Fig. 2

Mitophagy facilitates removal of damaged mitochondria in CuONPs-treated cells. A Representative immunofluorescence images of colocalization of GFP-LC3 and ATP5B in EA.hy926 cells treated with CuONPs (10 μg/ml). White arrows indicate colocalized dots of mitochondrial marker ATP5B with autophagosome marker GFP-LC3. Scale bars, 20 μm. B Western blotting analysis and quantification of protein level of LC3B, VDAC1, TIM23 and ATP5B in EA.hy926 treated with CuONPs (0, 5, 10, 15 and 20 μg/ml, respectively) for 12 h. C Western blotting analysis and quantification of protein level of LC3B, VDAC1, TIM23 and ATP5B in EA.hy926 treated with 10 μg/ml CuONPs for 0, 3, 6, 9 or 12 h. In (B) and (C), two-tailed unpaired Student’s t test was performed for statistical analysis. GAPDH was used as internal control. D The protein expression levels of VDAC1, TIM23 and ATP5B in EA.hy926 treated with or without wortmannin (Wort). Actin was used as internal control. E Representative FCM results of EA.hy926 cells labeled with MitoTracker Green FM. The cells were pretreated with or without chloroquine (CQ) for 1 h and then treated 10 μg/ml CuONPs for 12 h. MFI, mean fluorescence intensity. In D and E, one-way ANOVA with Tukey’s test was used for statistical analysis. All data are representative of at least three independent experiments. Data are mean ± S.D. *p < 0.05