Separated proteins were blotted onto nitrocellulose membranes (Whatman, Dassel, Germany) using a wet electro-blotting apparatus (Bio-Rad, USA).
Cell lysates were extracted, and LRP-5 (sc-21389; Santa Cruz Biotechnology, Santa Cruz, CA, USA), phospho-β-catenin (9561; Cell Signaling Technology, Boston, MA, USA), and total β-catenin protein levels (9582; Cell Signaling Technology, Boston, MA, USA) were evaluated by using Western blot analysis, as described earlier.
Antibodies specific for PTEN (138G6), phospho-PTEN (Ser), AKT (Cell Signaling Technology), phospho-AKT (Ser), PARP, cleaved PARP (Cell Signaling Technology, Beverly, MA), RAD51 (Millipore, MA, USA and Santa Cruz Biotechnology, CA, USA), and β-actin (Sigma-Aldrich, MO, USA) were used for western blotting, as recommended by the manufacturer.
For all other antibodies, Alexa Fluor 680 goat antibodies to mouse IgG (Invitrogen) or IR Dye 800CW donkey antibodies to rabbit IgG (LI-COR, Lincoln, NE, USA) were used as secondary antibodies, and blots were examined using the ODYSSEY infrared imager (LI-COR).
Enhanced chemiluminescence (Bio-Rad, Hercules, CA, USA) were used to visualize blots.
Primary antibodies against AR (1 2000) and FOXA1 (1 500; Santa Cruz Biotechnology, Dallas, TX, USA) were used for western blotting.
Mouse anti-human β-actin antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) and mouse anti-human FLG antibody (Vector Laboratories, Inc., Burlingame, CA, USA) were used for western blotting.
Recombinant topo I and polyclonal human anti-topo I IgG (scl-70) from Topogen (Columbus, USA) were used for western blot.
Antibodies against PPAR α (Abcam, Cambridge, MA, USA), phosphorylated ERK, phosphorylated JNK, phosphorylated p38, phosphorylated NF- κBp65, LC3B, Atg7, Atg5, Beclin-1, Lamp1 and β-actin (Cell Signaling Technology Inc., Santa Cruz, CA, USA) were used for western blot analysis.
Anti-β2-M antibody (self-government), anti-β-actin antibody (Rockland, Gilbertsville, PA, USA), anti-Bcl-2 antibody, anti-HER-2 antibody, anti-ER antibody, anti-PR antibody and horseradish peroxidase-conjugated secondary antibody (Invitrogen, Carlsbad, CA, USA) were used for western blotting.
