Keratinocyte sfm

This study used PZ-HPV-7 (an immortalized prostate epithelial cell line), CA-HPV-10 (an immortalized prostate adenocarcinoma cell line) and LNCaP, PC-3 and Du145 (metastatic prostate cancer cell lines). In addition, the HECV vascular endothelial cell line was used. All cell lines were purchased from the American Tissue Culture Collection (ATCC) (Manassas, VA, USA) at the commencement of this study. PC-3, Du145 and HECV cell lines were maintained in DMEM medium (Sigma-Aldrich, Gillingham, Dorset, UK). LNCaP clone FGC cell line was maintained in RPMI-1640 medium (Sigma-Aldrich, Gillingham, Dorset, UK). PZ-HPV-7 and CA-HPV-10 cell lines were maintained in Keratinocyte-SFM (Sigma-Aldrich, Gillingham, Dorset, UK).

Primary HFKs were grown in 6-cm dishes in Keratinocyte-SFM (Gibco, Paisley, UK) supplemented with EGF and bovine pituitary extract. They were transfected at 70% confluency with 50 nM 21-mer siRNA targeting NRF2 (Sigma-Aldrich, Buchs, Switzerland) or AHR (Life Technologies, Carlsbad, CA, USA) using INTERFERin (Polyplus, Illkirch, France). After 48 h at 100% confluency, HFKs were washed with PBS and cultured for 48 h in supplement-free Keratinocyte-SFM medium with 10⁻⁸M TCDD (Sigma-Aldrich) in toluene/DMSO (0.1% final concentration) or toluene/DMSO only.