The human UC cell lines RT-112, VM-CUB-1, UM-UC-3, T24, 639 V, 253 J, 5637, SW170, HT-1376, BFTC-905, and J82, kindly provided by M. A. Knowles (Leeds, UK), J. Fogh (New York, NY), B. Grossmann (Houston, TX), or the DSMZ (Braunschweig, Germany), were grown in DMEM GlutaMAX-I (Gibco, Darmstadt, Germany) containing 10 % fetal calf serum. All cell lines were recently verified by standard DNA fingerprint analysis. For short-term experiments a single dose of cis-diamminedichloroplatinum-II (cisplatin; Accord Healthcare, London, UK) was added for 72 h; long-term treated (LTT) UCCs were generated by adding cisplatin after every passage at escalating doses over 8–10 months. Niclosamide was purchased from Sigma-Aldrich (#N3510, St. Louis, MO) and dissolved in DMSO. Reporter plasmids TopFlash or FopFlash (Merck Millipore, Billerica, MA) and mutated β-catenin-S33Y, kindly provided by H. Clevers (Utrecht, The Netherlands), were transfected using X-tremeGENE9 DNA Transfection Reagent (Roche, Basel, Switzerland) according to the manufacturer’s instructions.
Free full text: Click here
