Fig. 3. Flubendazole induces DRP1-mediated mitophagy in MDA-MB-231 and MCF-7 cells.
A Immunoblotting of DRP1, p-DRP1ser616 and Mitofusin-2 in MDA-MB-231 and MCF-7 cells treated with the indicated concentrations of flubendazole for 24 h. β-actin was used as the loading control. B DRP1 mRNA expression in MDA-MB-231 and MCF-7 cells was analyzed by RT-qPCR. C, D MDA-MB-231 and MCF-7 cells were transfected with negative-control or DRP1 shRNA for 24 h, respectively. After treatment with or without flubendazole (0.5 μM) for 24 h. The autophagosomes are labeled by LC3 (green fluorescence) protein, and the mitochondria are labeled by TOM20 (red fluorescence) protein. The number of co-localized LC3 and TOM20 was quantified. Scale bar, 5 µm. E, F Colocalization of PINK1 (green fluorescence) protein and Parkin (red fluorescence) protein in MDA-MB-231 and MCF-7 cells following flubendazole (0.5 μM, 24 h) treatment. The number of co-localized PINK1 and Parkin was quantified. Scale bar, 10 µm. G Immunoblotting of Parkin, p-Parkinser65 and PINK1 expression. β-actin was measured as the loading control. Data represent mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001. Statistical significance compared with respective control groups (all P-values were obtained by one-way ANOVA).