Colo320

Four cell lines derived from human colorectal carcinoma were examined: HT-29, WiDr, CaCo-2 and Colo320. All cell lines were obtained from the American Type Culture Collection (Rockville, MD, USA). The HT-29 was cultured in McCoy’s supplemented with 10% fetal bovine serum (FBS). WiDr and CaCo-2 were maintained in minimum essential medium eagle (Sigma Chemical Co., St. Louis, MO, USA) with high glucose and 10% FBS. Colo320 was maintained in RPMI-1640 medium (Sigma Chemical Co.) supplemented with 10% FBS. Fibroblasts were obtained from Lonza Walkersville Inc. (Walkersville, MD) and maintained in FBM-2 medium supplemented with 2% FBS, 1 ng/ml bFGF, and 1 mg/ml insulin. Human umbilical vein endothelial cell (HUVEC) was obtained from Kurabo Co. (Osaka, Japan). HUVEC were culture in HuMedia-EB2 medium supplemented with 2% FBS, 5 ng/ml of basic fibroblast growth factor, 10 µg/ml heparin, 10 ng/ml epidermal growth factor, and 1 µg/ml hydrocortisone according to the supplier’s instructions (Kurabo Co.). Fibroblast was obtained from Lonza (Walkersville, MD) and maintained in FBM-2 medium supplemented with 2% FBS, 1 ng/mL basic fibroblast growth factor, and 1 µg/ml insulin according to the supplier’s instructions. All cells were incubated at 37 °C in a humidified atmosphere of 5% CO₂ in air.

COLO320 were purchased from ATCC (Manassas, VA, USA). A549, H1299 and COLO320 cancer cells were grown in RPMI-1640 medium supplemented with 2 mM L-glutamine, 100 IU/ml penicillin/streptomycin, and 10% fetal calf serum (FCS; Sigma-Aldrich, St Louis, MO, USA). The rescue experiments were performed in A549 and H1299 cancer cells grown in Miminum Essential Medium Eagle (Sigma-Aldrich) supplemented with 1X MEM Vitamin Solution, 2 mM L-glutamine, 25 mM glucose, 10% fetal bovine serum dialyzed, 100 IU/ml penicillin/streptomycin and 0.4 mM serine, 0.4 mM glycine and 0.02 mM 5-formyl-THF (Merck Eprova AG).