Figure 4
Qp site substitutions lead to reduced cell death induction and ROS generation in response to MitoVES, and the induced cell death is dependent on CII-derived ROS. (a) Variant cell lines were exposed to 20 μM MitoVES for 24 h and the percentage of annexin V-positive cells was determined by flow cytometry. n≥5, mean±S.E.M., * values significantly different from WT. (b) Variant cell lines were exposed to 2 μM MitoVES for 30 min and the level of ROS was determined by dihydroethidium (DHE) staining and flow cytometry. n≥5, mean±S.E.M., * values significantly different from WT. (c) Cells were transfected with catalase-coding or control vector, exposed to 20 μM MitoVES for 20 h and the percentage of annexin V-positive cells was determined by flow cytometry. n≥4, mean±S.E.M., * values significantly different between catalase and mock-transfected cells. Inset, catalase overexpression verified by western blot. (d) Cells were exposed to 30 μM MitoVES for 12 h in the presence or absence of 20 mM malonate (30 min pretreatment). The percentage of annexin V-positive cells was determined by flow cytometry. n≥3, mean±S.E.M., * values significantly different in the presence and absence of malonate. (e) Cells were exposed to 5 μM MitoVES for 30 min in the presence or absence of 50 mM malonate (30 min pretreatment) and the level of ROS was determined by DHE staining and flow cytometry. n=5, mean±S.E.M., * values significantly different in the presence and absence of malonate
