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Berteroin present in cruciferous vegetables exerts potent anti-inflammatory properties in murine macrophages and mouse skin.

Jung YJ, Jung JI, Cho HJ, Choi MS, Sung MK, Yu R, Kang YH, Park JH - Int J Mol Sci (2014)

Bottom Line: Berteroin decreased LPS-induced release of inflammatory mediators and pro-inflammatory cytokines in Raw 264.7 macrophages.Furthermore, berteroin suppressed degradation of IL-1 receptor-associated kinase and phosphorylation of transforming growth factor β activated kinase-1.Berteroin also inhibited LPS-induced phosphorylation of p38 MAPK, ERK1/2, and AKT.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science and Nutrition, Hallym University, Chuncheon 200-702, Korea. betyloving@naver.com.

ABSTRACT
Berteroin (5-methylthiopentyl isothiocyanate) is a sulforaphane analog present in cruciferous vegetables, including Chinese cabbage, rucola salad leaves, and mustard oil. We examined whether berteroin exerts anti-inflammatory activities using lipopolysaccharide (LPS)-stimulated Raw 264.7 macrophages and 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced mouse skin inflammation models. Berteroin decreased LPS-induced release of inflammatory mediators and pro-inflammatory cytokines in Raw 264.7 macrophages. Berteroin inhibited LPS-induced degradation of inhibitor of κBα (IκBα) and nuclear factor-κB p65 translocation to the nucleus and DNA binding activity. Furthermore, berteroin suppressed degradation of IL-1 receptor-associated kinase and phosphorylation of transforming growth factor β activated kinase-1. Berteroin also inhibited LPS-induced phosphorylation of p38 MAPK, ERK1/2, and AKT. In the mouse ear, berteroin effectively suppressed TPA-induced edema formation and down-regulated iNOS and COX-2 expression as well as phosphorylation of AKT and ERK1/2. These results demonstrate that berteroin exhibits potent anti-inflammatory properties and suggest that berteroin can be developed as a skin anti-inflammatory agent.

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Berteroin inhibits nuclear factor (NF)-κB translocation to the nucleus and its DNA binding activity in lipopolysaccharide (LPS)-stimulated Raw 264.7 cells. The cells were treated with various concentrations of berteroin for 40 min. LPS was added, and the incubation was continued an additional 20 min. (A) Cytosolic and nuclear fractions were prepared 20 min after the incubation with LPS, and the levels of p65 were analyzed by Western blotting. Photographs of chemiluminescent detection of the blots, which were representative of three independent experiments, are shown (upper panel). Relative density (mean ± SEM, n = 3) was calculated using Image J software (lower panel). Levels of α-tubulin and lamin B were used as cytosolic and nuclear loading controls, respectively; (B) The effect of berteroin on the DNA binding activity of p65 was evaluated by electrophoretic mobility shift assay. The nuclear extracts were incubated with the [γ-32P]-labeled NF-κB consensus oligonucleotides for 30 min. Each sample was subjected to 5% nondenaturing gel electrophoresis. (Left) An autoradiograph of the dried gel, which was representative of three independent experiments, is shown. (Right) The relative abundance of each band was quantified, and the LPS control levels (1 mg/L LPS + 0 μmol/L berteroin) were set to 100%. Each bar represents the mean ± SEM (n = 3). * Significantly different from the control group (p < 0.05). Means with different letter (a, b, c or d) are significantly different among the berteroin groups (p < 0.05).
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ijms-15-20686-f004: Berteroin inhibits nuclear factor (NF)-κB translocation to the nucleus and its DNA binding activity in lipopolysaccharide (LPS)-stimulated Raw 264.7 cells. The cells were treated with various concentrations of berteroin for 40 min. LPS was added, and the incubation was continued an additional 20 min. (A) Cytosolic and nuclear fractions were prepared 20 min after the incubation with LPS, and the levels of p65 were analyzed by Western blotting. Photographs of chemiluminescent detection of the blots, which were representative of three independent experiments, are shown (upper panel). Relative density (mean ± SEM, n = 3) was calculated using Image J software (lower panel). Levels of α-tubulin and lamin B were used as cytosolic and nuclear loading controls, respectively; (B) The effect of berteroin on the DNA binding activity of p65 was evaluated by electrophoretic mobility shift assay. The nuclear extracts were incubated with the [γ-32P]-labeled NF-κB consensus oligonucleotides for 30 min. Each sample was subjected to 5% nondenaturing gel electrophoresis. (Left) An autoradiograph of the dried gel, which was representative of three independent experiments, is shown. (Right) The relative abundance of each band was quantified, and the LPS control levels (1 mg/L LPS + 0 μmol/L berteroin) were set to 100%. Each bar represents the mean ± SEM (n = 3). * Significantly different from the control group (p < 0.05). Means with different letter (a, b, c or d) are significantly different among the berteroin groups (p < 0.05).

Mentions: NF-κB is a key transcriptional regulator of pro-inflammatory gene expression. It induces transcription of a wide variety of genes including COX-2, iNOS, IL-1, and TNF genes [25] (reviewed in [26]). The Western blotting results revealed that LPS reduced the levels of cytosolic NF-κB p65 and increased those of nuclear p65, which were prevented by berteroin in a dose-dependent manner (Figure 4A). These results clearly indicate that berteroin inhibits p65 translocation to the nucleus. The electrophoretic mobility shift assay (EMSA) results revealed that LPS greatly increased NF-κB DNA binding activity, which was markedly suppressed in a berteroin dose-dependent manner (Figure 4B).


Berteroin present in cruciferous vegetables exerts potent anti-inflammatory properties in murine macrophages and mouse skin.

Jung YJ, Jung JI, Cho HJ, Choi MS, Sung MK, Yu R, Kang YH, Park JH - Int J Mol Sci (2014)

Berteroin inhibits nuclear factor (NF)-κB translocation to the nucleus and its DNA binding activity in lipopolysaccharide (LPS)-stimulated Raw 264.7 cells. The cells were treated with various concentrations of berteroin for 40 min. LPS was added, and the incubation was continued an additional 20 min. (A) Cytosolic and nuclear fractions were prepared 20 min after the incubation with LPS, and the levels of p65 were analyzed by Western blotting. Photographs of chemiluminescent detection of the blots, which were representative of three independent experiments, are shown (upper panel). Relative density (mean ± SEM, n = 3) was calculated using Image J software (lower panel). Levels of α-tubulin and lamin B were used as cytosolic and nuclear loading controls, respectively; (B) The effect of berteroin on the DNA binding activity of p65 was evaluated by electrophoretic mobility shift assay. The nuclear extracts were incubated with the [γ-32P]-labeled NF-κB consensus oligonucleotides for 30 min. Each sample was subjected to 5% nondenaturing gel electrophoresis. (Left) An autoradiograph of the dried gel, which was representative of three independent experiments, is shown. (Right) The relative abundance of each band was quantified, and the LPS control levels (1 mg/L LPS + 0 μmol/L berteroin) were set to 100%. Each bar represents the mean ± SEM (n = 3). * Significantly different from the control group (p < 0.05). Means with different letter (a, b, c or d) are significantly different among the berteroin groups (p < 0.05).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4264190&req=5

ijms-15-20686-f004: Berteroin inhibits nuclear factor (NF)-κB translocation to the nucleus and its DNA binding activity in lipopolysaccharide (LPS)-stimulated Raw 264.7 cells. The cells were treated with various concentrations of berteroin for 40 min. LPS was added, and the incubation was continued an additional 20 min. (A) Cytosolic and nuclear fractions were prepared 20 min after the incubation with LPS, and the levels of p65 were analyzed by Western blotting. Photographs of chemiluminescent detection of the blots, which were representative of three independent experiments, are shown (upper panel). Relative density (mean ± SEM, n = 3) was calculated using Image J software (lower panel). Levels of α-tubulin and lamin B were used as cytosolic and nuclear loading controls, respectively; (B) The effect of berteroin on the DNA binding activity of p65 was evaluated by electrophoretic mobility shift assay. The nuclear extracts were incubated with the [γ-32P]-labeled NF-κB consensus oligonucleotides for 30 min. Each sample was subjected to 5% nondenaturing gel electrophoresis. (Left) An autoradiograph of the dried gel, which was representative of three independent experiments, is shown. (Right) The relative abundance of each band was quantified, and the LPS control levels (1 mg/L LPS + 0 μmol/L berteroin) were set to 100%. Each bar represents the mean ± SEM (n = 3). * Significantly different from the control group (p < 0.05). Means with different letter (a, b, c or d) are significantly different among the berteroin groups (p < 0.05).
Mentions: NF-κB is a key transcriptional regulator of pro-inflammatory gene expression. It induces transcription of a wide variety of genes including COX-2, iNOS, IL-1, and TNF genes [25] (reviewed in [26]). The Western blotting results revealed that LPS reduced the levels of cytosolic NF-κB p65 and increased those of nuclear p65, which were prevented by berteroin in a dose-dependent manner (Figure 4A). These results clearly indicate that berteroin inhibits p65 translocation to the nucleus. The electrophoretic mobility shift assay (EMSA) results revealed that LPS greatly increased NF-κB DNA binding activity, which was markedly suppressed in a berteroin dose-dependent manner (Figure 4B).

Bottom Line: Berteroin decreased LPS-induced release of inflammatory mediators and pro-inflammatory cytokines in Raw 264.7 macrophages.Furthermore, berteroin suppressed degradation of IL-1 receptor-associated kinase and phosphorylation of transforming growth factor β activated kinase-1.Berteroin also inhibited LPS-induced phosphorylation of p38 MAPK, ERK1/2, and AKT.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science and Nutrition, Hallym University, Chuncheon 200-702, Korea. betyloving@naver.com.

ABSTRACT
Berteroin (5-methylthiopentyl isothiocyanate) is a sulforaphane analog present in cruciferous vegetables, including Chinese cabbage, rucola salad leaves, and mustard oil. We examined whether berteroin exerts anti-inflammatory activities using lipopolysaccharide (LPS)-stimulated Raw 264.7 macrophages and 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced mouse skin inflammation models. Berteroin decreased LPS-induced release of inflammatory mediators and pro-inflammatory cytokines in Raw 264.7 macrophages. Berteroin inhibited LPS-induced degradation of inhibitor of κBα (IκBα) and nuclear factor-κB p65 translocation to the nucleus and DNA binding activity. Furthermore, berteroin suppressed degradation of IL-1 receptor-associated kinase and phosphorylation of transforming growth factor β activated kinase-1. Berteroin also inhibited LPS-induced phosphorylation of p38 MAPK, ERK1/2, and AKT. In the mouse ear, berteroin effectively suppressed TPA-induced edema formation and down-regulated iNOS and COX-2 expression as well as phosphorylation of AKT and ERK1/2. These results demonstrate that berteroin exhibits potent anti-inflammatory properties and suggest that berteroin can be developed as a skin anti-inflammatory agent.

Show MeSH
Related in: MedlinePlus