Ht 1197

RT4, BFTC905, BFTC909, TCCSUP cell lines were purchased from the Food Industry Research and Development Institute of Taiwan. Cell culture condition was performed as previously described [46 (link)]. HT1197, J82, were purchased from ATCC (Manassas, VA 20108, USA). We cultured J82 in Dulbeco's modified Eagle's medium containing 10% FBS, 1% Penicillin-Streptomycin, 100X (GIBCO). HT1197 cells were cultured in Eagle's Minimum Essential Medium containing 10% FBS, 1% MEM Non-Essential Amino Acids Solution, and 1% Antibiotic-Antimycotic, 100X(GIBCO). A non-tumoral urothelial primary cell, HUC (ScienCell Research Laboratories, San Diego, CA), was used as a control and maintained using the suggested medium and conditions. To generate stable CEBPD expression cell lines. TCCSUP cells were plated in 6-well plate at a density of 1×10⁶ per well, and then were infected by lentivirus for 48 hours. We purchased the pLKO-AS3w-puro expression vector from the National RNAi Core Facility (Academia Sinica, Taipei, Taiwan). Lentiviral vector express CEBPD (pLKO-AS3w-CEBPD-eGFP) was constructed according to protocol. The DNA sequence containing CEBPD's open-reading frame (ORF) was cloned from Hela cells using specific primers and inserted into the pLKO-AS3w-puro vector. The pLKO-AS3w-eGFP-puro plasmid was used as a negative control. Virus was produced as previously described [47 (link)].

The bladder cancer cell lines RT4, UM-UC-3, SCaBER, HT-1197, HT-1376, and 5637 were obtained from ATCC, and UM-UC-1 was obtained from Sigma-Aldrich. RT4 cells were maintained in McCoy’s 5A culture medium and all other cell lines were maintained in MEM with 2 mM glutamine (Corning, Corning, NY, USA). Culture medium for HT-1197 and HT-1376 was supplemented with 1× non-essential amino acids and 1 mM sodium pyruvate (Gibco, Waltham, MA, USA). All culture media were supplemented with 10% fetal bovine serum (Corning, Corning, NY, USA) and 1 μg/mL Gentamicin (Gibco, Waltham, MA, USA). Cells were grown in 60 mm tissue culture dishes and were incubated at 37 °C, 5% CO₂ with 95% humidity. The cells were examined under a microscope on a daily basis, and the medium was renewed every 2 days. Cells were passaged at 70% confluence using 0.25% trypsin, 0.53 mM EDTA solution (Corning, Corning, NY, USA).