Bj fibroblasts

The human pancreatic cancer cell lines AsPC1 (CRL-1682), BxPC3 (CRL-1687), CFPAC1 (CRL-1918), MIAPaCa2 (CRL-1420), PANC1 (CRL-1469), and BJ fibroblasts (CRL-2522) were acquired from ATCC, KP4 (JCRB0182) from JCRB, and cultured per supplier’s instructions. HPDE6C7 was provided by Tsao group (U. Toronto) and cultured as previously described^{31 (link)}. The spontaneously immortalized human pancreatic stellate cell line hPSC, ONO, and YAM1 were kindly provided by the Evans group (Salk) as previously described^{11 (link)}. Normal human fibroblast cells were a kind gift from the Gage group (Salk). MEF cells for Lifr gene knockout characterization were isolated from individual embryos at E13.5, and cultured in DMEM containing 10% FBS for less than five passages. For the low dosage Gem treatment assay, 3 nM gemcitabine or vehicle (PBS) were used.

IMR90, MEFs, HEK293T were described previously^{16 (link),31} . Primary BJ fibroblasts were purchased from ATCC. Cell line identities were not further authenticated. The cells were cultured in DMEM supplemented with 10% fetal bovine serum (FBS), 100 units/ml penicillin, and 100 µg/ml streptomycin (Invitrogen), and were intermittently tested for mycoplasma. IMR90 and BJ were cultured under physiological oxygen (3%), except for the H₂O₂ treatment, in which cells were cultured in 20% oxygen incubator, and the experiments involved in live-cell imaging. For primary cell cultures, cells were briefly washed with PBS, trypsinized at 37°C for 2–4 min, and passaged at no more than 1:4 dilutions. Cells were counted with Countess automated cell counter (Life Technologies), and the numbers were recorded where growth curves are generated. HEK293T were transfected using Lipofectamine 2000 (Invitrogen). For amino acid starvation, cells were incubated in Hank’s buffer (with calcium and glucose) supplemented with 10% dialyzed FBS and 1% HEPES (Invitrogen). For amino acid and serum deprivation, cells were cultured in Hank’s buffer plus 1% HEPES.