROS production and mitochondrial membrane potential. (A) HL-1 cells were given 1 μM simvastatin with or without the concurrent supplementation with 5 μM coenzyme Q₁₀ for 24 h. Cells were then loaded with 10 μM H₂DCFDA for 30 min at 37°C and subsequently analyzed via FACS for fluorescence intensity per cell as a measure of ROS. (B) Representative FACS histograms of HL-1 cells loaded with membrane potential-sensitive MitoTracker Red CMX Rosamine for 30 min at the end of statin and/or coenzyme Q₁₀ treatments, which were then fixed with 4% paraformaldehyde before analysis. Unstained cells, and cells given 10 μM FCCP for 1 h served as controls. (C) Mean average fluorescence intensity (of the MitoTracker Red CMX Rosamine) was quantified as a measure of relative mitochondrial membrane potential changes. n=3 plates per group; 10,000 cells were examined per plate. *p-value<0.05. (D) Cells were treated as in (A), then immunolabeled with anti-Tom70 and nuclear stain Hoechst 33432. n=3 plates per group; 50 cells per plate were analyzed for MitoTracker Red CMX Rosamine normalized to Tom70 fluorescence. *p-value<0.05. FACS, fluorescence-activated cell sorting; FCCP, carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone; ROS, reactive oxygen species; Tom70, translocase outer membrane 70 kDa subunit.