Pregm bullet kit

Two human PCa cell lines, LNCaP and PC3, were purchased from the American Type Culture Collection (Manassas, VA, USA) and cultured in RPMI 1640 medium (HyClone, Los Angeles, CA, USA) supplemented with 10% fetal bovine serum (GIBCO/Invitrogen, Carlsbad, CA, USA), 2 mM l-glutamine, and antibiotics. A normal human prostate epithelial cell line (PREC) was purchased from Lonza Company and cultured in PrEGM Bullet kit (Lonza, Allendale, NJ, USA) with antibiotics. All cell lines were maintained at 37 °C in a humidified chamber supplemented with 5% CO₂.

The human prostate epithelial cell line, PrEC, was purchased from Lonza (Walkersville, MD, United States). PrEC cells (passages 5–11) were cultured in PrEGM BulletKit medium (Lonza) supplemented with 10% fetal bovine serum (FBS; GIBCO/Invitrogen, Grand Island, NY, United States) and 100 U/ml penicillin plus 100 μg/ml streptomycin (GIBCO/Invitrogen) at 37°C. The human prostate cancer cell lines, LNCaP and DU145 (originally from the American Type Culture Collection, Manassas, VA, United States), were provided by Dr Hirotsugu Uemura (Kindai University, Osaka, Japan) (Muramatsu et al., 2019 (link)). The human prostate cancer cell line, PC-3, was obtained from the Cell Resource Center for Biomedical Research, the Institute of Development, Aging and Cancer, Tohoku University (Sendai, Japan). LNCaP, PC-3, and DU145 cells (passages 5–11) were cultured in RPMI-1640 medium (Sigma-Aldrich, St. Louis, MO, United States) supplemented with 10% FBS and 100 U/ml penicillin plus 100 μg/ml streptomycin at 37°C. Experiments using VEGF-C stimulation were performed after the exposure to RPMI-1640 medium containing 1% FBS for 4 h to prevent enhanced phosphorylation and starvation stress.