Culture of HaCaT keratinocytes

HaCaT human keratinocytes were seeded at 4×10⁴ cells/well in 12-well culture plates and were maintained in DMEM with 10% FBS at 37°C in a 5% CO₂ atmosphere. After overnight incubation, the medium was exchanged to DMEM without FBS and was incubated further for 24 hr, after which the cells were treated with BSG at final concentrations of 1, 3 and 10 μg/mL. The culture period was adjusted to 1, 3 or 6 hr for evaluation by real-time RT-PCR and to 48 hr for evaluation by western blotting analysis.