Fig. 1. Notoginseng suppresses LPS-induced TNF-α and IL-6 production by DC2.4 cells. DCs were treated with 0, 5, 25 or 50 μg/ml of notoginseng and/or LPS (1 μg/ml). Supernatants were collected after 24 h and assayed for TNF-α (A) and IL-6 (B) production as described in Section 2. Data represents mean ± S.E.M. of three samples. Hash (#) indicates significant differences between LPS-stimulated and unstimulated cells; asterisk (*) indicates significant differences between the LPS-stimulated control- and notoginseng-treated samples (p < 0.05). Data are representative of three independent experiments.

Fig. 2. Variable notoginseng treatment affects LPS-induced production of TNF-α by DC2.4 cells. Cells were treated with 50 μg/ml notoginseng (NG) at three different time points: −24, 0 and 8 h, relative to LPS stimulation. Supernatants were collected 24 h after LPS addition and TNF-α levels measured by ELISA. Data represents mean ± S.E.M. of three samples. Hash (#) indicates significant differences between LPS-stimulated and unstimulated cells; asterisk (*) indicates significant differences between LPS-stimulated controls and LPS-stimulated, notoginseng-treated samples (p < 0.05). Data are representative of three separate experiments.

Fig. 3. Notoginseng selectively reduces TLR ligand-induced production of TNF-α and IL-6 by DC2.4 cells. DCs were unstimulated or stimulated with LPS, CpG or poly(I:C) and concurrently treated with 50 μg/ml notoginseng (NG) for 24 h. Supernatants were collected and assayed for TNF-α and IL-6 production by ELISA. Data represents mean ± S.E.M. of three samples. Hash (#) indicates significant differences between stimulated and unstimulated cells; asterisk (*) indicates significant differences between the TLR ligand-stimulated control- and notoginseng-treated samples (p < 0.05). Data are representative of three independent experiments.

Fig. 4. Notoginseng inhibits the expression of costimulatory molecules on DC2.4 cells following stimulation with TLR ligands. Cells were unstimulated (dashed line) or stimulated (black line) with LPS, CpG or poly(I:C) and concomitantly treated with 50 μg/ml notoginseng (gray histogram) for 24 h. Representative histograms demonstrating the cell surface expression of CD40 (A) and CD86 (B) on activated DC2.4 cells were determined by FACS analysis. Data are representative of three independent experiments, each consisting of a minimum of three samples per treatment group.

Fig. 5. The production of TNF-α and IL-6 by DC2.4 cells is selectively suppressed by the whole notoginseng extract and purified ginsenosides. LPS-stimulated cells were treated with 50 μg/ml whole notoginseng extract (NG) or purified Rb1 and/or Rg1 ginsenosides. Supernatants were collected after 24 h and assayed for TNF-α (A) and IL-6 (B) production by ELISA. Data represents mean ± S.E.M. of three samples. Statistically significant differences (p < 0.05) between treatment groups are indicated by different letters while groups sharing letters are not significantly different. Data are representative of two separate experiments.

Notoginseng selectively alters TLR-induced costimulatory molecule expression on DC2.4 cells

DC2.4 cells were not stimulated or stimulated with LPS (1 μg/ml), CpG (0.5 μM) or poly(I:C) (12.5 μg/ml) and concomitantly treated with notoginseng (50 μg/ml) for 24 h.