Ha cells

Human astrocyte (HA) cells were purchased from ScienCell Research Laboratories (Carlsbad, CA, USA) and grown in RPMI-1640 culture medium (Gibco, Grand Island, NY, USA) with 10% fetal bovine serum (FBS, Gibco, Carlsbad, CA, USA). Human glioma cell lines (U87 and U251) and human embryonic kidney (HEK) 293 T cells were purchased from Shanghai Institutes for Biological Sciences Cell Resource Center, and grown in Dulbecco’s modified Eagle medium(DMEM)/high glucose with 10% FBS. All cells were maintained in a humidified incubator at 37 °C with 5% CO₂. Human glioma tissues and normal brain tissues (NBTs) were collected from patients at the Department of Neurosurgery of Shengjing Hospital of China Medical University (n = 5). All the tissue samples were immediately frozen in liquid nitrogen after surgical resection, and stored at −80 °C until use. Informed consent was obtained from all patients and the study was approved by the Ethics Committee of Shengjing Hospital of China Medical University. Glioma tissue samples were classified into five groups according to the 2007 WHO classification by neuropathologists: Grade I (n = 5), Grade II (n = 5), Grade III (n = 8) and Grade IV (n = 8).

The human GBM cell lines LN229, U118, U87, U251, and T98G were purchased from the American Type Culture Collection (ATCC, USA). All GBM cell lines were cultured with Dulbecco’s Modified Eagle’s Medium (DMEM, Gibco, USA) containing 10% fetal bovine serum (FBS, Gibco, USA) and antibiotics (100 units/mL penicillin and 100 mg/mL streptomycin, Gibco, USA) in a 37 °C incubator with 5% CO₂ and 100% humidity. The human astrocyte cell line HA cells (ScienCell, Cat. No. #1800) was cultured in basal astrocyte medium (ScienCell, Cat. No. #1801) supplemented with 2% FBS and 1% astrocyte growth supplement (ScienCell, Cat. No. #1852). Human umbilical vein endothelial cells (HUVEC, Cat. No. #8000, ScienCell) were cultured with Endothelial Cell Medium (ECM, Cat. No. #1001, ScienCell) containing 1% endothelial cell growth supplement (ECGS, Cat. No. #1052, ScienCell), 5% FBS (Cat. No. #0025, ScienCell) and 1% antibiotic solution (P/S, Cat. No #0503, ScienCell) in a 37 °C incubator.