Inos inhibitor 1400 w

For inhibition of NO production, cells were treated with 2.5 mM L-NAME (N_ω-Nitro-L-arginine methyl ester hydrochloride, Sigma-Aldrich, St. Louis, MO, USA); for induction of NO production, 2.5 μM SNAP (S-Nitroso-N-acetyl-DL-penicillamine, Sigma-Aldrich) or 2.5 μM GSNO (S-nitrosoglutathione, Sigma-Aldrich) was used. To inhibit ODC1, the rate-limiting enzyme of polyamine synthesis, DMFO (DL-α-Difluoromethylornithine, Sigma-Aldrich) was used at 5 mM. To compensate for the reduced BH₄ level in cancer cells and M2-type macrophages, 20 or 100 μM L-sepiapterin (BH₄ precursor, Sigma-Aldrich or Santa Cruz Biotech. (Santa Cruz, CA, USA)) was used. For iNOS inhibition, iNOS inhibitor (1400 W) was obtained from Cayman Chemical (Ann Arbor, MI, USA) and used at 50 and 100 μM for 2 days [40 (link)]. For inhibition of STAT3, 2.5 μM Stattic (Tocris Biosci., Minneapolis, MN, USA) was used. For inhibition of SMAD3, 25 μM SIS3 (Sigma-Aldrich) was used. For macrophage differentiation/polarization, 100 ng/ml phorbol 12-myristate 13-acetate (PMA, Invivogen, San Diego, CA, USA), 5 ng/ml lipopolysaccharide (LPS, Sigma-Aldrich), 20 ng/ml Interferon-γ (IFN-γ, PeproTech, Rocky Hill, NJ, USA), 20 ng/ml interleukin-4 (IL-4, PeproTech) and 20 ng/ml interleukin-13 (IL-13, Pepro-Tech) were used.