Figures & data
Figure 1. Chemical structure of 3-hydroxy-4,7-megastigmadien-9-one (comp).
Figure 2. Effects of comp on cell viability of bone marrow-derived dendritic cells (BMDCs). BMDCs were treated with comp (1–50 μM) for 18 h and viability was measured using MTT assay. Results shown are the mean ± SD of an experiment done in triplicate and are representative of three separate experiments. Comp, 3-hydroxy-4,7-megastigmadien-9-one.
Figure 3. Inhibitory effects of comp on pro-inflammatory cytokine production in CpG DNA-stimulated bone marrow-derived dendritic cells (BMDCs). BMDCs were treated with comp at the indicated doses for 1 h before stimulation with CpG DNA (1 μM). Enzyme-linked immunosorbent assay (ELISA) was used to measure the concentrations of murine IL-12 p40 (A), IL-6 (B) and TNF-α (C) in the culture supernatants. Data are representative of three independent experiments. Comp, 3-hydroxy-4,7-megastigmadien-9-one. *p < 0.05, **p < 0.01 vs. comp-untreated cells in the presence of CpG DNA.
Figure 4. Effects of comp on the phosphorylation of MAPK and IκBα by CpG DNA-stimulated BMDCs. (A) Cells were pretreated with or without comp (50 μM) for 1 h before stimulation with CpG DNA (1 μM). Total cell lysate was obtained at the indicated time intervals. Western blot analysis was performed on the cell lysate to assess phosphorylation of ERK, JNK, p38 and IκBα. β-Actin was taken as the loading control. Data are representative of three independent experiments. (B) Phosphorylation of ERK, JNK, p38 and IκBα protein expression was quantified using scanning densitometry, and the band intensities were normalized by that of β-actin. Comp: 3-hydroxy-4,7-megastigmadien-9-one. *p < 0.05 vs. comp-untreated cells in the presence of CpG DNA.
Figure 5. Effects of comp on AP-1 reporter activity in HEK293T cells. HEK293T cells were transfected with empty vector (pcDNA3) or a TLR9-expressing plasmid (pcDNA3-mTLR9) and then treated with comp for 1 h before stimulation with CpG DNA (1 μM). Cell lysates were prepared, luciferase activity was assayed by the dual luciferase reporter assay and the results were expressed as relative luciferase. Data are representative of three independent experiments. Comp, 3-hydroxy-4,7-megastigmadien-9-one. *p < 0.05 vs. comp-untreated cells in the presence of CpG DNA.
Figure 6. Effects of comp on NF-κB reporter activity in HECK293T cells. HEK293T cells were transfected with empty vector (pcDNA) or a TLR9-expressing plasmid (pcDNA3-mTLR9) and then treated with comp for 1 h before stimulation with CpG DNA (1 μM). Cell lysates were prepared, luciferase activity was assayed by the dual luciferase reporter assay and the results were expressed as relative luciferase. Data are representative of three independent experiments. Comp, 3-hydroxy-4,7-megastigmadien-9-one. *p < 0.05 vs. comp-untreated cells in the presence of CpG DNA.
