Tgfβr1 inhibitor

The human embryonic kidney cell line 293T (HEK293T), immortalized colon mucosa epithelial cell line (FHC), and human CRC cell lines HT29, LoVo, HCT116, SW480, DLD1, LS174t, SW620 and RKO were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA). A subclone named M5, which has enhanced metastatic abilities in the liver, was isolated by the in vivo selection of SW480 cells in our laboratory.¹⁶ All CRC cell lines were cultured in RPMI 1640 medium (Gibco, Gaithersburg, MD, USA) with 10% fetal bovine serum (FBS, Gibco) and 100 U/ml penicillin/streptomycin (Gibco). 293T cells were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% FBS. FHC cells were cultured in DMEM:F12 medium (Gibco) with 10% FBS, 25 mmol/L HEPES, 10 mg/L cholera toxin, 5 mg/L insulin, 5 mg/L transferrin, 100 mg/L hydrocortisone, and 20 mg/L human recombinant epidermal growth factor (EGF). To examine the effects of TGF‐β signaling, LoVo, HCT116, SW480 and HT29 cells were treated with 10 ng/mL recombinant human TGF‐β1 (PeproTech Inc, Rocky Hill, NJ, USA) for 48 h, or with 1 μΜ TGFβR1 inhibitor (SB525334, Selleckchem, Houston, TX, USA) for 48 h, respectively. All of the cell lines were cultured at 37°C and in 5% CO₂ in air in an humidified incubator.