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A novel chromone derivative with anti-inflammatory property via inhibition of ROS-dependent activation of TRAF6-ASK1-p38 pathway.

Liu H, Xu R, Feng L, Guo W, Cao N, Qian C, Teng P, Wang L, Wu X, Sun Y, Li J, Shen Y, Xu Q - PLoS ONE (2012)

Bottom Line: In the present study, we show that a novel chromone derivative, DCO-6, significantly reduced lipopolysaccharide (LPS)-induced production of nitric oxide, IL-1β and IL-6, decreased the levels of iNOS, IL-1β and IL-6 mRNA expression in both RAW264.7 cells and mouse primary peritoneal macrophages, and inhibited LPS-induced activation of p38 MAPK but not of JNK, ERK.LPS-induced production of intracellular reactive oxygen species (ROS) was remarkably impaired by DCO-6, which disrupted the formation of the TRAF6-ASK1 complex.Our results indicate that DCO-6 showed anti-inflammatory properties through inhibition of ROS-dependent activation of TRAF6-ASK1-p38 pathway.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China.

ABSTRACT
The p38 MAPK signaling pathway plays a pivotal role in inflammation. Targeting p38 MAPK may be a potential strategy for the treatment of inflammatory diseases. In the present study, we show that a novel chromone derivative, DCO-6, significantly reduced lipopolysaccharide (LPS)-induced production of nitric oxide, IL-1β and IL-6, decreased the levels of iNOS, IL-1β and IL-6 mRNA expression in both RAW264.7 cells and mouse primary peritoneal macrophages, and inhibited LPS-induced activation of p38 MAPK but not of JNK, ERK. Moreover, DCO-6 specifically inhibited TLR4-dependent p38 activation without directly inhibiting its kinase activity. LPS-induced production of intracellular reactive oxygen species (ROS) was remarkably impaired by DCO-6, which disrupted the formation of the TRAF6-ASK1 complex. Administering DCO-6 significantly protected mice from LPS-induced septic shock in parallel with the inhibition of p38 activation and ROS production. Our results indicate that DCO-6 showed anti-inflammatory properties through inhibition of ROS-dependent activation of TRAF6-ASK1-p38 pathway. Blockade of the upstream events required for p38 MAPK action by DCO-6 may provide a new therapeutic option in the treatment of inflammatory diseases.

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Effects of DCO-6 on p38 MAPK activation induced by different TLR ligands in RAW264.7 cells.(A) Cells were treated with DCO-6 in the absence or presence of indicated TLR ligands for 6 h. Whole cell lysates were prepared for Western blotting analysis. The protein levels of total and phosphorylated p38 were determined at least three times, and the representative data are shown. (B) Cells were stimulated by 500 ng/ml LPS for 6 h and the protein was collected. Phosphorylated p38 MAPK was immunoprecipitated. The immune complexes were used for testing the effects of DCO-6 and SB203580 on kinase activities. Representative data are shown. Bands from (A–B) were analyzed by densitometry. Quantitative data are shown. *P<0.05 vs control.
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pone-0037168-g004: Effects of DCO-6 on p38 MAPK activation induced by different TLR ligands in RAW264.7 cells.(A) Cells were treated with DCO-6 in the absence or presence of indicated TLR ligands for 6 h. Whole cell lysates were prepared for Western blotting analysis. The protein levels of total and phosphorylated p38 were determined at least three times, and the representative data are shown. (B) Cells were stimulated by 500 ng/ml LPS for 6 h and the protein was collected. Phosphorylated p38 MAPK was immunoprecipitated. The immune complexes were used for testing the effects of DCO-6 and SB203580 on kinase activities. Representative data are shown. Bands from (A–B) were analyzed by densitometry. Quantitative data are shown. *P<0.05 vs control.

Mentions: Next we used LPS (a ligand for TLR4), poly (I:C) (a synthetic double-stranded RNA and a ligand for TLR3) or unmethylated CpG (a ligand for TLR9) to stimulate RAW264.7 cells. As shown in Fig. 4A, p38 MAPK phosphorylation was substantially induced upon any stimuli. However, DCO-6 failed to reduce p38 phosphorylation induced by poly (I:C) or CpG, suggesting that DCO-6 specifically inhibited TLR4-dependent p38 activation. In an in vitro kinase assay to determine the direct effect of DCO-6 on the p38 MAPK, we found that, unlike p38 MAPK inhibitor SB203580, which can inhibit p38 MAPK at 10 µM, DCO-6 did not affect the kinase activity of p38 MAPK (Fig. 4B and Fig. S4).


A novel chromone derivative with anti-inflammatory property via inhibition of ROS-dependent activation of TRAF6-ASK1-p38 pathway.

Liu H, Xu R, Feng L, Guo W, Cao N, Qian C, Teng P, Wang L, Wu X, Sun Y, Li J, Shen Y, Xu Q - PLoS ONE (2012)

Effects of DCO-6 on p38 MAPK activation induced by different TLR ligands in RAW264.7 cells.(A) Cells were treated with DCO-6 in the absence or presence of indicated TLR ligands for 6 h. Whole cell lysates were prepared for Western blotting analysis. The protein levels of total and phosphorylated p38 were determined at least three times, and the representative data are shown. (B) Cells were stimulated by 500 ng/ml LPS for 6 h and the protein was collected. Phosphorylated p38 MAPK was immunoprecipitated. The immune complexes were used for testing the effects of DCO-6 and SB203580 on kinase activities. Representative data are shown. Bands from (A–B) were analyzed by densitometry. Quantitative data are shown. *P<0.05 vs control.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3376149&req=5

pone-0037168-g004: Effects of DCO-6 on p38 MAPK activation induced by different TLR ligands in RAW264.7 cells.(A) Cells were treated with DCO-6 in the absence or presence of indicated TLR ligands for 6 h. Whole cell lysates were prepared for Western blotting analysis. The protein levels of total and phosphorylated p38 were determined at least three times, and the representative data are shown. (B) Cells were stimulated by 500 ng/ml LPS for 6 h and the protein was collected. Phosphorylated p38 MAPK was immunoprecipitated. The immune complexes were used for testing the effects of DCO-6 and SB203580 on kinase activities. Representative data are shown. Bands from (A–B) were analyzed by densitometry. Quantitative data are shown. *P<0.05 vs control.
Mentions: Next we used LPS (a ligand for TLR4), poly (I:C) (a synthetic double-stranded RNA and a ligand for TLR3) or unmethylated CpG (a ligand for TLR9) to stimulate RAW264.7 cells. As shown in Fig. 4A, p38 MAPK phosphorylation was substantially induced upon any stimuli. However, DCO-6 failed to reduce p38 phosphorylation induced by poly (I:C) or CpG, suggesting that DCO-6 specifically inhibited TLR4-dependent p38 activation. In an in vitro kinase assay to determine the direct effect of DCO-6 on the p38 MAPK, we found that, unlike p38 MAPK inhibitor SB203580, which can inhibit p38 MAPK at 10 µM, DCO-6 did not affect the kinase activity of p38 MAPK (Fig. 4B and Fig. S4).

Bottom Line: In the present study, we show that a novel chromone derivative, DCO-6, significantly reduced lipopolysaccharide (LPS)-induced production of nitric oxide, IL-1β and IL-6, decreased the levels of iNOS, IL-1β and IL-6 mRNA expression in both RAW264.7 cells and mouse primary peritoneal macrophages, and inhibited LPS-induced activation of p38 MAPK but not of JNK, ERK.LPS-induced production of intracellular reactive oxygen species (ROS) was remarkably impaired by DCO-6, which disrupted the formation of the TRAF6-ASK1 complex.Our results indicate that DCO-6 showed anti-inflammatory properties through inhibition of ROS-dependent activation of TRAF6-ASK1-p38 pathway.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, China.

ABSTRACT
The p38 MAPK signaling pathway plays a pivotal role in inflammation. Targeting p38 MAPK may be a potential strategy for the treatment of inflammatory diseases. In the present study, we show that a novel chromone derivative, DCO-6, significantly reduced lipopolysaccharide (LPS)-induced production of nitric oxide, IL-1β and IL-6, decreased the levels of iNOS, IL-1β and IL-6 mRNA expression in both RAW264.7 cells and mouse primary peritoneal macrophages, and inhibited LPS-induced activation of p38 MAPK but not of JNK, ERK. Moreover, DCO-6 specifically inhibited TLR4-dependent p38 activation without directly inhibiting its kinase activity. LPS-induced production of intracellular reactive oxygen species (ROS) was remarkably impaired by DCO-6, which disrupted the formation of the TRAF6-ASK1 complex. Administering DCO-6 significantly protected mice from LPS-induced septic shock in parallel with the inhibition of p38 activation and ROS production. Our results indicate that DCO-6 showed anti-inflammatory properties through inhibition of ROS-dependent activation of TRAF6-ASK1-p38 pathway. Blockade of the upstream events required for p38 MAPK action by DCO-6 may provide a new therapeutic option in the treatment of inflammatory diseases.

Show MeSH
Related in: MedlinePlus