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. 2012 Dec 3;32(42):5101–5110. doi: 10.1038/onc.2012.523

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Copyright © 2013 Macmillan Publishers Limited

This work is licensed under the Creative Commons Attribution-NonCommercial-No Derivative Works 3.0 Unported License. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-nd/3.0/

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Combined AMPK and FASN inhibition in PC cells is associated with an increased NOX activity and associated ROS production. (a) NADPH on NADP cellular ratio was measured in LNCaP cells by colorimetric assay at 48 h following indicated treatments. (b) NOX activity was measured by chemiluminescence in the presence of NADPH substrate and lucigenin at 24 h following indicated treatments. (c) NOX1 and -2 isoforms mRNA levels were analyzed by RT-qPCR at 24 h following indicated treatments. (d, e) FACS analysis of cellular ROS production measuring superoxide radicals (O₂^•−) and H₂O₂ with DHE (d) and DCFDA (e) fluoroprobes, respectively. (f) Co-treatment of cells with 2 μM NOX inhibitor DPI abrogated cC+C75-induced ROS increase, as measured with DCFDA fluoroprobe by FACS analysis. (g) Rescue from NOX1 mRNA level induction following cC+C75 treatment was measured by RT-qPCR following LNCaP cell transfection with siRNA against ME1, as compared with control.