Pcs 440 030

All cell lines were obtained from ATCC. PC3 and DU145 cells were cultured in F-12K medium and Eagle’s minimum essential medium supplemented with fetal bovine serum (FBS) to a final concentration of 10%, respectively. C4-2B cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM)/F12 (4:1) supplemented with heat-inactivated FBS (to 10%) and T-medium supplement (to 1×). To prepare 10×T-medium supplement in 100 ml of 0.1% BSA (Sigma cat# A8022) in PBS, the following components were added: 50 mg insulin (Gibco cat# 12585-014), 136 ng triiodo-l-thyronine (Sigma cat# T2877), 50 mg transferrin (Sigma cat# T4382), 2.5 mg d-Biotin (Sigma cat# 47868), and 250 mg adenine (Sigma cat# A3159). RWPE-1 and PZ-HPV-7 cells were maintained in keratinocyte serum free medium (K-SFM, Gibco 17005-042), supplemented with 0.05 mg/ml of bovine pituitary extract and 5 ng/ml of human recombinant epidermal growth factor. HPrEC cells were maintained in prostate epithelial cell basal medium (ATCC, PCS-440-030) supplemented with prostate epithelial cell growth kit (ATCC, PCS-440-040). HeLa cells were cultured in DMEM supplemented with 5% FBS.

The immortalized human normal prostate epithelial cell line RWPE1 was purchased from the American Type Culture Collection (ATCC, CRL-11609) and maintained in culture in prostate epithelial cell basal medium (ATCC PCS-440-030) supplemented with the prostate epithelial cell growth kit (ATCC PCS-440-040) in T25 flasks. The RWPE1 cell lines were authenticated and characterized by ATCC, which uses morphology, karyotyping, and PCR-based approaches to confirm the identity of cell lines. All cells were maintained at lower passages.

Immortalized 957E/hTERT and 957E/hTERT-AR human prostate epithelial cells were generously provided by John T. Isaacs lab (Johns Hopkins, Baltimore, MD, USA) [9 (link),10 (link)]. The prostate epithelial cell line RWPE-1 was purchased from the American Type Culture Collection (ATCC, Cat. No. CRL-11609). The cells, 957E/hTERT, 957E/hTERT-AR and RWPE-1, were grown in Keratinocyte-SFM (serum-free medium) supplemented with bovine pituitary extract (BPE) and human recombinant epidermal growth factor (EGF) supplied as a kit (Cat. No. 17005-042, Thermo Fisher Scientific, Waltham, MA, USA), and 1% penicillin-streptomycin. Primary prostate epithelial cells (PrEC) were purchased from ATCC, Manassas, VA, USA (Cat. No. PCS-440-010) and were grown in prostate epithelial cell basal medium (ATCC, PCS-440-030), supplemented with prostate epithelial cell growth kit contents (ATCC, PCS-440-040). The prostate epithelial cell line EP156T was grown in MCDB153 medium (Biological Ind. Ltd., Beit Haemek, Israel), supplemented with 1% fetal calf serum (FCS), growth factors and antibiotics as described [3 (link),5 (link)]. Complete growth media were changed every 2–3 days.