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Dimethylfumarate protects against TNF-α-induced secretion of inflammatory cytokines in human endothelial cells.

Gerhardt S, König V, Doll M, Hailemariam-Jahn T, Hrgovic I, Zöller N, Kaufmann R, Kippenberger S, Meissner M - J Inflamm (Lond) (2015)

Bottom Line: Furthermore, we found that DMF slightly inhibited the early degradation of IκBα.This action is regulated by reduced p65 activity and nuclear translocation, which can be explained in part by the reduced early degradation of IκBα and more important the reduced phosphorylation of p65 at Serine 536.These effects were independent of the p38, PI3K and p42/44 signaling pathways.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, Venereology and Allergology, Goethe-University, Theodor-Stern-Kai 7, D-60590 Frankfurt am Main, Germany ; Department of Cardiology, Gutenberg-University, Mainz, Germany.

ABSTRACT

Background: Inflammation, angiogenesis and oxidative stress have been implicated in the pathogenesis of various vascular diseases. Recent evidence suggests that dimethylfumarate (DMF), an antiposriatic and anti-multiple sclerosis agent, possesses anti-inflammatory, anti-oxidative and anti-angiogenic properties. Here, we analyze the influence of DMF on TNF-α-induced expression of the important pro-inflammatory and pro-atherogenic chemokine MCP-1 and investigate the underlying mechanisms of this expression.

Findings: We analyzed constitutive and TNF-α-induced expression of MCP-1 in human umbilical vascular endothelial cells (HUVEC) +/- DMF treatment via enzyme-linkes immunosorbent assay (ELISA). DMF significantly inhibited the protein expression levels in a time- and concentration-dependent manner. Furthermore, MCP-1 mRNA expression was also reduced in response to DMF, as demonstrated by RT-PCR. Thus, the regulation occurs at the transcriptional level. Interestingly, DMF prolonged the TNF-α-induced p38 and JNK phosphorylation in HUVEC, as demonstrated by Western blot analysis; however, the p38 and JNK inhibitor SB203580 did not affect the DMF-conveyed suppression of TNF-α-induced MCP-1 expression. DMF suppressed the TNF-α-induced nuclear translocation and phosphorylation (Serine 536) of p65 in these cells. These results were additionally approved by p65 luciferase promoter assays. Furthermore, we found that DMF slightly inhibited the early degradation of IκBα. In addition, we verified our results using other important inflammatory cytokines such as CCL-5, PDGF-BB, GM-CSF and IL-6.

Conclusion: DMF suppresses various TNF-α-induced pro-inflammatory and pro-atherogenic cytokines/chemokines in human endothelial cells. This action is regulated by reduced p65 activity and nuclear translocation, which can be explained in part by the reduced early degradation of IκBα and more important the reduced phosphorylation of p65 at Serine 536. These effects were independent of the p38, PI3K and p42/44 signaling pathways. As a result, DMF might be suitable for treating patients with vascular diseases.

No MeSH data available.


Related in: MedlinePlus

Analysis of constitutive and TNF-α-induced MCP-1 expression during the treatment with DMF in HUVECs. We assayed the MCP-1 protein content in culture supernatants by MCP-1 ELISA according to the manufacturer’s instructions. a HUVECs were mock-treated (solvent only) or treated with DMF at the indicated concentrations for 24 h. b HUVECs were mock-treated (solvent only) or treated with DMF (80 μM) for the indicated times. c HUVECs were mock-treated (solvent only), treated with TNF-α (20 ng/ml) or DMF at the indicated concentrations + TNF-α for 24 h. d HUVECs were mock-treated (solvent only) or treated with TNF-α (20 ng/ml) or DMF (80 μM) + TNF-α for the indicated times. The mean values from three triplicate experiments are presented as the mean ± SEM. We analyzed the data using the Student’s t-test. *p < 0.05
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Fig1: Analysis of constitutive and TNF-α-induced MCP-1 expression during the treatment with DMF in HUVECs. We assayed the MCP-1 protein content in culture supernatants by MCP-1 ELISA according to the manufacturer’s instructions. a HUVECs were mock-treated (solvent only) or treated with DMF at the indicated concentrations for 24 h. b HUVECs were mock-treated (solvent only) or treated with DMF (80 μM) for the indicated times. c HUVECs were mock-treated (solvent only), treated with TNF-α (20 ng/ml) or DMF at the indicated concentrations + TNF-α for 24 h. d HUVECs were mock-treated (solvent only) or treated with TNF-α (20 ng/ml) or DMF (80 μM) + TNF-α for the indicated times. The mean values from three triplicate experiments are presented as the mean ± SEM. We analyzed the data using the Student’s t-test. *p < 0.05

Mentions: First, we analyzed the effect of DMF on constitutive MCP-1 expression in HUVECs. DMF inhibited MCP-1 expression significantly in a time- and concentration-dependent manner (Fig. 1a,b). The concentrations and incubation times that we adopted have been published recently and have been proven to have no cytotoxic effects in HUVEC [14]. DMF in concentrations less than 20 μM reduced the MCP-1 expression level by almost 50 %; within 3 h of treatment, the MCP-1 expression was significantly reduced compared with that of the matching controls. To determine whether DMF was also able to reduce TNF-α-induced MCP-1 expression, we analyzed the effects of the combinational treatment with DMF and TNF-α (20 ng/ml) in a time- and concentration-dependent manner. We found that 80 μM DMF reduced the TNF-α-induced MCP-1 expression by more than 50 % (Fig. 1c). A first significant reduction of MCP-1 expression could be observed as early as 6 h after treatment with 80 μM DMF and TNF-α (Fig. 1d).Fig. 1


Dimethylfumarate protects against TNF-α-induced secretion of inflammatory cytokines in human endothelial cells.

Gerhardt S, König V, Doll M, Hailemariam-Jahn T, Hrgovic I, Zöller N, Kaufmann R, Kippenberger S, Meissner M - J Inflamm (Lond) (2015)

Analysis of constitutive and TNF-α-induced MCP-1 expression during the treatment with DMF in HUVECs. We assayed the MCP-1 protein content in culture supernatants by MCP-1 ELISA according to the manufacturer’s instructions. a HUVECs were mock-treated (solvent only) or treated with DMF at the indicated concentrations for 24 h. b HUVECs were mock-treated (solvent only) or treated with DMF (80 μM) for the indicated times. c HUVECs were mock-treated (solvent only), treated with TNF-α (20 ng/ml) or DMF at the indicated concentrations + TNF-α for 24 h. d HUVECs were mock-treated (solvent only) or treated with TNF-α (20 ng/ml) or DMF (80 μM) + TNF-α for the indicated times. The mean values from three triplicate experiments are presented as the mean ± SEM. We analyzed the data using the Student’s t-test. *p < 0.05
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4525722&req=5

Fig1: Analysis of constitutive and TNF-α-induced MCP-1 expression during the treatment with DMF in HUVECs. We assayed the MCP-1 protein content in culture supernatants by MCP-1 ELISA according to the manufacturer’s instructions. a HUVECs were mock-treated (solvent only) or treated with DMF at the indicated concentrations for 24 h. b HUVECs were mock-treated (solvent only) or treated with DMF (80 μM) for the indicated times. c HUVECs were mock-treated (solvent only), treated with TNF-α (20 ng/ml) or DMF at the indicated concentrations + TNF-α for 24 h. d HUVECs were mock-treated (solvent only) or treated with TNF-α (20 ng/ml) or DMF (80 μM) + TNF-α for the indicated times. The mean values from three triplicate experiments are presented as the mean ± SEM. We analyzed the data using the Student’s t-test. *p < 0.05
Mentions: First, we analyzed the effect of DMF on constitutive MCP-1 expression in HUVECs. DMF inhibited MCP-1 expression significantly in a time- and concentration-dependent manner (Fig. 1a,b). The concentrations and incubation times that we adopted have been published recently and have been proven to have no cytotoxic effects in HUVEC [14]. DMF in concentrations less than 20 μM reduced the MCP-1 expression level by almost 50 %; within 3 h of treatment, the MCP-1 expression was significantly reduced compared with that of the matching controls. To determine whether DMF was also able to reduce TNF-α-induced MCP-1 expression, we analyzed the effects of the combinational treatment with DMF and TNF-α (20 ng/ml) in a time- and concentration-dependent manner. We found that 80 μM DMF reduced the TNF-α-induced MCP-1 expression by more than 50 % (Fig. 1c). A first significant reduction of MCP-1 expression could be observed as early as 6 h after treatment with 80 μM DMF and TNF-α (Fig. 1d).Fig. 1

Bottom Line: Furthermore, we found that DMF slightly inhibited the early degradation of IκBα.This action is regulated by reduced p65 activity and nuclear translocation, which can be explained in part by the reduced early degradation of IκBα and more important the reduced phosphorylation of p65 at Serine 536.These effects were independent of the p38, PI3K and p42/44 signaling pathways.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, Venereology and Allergology, Goethe-University, Theodor-Stern-Kai 7, D-60590 Frankfurt am Main, Germany ; Department of Cardiology, Gutenberg-University, Mainz, Germany.

ABSTRACT

Background: Inflammation, angiogenesis and oxidative stress have been implicated in the pathogenesis of various vascular diseases. Recent evidence suggests that dimethylfumarate (DMF), an antiposriatic and anti-multiple sclerosis agent, possesses anti-inflammatory, anti-oxidative and anti-angiogenic properties. Here, we analyze the influence of DMF on TNF-α-induced expression of the important pro-inflammatory and pro-atherogenic chemokine MCP-1 and investigate the underlying mechanisms of this expression.

Findings: We analyzed constitutive and TNF-α-induced expression of MCP-1 in human umbilical vascular endothelial cells (HUVEC) +/- DMF treatment via enzyme-linkes immunosorbent assay (ELISA). DMF significantly inhibited the protein expression levels in a time- and concentration-dependent manner. Furthermore, MCP-1 mRNA expression was also reduced in response to DMF, as demonstrated by RT-PCR. Thus, the regulation occurs at the transcriptional level. Interestingly, DMF prolonged the TNF-α-induced p38 and JNK phosphorylation in HUVEC, as demonstrated by Western blot analysis; however, the p38 and JNK inhibitor SB203580 did not affect the DMF-conveyed suppression of TNF-α-induced MCP-1 expression. DMF suppressed the TNF-α-induced nuclear translocation and phosphorylation (Serine 536) of p65 in these cells. These results were additionally approved by p65 luciferase promoter assays. Furthermore, we found that DMF slightly inhibited the early degradation of IκBα. In addition, we verified our results using other important inflammatory cytokines such as CCL-5, PDGF-BB, GM-CSF and IL-6.

Conclusion: DMF suppresses various TNF-α-induced pro-inflammatory and pro-atherogenic cytokines/chemokines in human endothelial cells. This action is regulated by reduced p65 activity and nuclear translocation, which can be explained in part by the reduced early degradation of IκBα and more important the reduced phosphorylation of p65 at Serine 536. These effects were independent of the p38, PI3K and p42/44 signaling pathways. As a result, DMF might be suitable for treating patients with vascular diseases.

No MeSH data available.


Related in: MedlinePlus