Fig. 4
Effects of AGEs on oxidative stress in HCAECs. a O2 − levels. Cells were treated with 50, 100 or 200 μg/ml of AGEs for 24 h, and intracellular O2 − levels were determined by DHE (5 μM) staining and flow cytometric analysis. b ROS production (glutathione assay). HCAECs were treated with AGEs (100 μg/ml) and/or other molecules for 24 h. RLU, relative light units; TFA, thenoyltrifluoroacetone. c Mitochondrial membrane potential (JC-1 staining and flow cytometry). Cells were treated with AGEs for 24 h, and mitochondrial membrane potential was determined by JC-1 staining and flow cytometric analysis. d ATP content. Cells were treated with 100 μg/ml AGEs for 24 h, and ATP content was determined by an ATPLite kit. * P < 0.05, compare with control. e Effect of mitochondrial complexII inhibitor TTFA on eNOS mRNA levels. HCAECs were treated with AGEs and/or TFA (10 μM) for 24 h, and eNOS mRNA levels were determined by real-time PCR analysis. F: Effect of mitochondrial complexII inhibitor TTFA on eNOS protein levels. HCAECs were treated with AGEs and/or TTFA (10 μM) for 24 h, and eNOS protein levels were determined by Western blot. Full-length blots are presented. * P < 0.05, compare with control, # P < 0.05, compare with AGEs treatment, n = 3. Data are means and SE of multiple experiments (n)