25 cm3 cell culture flask

To generate PDPC, necrotic areas and blood vessels were removed from the intra-operatively obtained tumor tissue and the latter then separated using a homogenizer. The homogenized tumor material was cultured in 25 cm³ cell culture flasks (Corning, New York, NY, USA) in Dulbecco’s Modified Eagle’s Medium (DMEM) containing 1 g/L glucose, sodium pyruvate, 3.7 g/L NaHCO₃ and L-glutamine and supplemented with 20% v/v heat-inactivated fetal calf serum (FCS), 2 × non-essential amino acids (NEAA, 100× stock, add 10 mL to 500 mL medium) (all from Gibco, Carlsbad, CA, USA) and 1.5% vitamin C (Sigma-Aldrich, St. Louis, MO, USA) at 37 °C, 5% CO₂, and 95% humidity until an adherent cell layer was formed [27 (link)]. A GBM cell line U87MG (CLS, Eppelheim, Germany) was cultured in DMEM supplemented with 10% v/v FCS, 2 × NEAA, 3 mM L-glutamine, 100 U/mL penicillin, and 100 mg/mL streptomycin (Invitrogen, Carlsbad, CA, USA) as a monolayer under the same conditions.

Ethical approval was obtained from the Ethics Committee of the 980th Hospital of the PLA Joint Logistics Support Force. Ibuprofen was obtained from the Institute of Chinese Materia Medica with a concentration of 96% and diluted in DMSO to a final concentration of 2 mM. The solution was filtered through a 0.22-µm filter and stored at 4°C before being further diluted in cell culture medium. The cells (1.5 × 10⁶) were inoculated in to 90% confluency in 25 cm³ cell culture flasks (Corning Inc.). Ibuprofen was treated at different concentrations (0, 2.5, 5, 10, 20 µM) for 24 h [4 (link)]. The control group received only the same amount of DMSO. Cells were measured after treatments. The human gastric cancer cell lines SGC7901, MKN45were maintained at 37°C in 5% CO₂ in DMEM (Gibco) supplemented with 10% FBS, 100 U/mL penicillin, and 0.1 mg/mL streptomycin. All results were validated by conducting two independent experiments, in triplicate.

Human breast adenocarcinoma MCF-7 (ATCC® HTB-22™), MDA-MB-231 (ATCC® HTB-26™), and murine breast adenocarcinoma 4T1 (ATCC® CRL-2539™) cells were obtained from the American Type Culture Collection (ATCC) and maintained at 37 °C in a humidified incubator containing 5 % CO₂. MCF-7 and MDA-MB-231 cells were cultured in DMEM-F12 and 4T1 cells in RPMI-1640, both supplemented with 10 % fetal bovine serum (FBS) and 1 % penicillin-streptomycin (GIBCO) referred as complete DMEM or complete RPMI, respectively, and were routinely grown in 25-cm³ cell culture flasks (CORNING Enterprices, Corning, NY).