Triptolide attenuates lipopolysaccharide-induced inflammatory responses in human endothelial cells: involvement of NF-κB pathway

Figures

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  Fig. 1. The effects of triptolide on the viability of human umbilical vein endothelial cells (HUVECs). HUVECs were treated with triptolide (25, 50 or 100 nM) for the indicated times. Cell viability was determined by an MTT assay. Data are expressed as the mean ± standard error of the mean. *p < 0.05 and **p < 0.01, compared with the corresponding control group

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  Fig. 2. The effects of triptolide on lipopolysaccharide (LPS)-induced production of pro-inflammatory (a) TNF-α, (b) IL-6 and (c) MCP-1 in human umbilical vein endothelial cells (HUVECs). HUVECs were treated with LPS (1 μg/ml) in the presence or absence of triptolide (25, 50 or 100 nM) for 6 h. The ELISA assay was used to measure the concentrations of pro-inflammatory cytokines and chemokines in the culture medium. Data are expressed as the mean ± standard error of the mean. *p < 0.05 and **p < 0.01, compared with LPS treatment alone. #p < 0.05, compared with the control group

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  Fig. 3. The effects of triptolide on lipopolysaccharide (LPS)-induced (a) ICAM-1 and (b) VCAM-1 expression in human umbilical vein endothelial cells (HUVECs). HUVECs were treated with LPS (1 μg/ml) in the presence or absence of triptolide (25, 50 or 100 nM) for 6 h. The cell-based ELISA assay was used to quantify the expression of ICAM-1 and VCAM-1 in HUVECs. Data are expressed as the mean ± standard error of the mean. *p < 0.05 and **p < 0.01, compared with LPS treatment alone. #p < 0.05, compared with the control group

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  Fig. 4. The effects of triptolide on lipopolysaccharide (LPS)-induced monocyte adhesion to human umbilical vein endothelial cells (HUVECs). HUVECs were treated with LPS (1 μg/ml) in the presence or absence of triptolide (25, 50 or 100 nM) for 6 h, and subsequently co-cultured with fluorescence-labeled THP-1 cells for 30 min. (a) Representative images of fluorescence staining of monocyte adhesion to HUVECs. Magnification, × 200. HUVECs were exposed to (a) vehicle (control), (b) LPS or LPS with triptolide at the concentration of (c) 25, (d) 50 or (e) 100 nM prior to incubation with fluorescence-labeled THP-1 cells. (b) The adhesion ability of THP-1 cells was assayed using a spectrofluorometer. Data are expressed as the mean ± standard error of the mean. *p < 0.05 and **p < 0.01, compared with LPS treatment alone. #p < 0.05, compared with the control group

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  Fig. 5. The effects of triptolide on lipopolysaccharide (LPS)-induced changes in the protein levels of (a and b) phosphorylated IκBα (p-IκBα) and (c and d) IκBα in human umbilical vein endothelial cells (HUVECs). Full-length blots for p-IκBα and IκBα are presented in Additional file Data. HUVECs were treated with LPS (1 μg/ml) in the presence or absence of triptolide (25, 50 or 100 nM) for 30 min. Cell lysates were subjected to immunoblot analysis using the antibodies against p-IκBα (Ser32) and IκBα. The data are presented as the relative protein expression normalized to β-actin expression. Data are expressed as the mean ± standard error of the mean. *p < 0.05 and **p < 0.01, compared with LPS treatment alone. #p < 0.05, compared with the control group

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  Fig. 6. The effects of triptolide on lipopolysaccharide (LPS)-induced increases in NF-κB p65 DNA binding activity in human umbilical vein endothelial cells (HUVECs). HUVECs were treated with LPS (1 μg/ml) in the presence or absence of triptolide (25, 50 or 100 nM) for 30 min. The DNA binding activity of NF-κB p65 in HUVECs was determined using a non-radioactive, ELISA-based assay kit. Data are expressed as the mean ± standard error of the mean. *p < 0.05 and **p < 0.01, compared with LPS treatment alone. #p < 0.05, compared with the control group

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  Fig. 7. The effects of triptolide on lipopolysaccharide (LPS)-induced NF-κB transcriptional activation in human umbilical vein endothelial cells (HUVECs). HUVECs were treated with LPS (1 μg/mL) in the presence or absence of triptolide (25, 50, or 100 nM) for 1 h. The NF-κB transcriptional activity in HUVECs was determined using a dual luciferase reporter assay system as described in the method section. The data were presented as a ratio of firefly to Renilla luciferase activities. Data are expressed as the mean ± standard error of the mean. *p < 0.05, **p < 0.01 compared with LPS treatment alone. #p < 0.05 compared with control group