Fig. 5.

Effect of carnosol on LPS-induced NO release and on protein expressions of iNOS and COX-2 in RAW 264.7 cells. NO level was determined by Griess reaction as described in Methods, in supernatants from RAW 264.7 cells (A). Cells were treated with 1 μg/ml of LPS alone, or with LPS plus different concentrations (1, 2, 5 μM) of carnosol, at 37°C for 24 h. Equal amounts of total protein (20 μg/lane) were subjected to 8% SDS-PAGE, and the expression of iNOS, COX-2, STAT3 and p-STAT3 were detected by Western blotting using specific antibodies in RAW 264.7 cells (B and E). β-actin protein was used here as an internal control. Similar results were obtained from at least three different sets of experiment. Cell viability was evaluated using a MTT assay as described in Methods. RAW 264.7 cells (C). RAW 264.7 cells were transfected with a p-STAT3-Luc plasmid (5×STAT3), and then treated with LPS (1 μg/ml) either alone or with KRICT NO.9 (1, 2, 5 μM) for 37°C for 24 h. Luciferase activity was then determined as described in Methods (D). The activation of STAT3 was investigated using EMSA (F). Values represent means ± SD for three independent experiments performed in triplicate. ^*p<0.05. ^# indicates significantly different from LPS treated group (p<0.05).