Epinephrine

MCF-7 cells were grown in low glucose Dulbecco modified eagle medium (DMEM), U87 cells were grown in high glucose DMEM, MDA-MB-231 and BT-549 cells in Roswell Park Memorial Institute (RPMI) 1640. Cell line media were supplemented with 10% fetal bovine serum (FBS), penicillin and streptomycin from Biological Industries Kibbutz Beit-Haemek, Il. Growth medium of BT-549 was also supplemented with 0.023 IU/ml of human recombinant insulin (Sigma, St. Louis, MO.). Fibroblasts were grown in high glucose DMEM supplemented with 20% FBS, penicillin-streptomycin, 1% non-essential amino acids and 0.2% β-mercaptoethanol (Invitrogen, Life Technologies Grand Island, NY). Primary normal human breast cells were grown in mammary epithelial cell basal medium containing recombinant human insulin, and TGFα, L-glutamine, epinephrine, apo-transferrin, pituitary extract and hydrocortisone hemisuccinate all from ATCC. For western blot analysis of cellular proteins cells were plated at a density of 4·10³ per cm² and drugs were added 48 hours post-plating from stock solutions in dimethyl sulfoxide (DMSO) (Sigma, St. Louis MO). Controls received the vehicle. The concentration of DMSO in the growth medium did not exceed 0.08%.

Human prostate cancer cell lines (LNCaP, VCaP, PC3 and 22Rv1), murine prostate cancer line (Myc-CaP), normal cultured prostate epithelial cells (HPrEC), benign prostatic hypertrophy cells (BPH-1) and murine Lewis lung carcinoma cells (LLC-1) were all obtained from ATCC, after authentication by short tandem repeat (STR) profiling. All cell lines used in the paper were derived from male mice or male human patients. They were cultured in the following media at 37°: RPMI-1640 (ATCC) medium supplemented with 10% FBS (Gibco) and 1X Pen/Strep (Gibco) (for LNCaP, VCaP, PC3, 22Rv1 and BPH-1 cells); Prostate Epithelial Cell medium (ATCC) with 6 nM L-glutamine (ATCC), 0.4% Extract P (ATCC), 1.0 mM Epinephrine (ATCC), 0.5 ng/ml rh-TGFα (ATCC), 100ng/ml hydrocortisone hemisuccinate (ATCC), 5 mg/ml rh-Insulin (ATCC), 5 mg/ml Apo-transferrin (ATCC), 33 μM Phenol red (ATCC) and 1X Pen/Strep/Ampho Solution (ATCC) (for HPrEC cells); DMEM high glucose medium (Gibco) with 10% FBS (Gibco) and 1X Pen/Strep (Gibco) (for Myc-CaP cells and LLC-1 cells). All the cell lines used in this study were checked for mycoplasma every 4 months using Mycoalert kit (Lonza).