Figure 1. Effects of iberin on cell viability of TR146 cells. TR146 cells were seeded on 96-well cell culture plates, grown for two days, and then treated for 24 h with iberin (1.875–30 μM). The vitality of cells was determined using Cell Count Reagent SF. The data are presented as the mean SD of four independent experiments. * = p < 0.05, significantly different from TR146 cells not treated with iberin.

Figure 1. Effects of iberin on cell viability of TR146 cells. TR146 cells were seeded on 96-well cell culture plates, grown for two days, and then treated for 24 h with iberin (1.875–30 μM). The vitality of cells was determined using Cell Count Reagent SF. The data are presented as the mean SD of four independent experiments. * = p < 0.05, significantly different from TR146 cells not treated with iberin.

Figure 2. Effects of iberin on TNF-α-induced production of CXCL10 and IL-6. TR146 cells were grown for 24 h with TNF-α (100 ng/mL) with or without iberin (1.875–15 μM). CXCL10 and IL-6 levels in the supernatant were measured using the ELISA kits indicated in the Materials and Methods section. The data are presented as the mean SD of three independent experiments. * = p < 0.05, significantly different from TNF-α-stimulated TR146 cells without iberin.

Figure 2. Effects of iberin on TNF-α-induced production of CXCL10 and IL-6. TR146 cells were grown for 24 h with TNF-α (100 ng/mL) with or without iberin (1.875–15 μM). CXCL10 and IL-6 levels in the supernatant were measured using the ELISA kits indicated in the Materials and Methods section. The data are presented as the mean SD of three independent experiments. * = p < 0.05, significantly different from TNF-α-stimulated TR146 cells without iberin.

Figure 3. Effects of iberin on VCAM-1, iNOS, and COX-2 expression in TNF-α-stimulated TR146 cells. TR146 cells were pretreated for 1 h with iberin (3.75, 7.5, or 15 μM) before being stimulated with TNF-α (100 ng/mL). After 24 h of stimulation, the lysates were collected. Western blot analysis was used to investigate at the expression of VCAM-1, iNOS, and COX-2. (A) Representative Western blot image of the expression of VCAM-1, iNOS, COX-2, and GAPDH. (B–D) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of thre independent experiments. (* = p < 0.05 vs. TNF-α-stimulated TR146 cells without iberin).

Figure 3. Effects of iberin on VCAM-1, iNOS, and COX-2 expression in TNF-α-stimulated TR146 cells. TR146 cells were pretreated for 1 h with iberin (3.75, 7.5, or 15 μM) before being stimulated with TNF-α (100 ng/mL). After 24 h of stimulation, the lysates were collected. Western blot analysis was used to investigate at the expression of VCAM-1, iNOS, and COX-2. (A) Representative Western blot image of the expression of VCAM-1, iNOS, COX-2, and GAPDH. (B–D) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of thre independent experiments. (* = p < 0.05 vs. TNF-α-stimulated TR146 cells without iberin).

Figure 4. Effects of iberin on HO-1 and NQO1 expression in TNF-α-stimulated TR146 cells. TR146 cells were pretreated for 1 h with iberin (3.75, 7.5, or 15 μM) before being stimulated with TNF-α (100 ng/mL). After 24 h of stimulation, the lysates were collected. Western blot analysis was used to investigate at the expression of HO-1 and NQO1. (A) Representative Western blot image of the expression of HO-1, NQO-1, and GAPDH. (B,C) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of thre independent experiments. (* = p < 0.05 vs. TNF-α-stimulated TR146 cells without iberin).

Figure 4. Effects of iberin on HO-1 and NQO1 expression in TNF-α-stimulated TR146 cells. TR146 cells were pretreated for 1 h with iberin (3.75, 7.5, or 15 μM) before being stimulated with TNF-α (100 ng/mL). After 24 h of stimulation, the lysates were collected. Western blot analysis was used to investigate at the expression of HO-1 and NQO1. (A) Representative Western blot image of the expression of HO-1, NQO-1, and GAPDH. (B,C) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of thre independent experiments. (* = p < 0.05 vs. TNF-α-stimulated TR146 cells without iberin).

Figure 5. The effects of iberin treatment on the activation a NF-κB pathway in TNF-α stimulated TR146 cells. TR146 cells were pretreated with iberin (7.5 or 15 μM) for 1 h before being stimulated with TNF-α for 15, 30, or 60 min, and the phosphorylation of NF-κB p65 and IκB-α was measured using Western immunoblotting. (A) Representative Western blot image of the expression of phospho-NF-κB p65, total NF-κB p65, phospho-IκB-α, and GAPDH. (B,C) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of three independent experiments. (* = p < 0.05).

Figure 5. The effects of iberin treatment on the activation a NF-κB pathway in TNF-α stimulated TR146 cells. TR146 cells were pretreated with iberin (7.5 or 15 μM) for 1 h before being stimulated with TNF-α for 15, 30, or 60 min, and the phosphorylation of NF-κB p65 and IκB-α was measured using Western immunoblotting. (A) Representative Western blot image of the expression of phospho-NF-κB p65, total NF-κB p65, phospho-IκB-α, and GAPDH. (B,C) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of three independent experiments. (* = p < 0.05).

Figure 6. The effect of iberin treatment on the activation of a STAT3 pathway in TNF-α stimulated TR146 cells. TR146 cells were pretreated with iberin (7.5 or 15 μM) for 1 h before being stimulated with TNF-α for 15, 30, or 60 min, and the phosphorylation of STAT3 was evaluated using Western immunoblotting. (A) Representative Western blot image of the expression of phospho-STAT3, total STAT3, and GAPDH. (B) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of three independent experiments. (* = p < 0.05).

Figure 6. The effect of iberin treatment on the activation of a STAT3 pathway in TNF-α stimulated TR146 cells. TR146 cells were pretreated with iberin (7.5 or 15 μM) for 1 h before being stimulated with TNF-α for 15, 30, or 60 min, and the phosphorylation of STAT3 was evaluated using Western immunoblotting. (A) Representative Western blot image of the expression of phospho-STAT3, total STAT3, and GAPDH. (B) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of three independent experiments. (* = p < 0.05).

Figure 7. The effect of iberin treatment on the activation of a p70S6K-S6 pathway in TNF-α stimulated TR146 cells. TR146 cells were pretreated with iberin (7.5 or 15 μM) for 1 h before being stimulated with TNF-α for 15, 30, or 60 min, and the phosphorylation of p70S6K and S6 was measured using Western immunoblotting. (A) Representative Western blot image of the expression of phospho-p70S6K, total p70S6K, phospho-S6, total S6, and GAPDH. (B,C) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of thre independent experiments. (* = p < 0.05).

Figure 7. The effect of iberin treatment on the activation of a p70S6K-S6 pathway in TNF-α stimulated TR146 cells. TR146 cells were pretreated with iberin (7.5 or 15 μM) for 1 h before being stimulated with TNF-α for 15, 30, or 60 min, and the phosphorylation of p70S6K and S6 was measured using Western immunoblotting. (A) Representative Western blot image of the expression of phospho-p70S6K, total p70S6K, phospho-S6, total S6, and GAPDH. (B,C) Quantification of protein expression by densitometry analysis of Western blots. Data are expressed as the mean ± SD of thre independent experiments. (* = p < 0.05).