Figure 3.

Quantification of ROS and Mitochondrial OCR in miR-17-3p-Expressing PC-3 Cells

(A) After treatment, the levels of cellular ROS were determined using a total cellular ROS detection kit. ROS inhibitor NAC was included to eliminate the IR-induced ROS. (B) The levels of ROS were calculated. The cells were treated with or without NAC followed by IR treatment as indicated. (C) The cell viability was quantified by MTT, and (D) the cell surviving fraction was determined by MTT and colony formation. (E) Mitochondrial OCR in the treated cells was measured using a Seahorse XF-96 analyzer. Supplemental reagents, including oligomycin, FCCP, and antimycin or rotenone, were automatically injected into the analyzing plates to determine the OCR of basal, ATP-linked, maximum, and background image, respectively. (F) The relative OCR was calculated, and the OCR reverse capacity, an important mitochondrial respiration index, was determined by the maximal OCR subtracted from the basal OCR. Values are presented as the mean ± SD (n ≥ 3) and statistical significances between two groups are as described in Figure 2.