Fig. 8.

MiR-210 mimic and ISCU siRNA increase mitochondrial ROS and activated caspase-dependent death pathway. Rat primary cortical neurons at DIV5 were transfected with 50 μM miR-210 mimic, scramble (Neg. Ctrl), ISCU siRNA (si. Iscu), or scramble siRNA (si. Ctrl). Mitochondrial ROS (a, b; immunocytochemistry), intracellular ROS (c, d; DCF), caspase 3 activity (e, f), and caspase 9 activity (g, h) were detected 72 h after transfection. In a, b, representative confocal images of the colocalization of mitochondrial ROS indicator (MitoSOX, red) and cortical neurons (MAP 2; green). Scale bar, 20 μm. Cortical neurons were cultured on PDL-coated 12 mm glass coverslips in 24-well plates. After transfection, cells were washed with pre-warmed growth medium and then incubated with MitoSOX Red at a final concentration of 5 μM for 10 min at 37 °C. Then neurons were briefly washed with PBS followed by immunostaining for neurons (MAP 2) and nuclei (DAPI). In c–h, data are expressed as mean ± SEM. n =3 independent experiments. **, p < 0.005 vs Neg. Ctrl or si. Ctrl. Two-tailed Student’s t test