Induction of iNOS Expression

To induce iNOS expression, subconfluent monolayers were cultured in serum-free medium for 24 h. Growth-arrested cultures were treated with pro-inflammatory cytokines, 100 U/ml interferon γ (IFN-γ) (Sigma-Aldrich, St. Louis, MO, USA), 10 ng/ml interleukin-1 α (IL-1α) (PeproTech, Inc., Rocky Hill, NJ, USA) and 25 ng/ml tumor necrosis factor-α (TNF-α) (R&D Systems, Minneapolis, MN, USA), pro-inflammatory cytokines and 0.1–5 mg/ml water extract of Cnidii Rhizoma or 0.5 mM 1400W (Sigma-Aldrich) in fresh medium without fetal bovine serum. After 48 h, the supernatants were collected and the cells were harvested and lysed as previously described (18 (link)).