Nigg, Basel) cells were cultured in DMEM containing 4.5 g/l glucose supplemented with 10% fetal bovine serum (Sigma), 2 mM L-glutamine (Sigma), 100 U/ml penicillin and 0.1 mg/ml streptomycin (Sigma) at 37°C in 5% CO2.
COS7 cells were cultured in DMEM supplemented with 10% FBS and transfected using the FuGene Transfection Reagent (Roche, Basel, Switzerland) Cells were lysed in ice-cold M-PER buffer (PIERCE, Rockford, IL) containing protease inhibitors for 15 min on ice.
MRMT-1 rat mammary gland carcinoma cells (Novartis Oncology Research, Basel) and H1299 human lung cancer cells were cultured in RPMI 1640 (Gibco) medium, human SY5Y cell lines were cultured in Dulbecco's modified Eagle's medium (DMEM).
In some experiments, BM specimens of 6 CML patients at diagnosis were cultured in vitro for 3 to 14 days with 5 µM of the purified imatinib mesylate (Novartis Pharma AG, Basel, Switzerland, MTA31812); cells were recovered at day 3, 5, 7 and 14 for RNA extraction.
Transfections were performed using Mammalian Epithelial Cells Amaxa Basic Nucleofector Kit (Lonza, Basel, Switzerland) and cells were electroporated according to manufacturer's instructions.
Human umbilical artery smooth muscle cells (Lonza, Basel, Switzerland) were cultured in Smooth Muscle Cell Growth Medium 2 (PromoCell, Heidelberg, Germany) supplemented with 50 U/ml penicillin and 50 μg/ml streptomycin.
Cell viability was quantified using the WST-1 proliferation reagent (Roche, Basel, Switzerland) or alamarBlue cell viability assay (Invitrogen) and a plate reader (Tecan, Männedorf, Switzerland).
Additionally, PREC cells (Lonza, Basel, Switzerland) were used as a normal human prostate cell line control.
Human skeletal muscle cells (Lonza, Basel, Switzerland) were cultured at a density of 100,000 cells in a six-well plate in 2 ml DMEM (Gibco, Carlsbad, CA, USA).
Cell survival and death were quantified by measuring absorbance of the dye product from the nonradioactive quantitative reagent WST-1 (Roche, Basel, Switzerland) and by cell counts using a hemocytometer.
