Co-culture of Glioma Stem Cells and MSCs

Glioma stem cell line, GSC23 was kindly provided by the MD Anderson Cancer Center, University of Texas^{35 (link)}. HMSCs were bought from ScienCell Research Laboratories (Sciencell, Cat. #7500, CA, USA). Similar to previously reported methods^{36 (link)}, GSC23 cells were transfected with red fluorescence protein (RFP) gene, and HMSCs were transfected with green fluorescence protein (GFP) gene. In another method, GSC23 cells were also transfected with a lentiviral vector GV348 (sequence element: Ubi-MCS-SV40-puromycin) containing LOXP-STOP-LOXP-RFP gene, and MSCs were transfected with a lentiviral vector GV348 containing a CRE enzyme gene. The RFP, GFP and CRE-LOXP lentiviral vectors were packaged by Shanghai Genechem Co., Ltd (Shanghai, China). MSCs were cultured in Mesenchymal Stem Cell Medium (MSCM, Sciencell, Cat. #7501). GSC23 cells were maintained in Dulbecco’s Modified Eagle Medium/F12 (DMEM/F12), containing 20 ng/ml basic fibroblast growth factor (bFGF), 20 ng/ml epidermal growth factor (EGF), B27 supplement (50X), 2 mmol/l L-glutamine (100X), MEM vitamin solution (100X) and 100 mM sodium pyruvate (100X) (all from Gibco, Carlsbad, CA, USA).