Transient Transfection and EDA-A1 CM Preparation

HaCaT or HEK293 cells were cultured in DMEM containing 10% FBS and maintained in an incubator containing 5% CO₂ at 37 °C. Transient transfection in HaCaT or HEK293 cells with plasmids or SNAP23 siRNA were carried out using Lipo8000™ reagent (Beyotime) by following the manufacturer’s instruction. EDA-A1 CM was prepared as described previously [14 (link)]. Briefly, HEK293 cells (2 × 10⁸) were transfected with 100 µg of EDA-A1 expressing plasmid for 16 h and then changed to culture in DMEM containing 1% FBS. After 24 h, the medium was collected and concentrated tenfold using a Centricon Plus-10 filter (Millipore) and stored at − 80 °C until use.