Figure 7

Mitochondrial-derived ROS production was involved in rhArg-induced autophagy and cytotoxicity in H1975 cells. (a) H1975 cells were treated with 2 U/ml of rhArg in the presence or absence of 5 mM NAC for 72 h and stained by MitoSox and Cyto-ID green dye to detect the ROS and autophagy. Confocal micrographs were taken at × 40. Green (Cyto-ID) and red (MitoSox) dots in cells were counted by the ImageJ software (n=3, means±S.D.). (b) H1975 cells were incubated with 2 U/ml of rhArg for 24 h in the presence or absence of 5 mM NAC and the expression of LC3-I/II, p-Akt, p-mTOR and p-Erk 1/2 was measured by western blot analysis. Densitometric values were quantified using the ImageJ software and normalized to control. The values of control were set to 1. The data are presented as means±S.D. of three independent experiments. (c) H1975 cells were treated with 2 U/ml of rhArg in the presence or absence of 5 mM NAC for 72 h, and cell viability was analyzed by MTT assay. Results were expressed as mean±S.D. and analyzed by Student’s t-test (two-tailed). **P<0.01 (d) H1975 cells were incubated with 2 U/ml of rhArg for 24 h in the presence or absence of 5 mM NAC and the expression of PARP, cleaved-PARP and cleaved-caspase 3 was measured by western blot analysis