Fig. 2.

Inhibition of NO and PGE₂ production by spermidine in LPS-stimulated RAW 264.7 macrophages. The cells were pretreated with the indicated concentrations of spermidine for 1 h prior to incubation with 500 ng/mL LPS for 24 h. The levels of NO (A) and PGE₂ (B) in the culture media were measured by Griess assay and a commercial ELISA kit, respectively. Each value indicates the mean ± SD and is representative of the results obtained from three independent experiments (^#p<0.05 compared to the control; ^*p<0.05 compared to cells cultured with 500 ng/mL LPS). (C) Cell lysates were prepared for Western blot analysis with antibodies specific for murine iNOS and COX-2, and an ECL detection system. (D) The total RNAs were prepared for RT-PCR analysis of the iNOS and COX-2 mRNA expression using the indicated primers. The experiment was repeated thrice, and similar results were obtained. β-actin and GAPDH were used as the internal controls for the Western blot analysis and RT-PCR, respectively. The relative ratios of expression from Western blot analysis and RT-PCR are presented at the bottom of each of the results as relative values of the β-actin and GAPDH expression, respectively.