Characterization of AD-293 Cell Line

Cells from the AD-293 cell line, a derivative of the HEK293 cell line with improved adherence and plaque formation properties, were purchased from Stratagene (La Jolla, CA, USA). The protocols were adopted and modified from our previous report.^{25 (link)} All compounds were dissolved in DMSO at 10 mM as a stock solution. The final concentration of DMSO was 0.1% in the culture medium. AD-293 cells bearing luciferase reporters with promoter regions of IL-6/STAT3 and TNF-α/NF-κB in pGL4.20 vector (Promega) were maintained in DMEM high glucose medium supplemented with 10% FBS and 1% penicillin streptomycin in the presence of 1 μg mL⁻¹ puromycin. AD-293 cells overexpressing TLR4 and stably transfected with NF-κB reporter were maintained in DMEM high glucose medium in the presence of 1 μg mL⁻¹ puromycin and 10 μg mL⁻¹ blasticidin. All cells were maintained in a humidified incubator at 37 °C in 95% air and 5% CO₂.