Cell culture

The immortalized human oral keratinocyte cell line RT7 was maintained in Keratinocyte-SFM (Gibco BRL, Gaithersburg, MD, USA) supplemented with 25 μg/ml of bovine pituitary extract, 0.5 ng/ml of epidermal growth factor, 100 U/ml of penicillin, and 100 μg/ml of streptomycin. Human epithelial OSCC cell lines; HSC2, HSC3, and HSC4 were provided by Japanese Collection of Research Bioresources Cell Bank. These cell lines were cultured in RPMI-1640 (Nacalai Tesque, Inc., Tokyo, Japan) supplemented with 10% heat inactivated FBS (Biowest, France) and 100 U/ml of penicillin-streptomycin (Sigma-Aldrich, MO, USA). All cell lines were maintained in a humidified incubator at 37°C with 5% CO₂.