Animal studies were approved by the Animal Ethical and Welfare Committee of Sun Yat-sen University. The hind flanks of 4-week-old female BALB/c-nu/nu mice were subcutaneously inoculated with 2 × 105 GSC-1 cells. The mice were randomly divided into 6 groups after tumor cell injection and were injected intraperitoneally once per day with vehicle, 20 mg/kg dbcAMP (Sigma-Aldrich, USA), 40 mg/kg luteolin (Selleck, USA), 20 mg/kg MS275 (Selleck), 100 mg/kg temozolomide (Topscience, USA) or a combination of these drugs. Tumor diameters were measured every other day with a caliper, and tumor volumes were estimated using the following formula: width2 × length/2 = V (mm3).
The orthotopic implantation of glioma cells was performed using 1 × 105 GSCs. In brief, cells were injected 2 mm lateral and 0.5 mm anterior to the bregma and 2.5 mm below the skull of 4-week-old athymic nude mice. The mice were randomly divided into four groups and injected intraperitoneally with vehicle, 100 mg/kg RGFP109 (Selleck) plus 40 mg/kg luteolin, 100 mg/kg temozolomide or a combination once per day for 14 days (n = 8 animals for each group). The mice were monitored daily and killed when neurological symptoms were observed. Their brains were then dissected and fixed in formalin for H&E staining.
The orthotopic implantation of glioma cells was performed using 1 × 105 GSCs. In brief, cells were injected 2 mm lateral and 0.5 mm anterior to the bregma and 2.5 mm below the skull of 4-week-old athymic nude mice. The mice were randomly divided into four groups and injected intraperitoneally with vehicle, 100 mg/kg RGFP109 (Selleck) plus 40 mg/kg luteolin, 100 mg/kg temozolomide or a combination once per day for 14 days (n = 8 animals for each group). The mice were monitored daily and killed when neurological symptoms were observed. Their brains were then dissected and fixed in formalin for H&E staining.