U118mg

U251, U118MG and LN229 cell lines were purchased from the American Type Culture Collection. Patient-derived GBM stem-like cells (GSCs) P3, BG5 and BG7 were previously isolated and characterized from GBM surgical specimens. All human cell lines were validated through short tandem repeat analysis (Cell Cook Biotech Co., Ltd; Guangzhou, China). Primary GBM cell lines P3, BG5 and BG7 were kindly provided by Prof. Rolf Bjerkvig (University of Bergen, Bergen, Norway). U118MG and LN229 cell lines were maintained in Dulbecco's modified Eagle's medium (DMEM; Life Technologies/Thermo Fisher Scientific; Waltham, MA, USA) supplemented with 10% fetal bovine serum (Thermo Fisher Scientific). GBM#P3 cells were cultured in serum-free DMEM/F12 medium (Gibco/Thermo Fisher Scientific) supplemented with 2% B27 neuromix (Thermo Fisher Scientific), epidermal growth factor (20 ng/mL; and basic fibroblast growth factor (10 ng/mL; Thermo Fisher Scientific). Normal human astrocytes (NHA) were obtained from Lonza (Walkersville, MD, USA) and cultured in Astrocyte Medium (ScienCell; Carlsbad, CA, USA) supplemented with the Astrocyte Growth Medium Bullet Kit (ScienCell). Cells were incubated in 5% CO2 at 37 °C.

Human GBM cell lines (U87MG, U118MG and LN229) and the human embryonic kidney cell line 293 (HEK293) were purchased from the American Type Culture Collection. All human cell lines were authenticated and submitted for short tandem repeat analysis (Cell Cook Biotech Co. LTD; Guangzhou, China). Primary GBM#P3 cells and BG5 glioma stem cells (GSC) were kind gifts provided by Prof. Rolf Bjerkvig (University of Bergen, Norway). Cells (U87MG, U118MG, LN229, and HEK293) were maintained in the Dulbecco’s modified Eagle’s medium (DMEM; Life Technologies/Thermo Fisher Scientific; Waltham, MA, USA) supplemented with 10% fetal bovine serum (Thermo Fisher Scientific). Cells (GBM#P3 and GSC BG5) were cultured in serum-free DMEM/F12 medium (Gibco/Thermo Fisher Scientific) supplemented with 2% B27 Neuro Mix (Thermo Fisher Scientific), epidermal growth factor (20 ng/mL; Thermo Fisher Scientific), and basic fibroblast growth factor (10 ng/mL; Thermo Fisher Scientific). Cells were maintained at 37 °C in a humidified chamber containing 5% CO₂.

The human GBM cell lines U87MG and U118MG were purchased from the American Type Culture Collection (ATCC). U87MG and U118MG was maintained in Minimum Essential Medium (MEM) and Dulbecco’s modified Eagle’s medium (DMEM), respectively, supplemented with 10% fetal bovine serum (Gibco, United States), 100 μg/ml streptomycin, and 100 U/ml penicillin, at 37°C in 5% CO₂. Three small interfering RNAs (siRNAs) against METTL3 were used to inhibit METTL3 expression. siCrtl was used as a control. METTL3 siRNA sequences were shown in Supplementary Table S1. All siRNAs were designed and chemically synthesized by Obio Technology (Obio Technology Co. Ltd., China). siRNAs were transfected into U87MG and U118MG using X-treme GENE^TM HP DNA Transfection Reagent (Roche, Switzerland) according to the manufacturer’s instructions. Cells were harvested for RNA and protein extraction assay at 48 h after the transfection.