Fgf 2 growth factor

Human placental mesenchymal stem cells (hpMSCs) were purchased to Cellular engineering Technologies (CET (Coraville, IA, USA)). Metastatic murine skin melanoma cells (B16-F10 cells) and glioblastoma cells (U251-MG) were provided by cell services from Cancer Research-UK. They were grown in Dulbecco’s modified Eagle’s medium (DMEM, Biowest, France) supplemented with 10% fetal bovine serum (FBS, GIBCO, USA), 1% penicillin/streptomycin and 1% amphotericin (Biowest, France). For culturing hpMSCs, 5 µg mL⁻¹ of FGF-2 growth factor (PeproTech, USA) were also added to the cell culture media. hpMSCs were maintained under hypoxic conditions whereas U251-MG cells and B16-F10 cells were cultured under and normoxic conditions.
To obtain the culture media free of exosomes, they were depleted from FBS by ultracentrifugation (100,000g, 8 h, 4 °C).

Human placental mesenchymal stem cells (hpMSCs) were obtained from Cellular Engineering Technologies (CET) (Caralville, IA, USA), while B16-F1 (low metastatic variant) and B16-F10 (high metastatic variant) murine skin melanoma cells, were provided by cell services from Cancer Research-UK. NIH-3T3 murine healthy fibroblasts were obtained from Dr. Antonio de la Vieja’s group (Instituto de Salud Carlos lll). hpMSCs were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Biowest, France) supplemented with 5 μg mL−1 of FGF-2 growth factor (PeproTech, Cranbury, NJ, USA), with 10% of fetal bovine serum (FBS, GIBCO, Waltham, MA, USA), 1% penicillin/streptomycin and 1% amphotericin (Biowest, France) and maintained at 37 °C in a 5% CO₂-humidified atmosphere under hypoxic conditions (3% O₂). For culturing B16-F1, B16-F10, and NIH-3T3 cells, DMEM with 10% of FBS (GIBCO), supplemented with 1% penicillin/streptomycin and 1% amphotericin (Biowest, France) was used. Finally, monocytes were cultured in RMPI Medium 1640 (Biowest, France) supplemented with 10% FBS (GIBCO), 1% penicillin/streptomycin and 1% amphotericin (Biowest, France). B16-F1, B16-F10 and NIH-3T3 cells were maintained under normoxic conditions.
To obtain culture media free of EVs, they were depleted from serum by ultracentrifugation at 100,000× g for 8 h at 4 °C.