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The saponin DT-13 attenuates tumor necrosis factor-α-induced vascular inflammation associated with Src/NF-кB/MAPK pathway modulation.

Zhang Y, Sun M, Han Y, Zhai K, Tang Y, Qin X, Cao Z, Yu B, Kou J - Int. J. Biol. Sci. (2015)

Bottom Line: This study aimed to explore the effect of DT-13 (25(R,S)-ruscogenin- 1-O- [β-d-glucopyranosyl- (1→2)][β-d-xylopyranosyl-(1→3)]-β -d- fucopyranoside) on tumor necrosis factor (TNF)-α-induced vascular inflammation and the potential molecular mechanisms.These findings suggest that DT-13 abrogates vascular inflammation by down-regulating adhesion molecules associated with modulating the NF-кB, p38MAPK, Src signaling pathways, and NF-κB binding site is at least one of the targets of DT-13.This study provides novel information regarding the mechanism by which DT-13 exerts its effects on vascular inflammation, which is important for the onset and progression of various diseases.

View Article: PubMed Central - PubMed

Affiliation: Jiangsu Key Laboratory of TCM Evaluation and Translational Research, Department of Complex Prescription of TCM, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China.

ABSTRACT
This study aimed to explore the effect of DT-13 (25(R,S)-ruscogenin- 1-O- [β-d-glucopyranosyl- (1→2)][β-d-xylopyranosyl-(1→3)]-β -d- fucopyranoside) on tumor necrosis factor (TNF)-α-induced vascular inflammation and the potential molecular mechanisms. In vitro, DT-13 suppressed TNF-α-induced adhesion and migration of human umbilical vein endothelial cells (HUVECs) by inhibiting the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). DT-13 markedly suppressed NF-кB p65 phosphorylation, and when NF-кB p65 was over-expressed, the inhibitory effect of DT-13 on adhesion molecular decreased. DT-13 also suppressed TNF-α induced luciferase activities of ICAM-1 and VCAM-1 promoter containing NF-κB binding sites. Furthermore, DT-13 markedly suppressed p38 phosphorylation and Src degradation induced by TNF-α, whereas had no significant effect on ERK and JNK activation. In vivo, DT-13 at 4 mg/kg prevented vascular inflammation and the expression of adhesion molecules induced by TNF-α in mice. These findings suggest that DT-13 abrogates vascular inflammation by down-regulating adhesion molecules associated with modulating the NF-кB, p38MAPK, Src signaling pathways, and NF-κB binding site is at least one of the targets of DT-13. This study provides novel information regarding the mechanism by which DT-13 exerts its effects on vascular inflammation, which is important for the onset and progression of various diseases.

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DT-13 inhibited TNF-α-induced ICAM-1 & VCAM-1 activity dependent with NF-κB pathway. (A). DT-13 inhibited TNF-α-induced p-65 phosphorylation in HUVECs. HUVECs were pretreated with DT-13 (0.01, 0.1 or 1 μM) for 1 h followed by TNF-α (10 ng/mL) exposure. Expression and activation of p-65 were detected by western blotting. ## P<0.01 νs. the control group; * P<0.05,** P<0.01 νs. the TNF-α group. (B) & (C). The over-expression of p65 overcomes the inhibitory effect of DT-13 on ICAM-1 (B) and VCAM-1 (C) expression. (D) Human ICAM-1 promoter or (E) VCAM-1 promoter were ligated into pGL3 basic luciferase vectors. The binding sites for transcription factors are also shown. Right: Cells were transfected with indicated forms of (D) ICAM-1 or (E) VCAM-1 promoter for 24 h, and then incubated with DT-13 for 1 h followed by TNF-α (10 ng/mL) stimulation for 4 h. The promoter activity is represented by the level of luciferase activity indicated by relative light units (RLU). pRL-TK plasmid was also transfected into cells and used as an internal control. The data represent the mean ± SD of three experiments. # P<0.05, ## P<0.01 νs. the control group; * P<0.05,** P<0.01 νs. the TNF-α group.
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Figure 3: DT-13 inhibited TNF-α-induced ICAM-1 & VCAM-1 activity dependent with NF-κB pathway. (A). DT-13 inhibited TNF-α-induced p-65 phosphorylation in HUVECs. HUVECs were pretreated with DT-13 (0.01, 0.1 or 1 μM) for 1 h followed by TNF-α (10 ng/mL) exposure. Expression and activation of p-65 were detected by western blotting. ## P<0.01 νs. the control group; * P<0.05,** P<0.01 νs. the TNF-α group. (B) & (C). The over-expression of p65 overcomes the inhibitory effect of DT-13 on ICAM-1 (B) and VCAM-1 (C) expression. (D) Human ICAM-1 promoter or (E) VCAM-1 promoter were ligated into pGL3 basic luciferase vectors. The binding sites for transcription factors are also shown. Right: Cells were transfected with indicated forms of (D) ICAM-1 or (E) VCAM-1 promoter for 24 h, and then incubated with DT-13 for 1 h followed by TNF-α (10 ng/mL) stimulation for 4 h. The promoter activity is represented by the level of luciferase activity indicated by relative light units (RLU). pRL-TK plasmid was also transfected into cells and used as an internal control. The data represent the mean ± SD of three experiments. # P<0.05, ## P<0.01 νs. the control group; * P<0.05,** P<0.01 νs. the TNF-α group.

Mentions: TNF-α activates NF-кB, which subsequently translocates into the nucleus, and promotes the gene transcription of adhesion molecules 33. We therefore evaluated the influence of DT-13 on TNF-⍺ activated NF-кB using Western blot analysis. HUVECs exposed to TNF-α (10 ng/mL) exhibited dramatic increases in NF-кB p65 and IкB-α phosphorylation, whereas the expression of total p65 and IкB-α remained unchanged. DT-13 inhibited p65 (Fig. 3A) phosphorylation, with maximum inhibitory rates of 43.3%. Meanwhile, DT-13 inhibited the IкB-α (Supplementary Figure S2.A&B) phosphorylation with maximum inhibitory rates of 41.2%.


The saponin DT-13 attenuates tumor necrosis factor-α-induced vascular inflammation associated with Src/NF-кB/MAPK pathway modulation.

Zhang Y, Sun M, Han Y, Zhai K, Tang Y, Qin X, Cao Z, Yu B, Kou J - Int. J. Biol. Sci. (2015)

DT-13 inhibited TNF-α-induced ICAM-1 & VCAM-1 activity dependent with NF-κB pathway. (A). DT-13 inhibited TNF-α-induced p-65 phosphorylation in HUVECs. HUVECs were pretreated with DT-13 (0.01, 0.1 or 1 μM) for 1 h followed by TNF-α (10 ng/mL) exposure. Expression and activation of p-65 were detected by western blotting. ## P<0.01 νs. the control group; * P<0.05,** P<0.01 νs. the TNF-α group. (B) & (C). The over-expression of p65 overcomes the inhibitory effect of DT-13 on ICAM-1 (B) and VCAM-1 (C) expression. (D) Human ICAM-1 promoter or (E) VCAM-1 promoter were ligated into pGL3 basic luciferase vectors. The binding sites for transcription factors are also shown. Right: Cells were transfected with indicated forms of (D) ICAM-1 or (E) VCAM-1 promoter for 24 h, and then incubated with DT-13 for 1 h followed by TNF-α (10 ng/mL) stimulation for 4 h. The promoter activity is represented by the level of luciferase activity indicated by relative light units (RLU). pRL-TK plasmid was also transfected into cells and used as an internal control. The data represent the mean ± SD of three experiments. # P<0.05, ## P<0.01 νs. the control group; * P<0.05,** P<0.01 νs. the TNF-α group.
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Figure 3: DT-13 inhibited TNF-α-induced ICAM-1 & VCAM-1 activity dependent with NF-κB pathway. (A). DT-13 inhibited TNF-α-induced p-65 phosphorylation in HUVECs. HUVECs were pretreated with DT-13 (0.01, 0.1 or 1 μM) for 1 h followed by TNF-α (10 ng/mL) exposure. Expression and activation of p-65 were detected by western blotting. ## P<0.01 νs. the control group; * P<0.05,** P<0.01 νs. the TNF-α group. (B) & (C). The over-expression of p65 overcomes the inhibitory effect of DT-13 on ICAM-1 (B) and VCAM-1 (C) expression. (D) Human ICAM-1 promoter or (E) VCAM-1 promoter were ligated into pGL3 basic luciferase vectors. The binding sites for transcription factors are also shown. Right: Cells were transfected with indicated forms of (D) ICAM-1 or (E) VCAM-1 promoter for 24 h, and then incubated with DT-13 for 1 h followed by TNF-α (10 ng/mL) stimulation for 4 h. The promoter activity is represented by the level of luciferase activity indicated by relative light units (RLU). pRL-TK plasmid was also transfected into cells and used as an internal control. The data represent the mean ± SD of three experiments. # P<0.05, ## P<0.01 νs. the control group; * P<0.05,** P<0.01 νs. the TNF-α group.
Mentions: TNF-α activates NF-кB, which subsequently translocates into the nucleus, and promotes the gene transcription of adhesion molecules 33. We therefore evaluated the influence of DT-13 on TNF-⍺ activated NF-кB using Western blot analysis. HUVECs exposed to TNF-α (10 ng/mL) exhibited dramatic increases in NF-кB p65 and IкB-α phosphorylation, whereas the expression of total p65 and IкB-α remained unchanged. DT-13 inhibited p65 (Fig. 3A) phosphorylation, with maximum inhibitory rates of 43.3%. Meanwhile, DT-13 inhibited the IкB-α (Supplementary Figure S2.A&B) phosphorylation with maximum inhibitory rates of 41.2%.

Bottom Line: This study aimed to explore the effect of DT-13 (25(R,S)-ruscogenin- 1-O- [β-d-glucopyranosyl- (1→2)][β-d-xylopyranosyl-(1→3)]-β -d- fucopyranoside) on tumor necrosis factor (TNF)-α-induced vascular inflammation and the potential molecular mechanisms.These findings suggest that DT-13 abrogates vascular inflammation by down-regulating adhesion molecules associated with modulating the NF-кB, p38MAPK, Src signaling pathways, and NF-κB binding site is at least one of the targets of DT-13.This study provides novel information regarding the mechanism by which DT-13 exerts its effects on vascular inflammation, which is important for the onset and progression of various diseases.

View Article: PubMed Central - PubMed

Affiliation: Jiangsu Key Laboratory of TCM Evaluation and Translational Research, Department of Complex Prescription of TCM, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China.

ABSTRACT
This study aimed to explore the effect of DT-13 (25(R,S)-ruscogenin- 1-O- [β-d-glucopyranosyl- (1→2)][β-d-xylopyranosyl-(1→3)]-β -d- fucopyranoside) on tumor necrosis factor (TNF)-α-induced vascular inflammation and the potential molecular mechanisms. In vitro, DT-13 suppressed TNF-α-induced adhesion and migration of human umbilical vein endothelial cells (HUVECs) by inhibiting the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). DT-13 markedly suppressed NF-кB p65 phosphorylation, and when NF-кB p65 was over-expressed, the inhibitory effect of DT-13 on adhesion molecular decreased. DT-13 also suppressed TNF-α induced luciferase activities of ICAM-1 and VCAM-1 promoter containing NF-κB binding sites. Furthermore, DT-13 markedly suppressed p38 phosphorylation and Src degradation induced by TNF-α, whereas had no significant effect on ERK and JNK activation. In vivo, DT-13 at 4 mg/kg prevented vascular inflammation and the expression of adhesion molecules induced by TNF-α in mice. These findings suggest that DT-13 abrogates vascular inflammation by down-regulating adhesion molecules associated with modulating the NF-кB, p38MAPK, Src signaling pathways, and NF-κB binding site is at least one of the targets of DT-13. This study provides novel information regarding the mechanism by which DT-13 exerts its effects on vascular inflammation, which is important for the onset and progression of various diseases.

No MeSH data available.


Related in: MedlinePlus