Nih3t3 mouse fibroblast cells

KB human oral squamous carcinoma cells and NIH3T3 mouse fibroblast cells were obtained from the Korean Cell Line Bank (KCLB, Seoul, Korea) and maintained with DMEM medium supplemented with 10% fetal bovine serum (FBS) and 1% antibiotics in a 5% CO₂ incubator operating at 37°C.

NIH3T3 mouse fibroblast cells, U87MG human malignant glioma cells, and HCT116 mouse colon carcinoma cells were obtained from Korean Cell Line Bank (Seoul, Korea). NIH3T3 cells and U87MG cells were cultured in DMEM (Gibco, Grand Island, NY, USA) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS) (Invitrogen) and 1% (v/v) penicillin-streptomycin at 37 °C in a 5% CO₂ incubator. The HCT116 cells were cultured in RPMI1640 (Gibco, Grand Island, NY, USA) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS) (Invitrogen) and 1% (v/v) penicillin-streptomycin.

MDA-MB231 human breast carcinoma cells and NIH3T3 mouse fibroblast cells were obtained from Korea Cell Line Bank Co. (KCLB, Seoul, Korea). MDA-MB231 human breast carcinoma cells and NIH3T3 mouse fibroblast cells were maintained in RPMI1640 and DMEM supplemented with 10 % fetal bovine serum (5 % CO₂ at 37 °C), respectively.
Cell viability test was performed as follows: MDA-MB231 (1 × 10⁴ cell/well) cells were seeded into 96-well plates and then incubated overnight (5 % CO₂ at 37 °C). DOX or nanoparticles were treated to these cells. The CD44 receptor of the cells was blocked with 10 equivalents of free HA 1 h before nanoparticle treatment, and then the cells were washed with PBS. After that, DOX or nanoparticles were added to the cells in the 96-well plates and incubated in a CO₂ incubator (5 % CO₂ at 37 °C) for 4 or 24 h. After that, media were discarded and then replaced with 100 μl of fresh media. To this solution, 30 μl of MTT (5 mg/ml) was added, and then the solution was further incubated in a CO₂ incubator (5 % CO₂ at 37 °C) for 4 h. Since the contents of the generated formazan were proportional to the number of viable cells, the generated formazan was solubilized with DMSO and determined with an automated computer-linked microplate reader (Molecular Device Co., USA) (560 nm test/630 nm reference).