Fig. 3

MnCl₂ induced mitophagy by regulating ROS generation.  a, b SH-SY5Y cells were treated with 200, 400, and 800 µM MnCl₂ (a) and 400 µM MnCl₂ with or without pretreatment with the antioxidant NAC (b) for 24 h. ROS levels were measured using a microplate reader. c BNIP3, LC3 and TOMM20 protein levels were measured by western blot after treatment with 400 µM MnCl₂ with or without pretreatment with the antioxidant NAC for 24 h. β-actin served as the internal reference. Compared with control, *P < 0.05, **P < 0.01. d Mitophagy bodies and mitochondria were observed by transmission electron microscopy (TEM) in cells treated with 400 μM MnCl2 alone or co-treated with the mitophagy inhibitor NAC for 24 h. N, nucleus. White arrow, expanded, condensed or dissolved, mitochondria aggregates. Black arrow ( ), relatively normal mitochondrial morphology. Black arrow ( ), mitophagy bodies. e After co-treatment with or without the autophagy inhibitor 3-MA, ROS levels in SH-SY5Y cells were also measured using a microplate reader