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Apigenin Inhibits Tumor Necrosis Factor-α-Induced Production and Gene Expression of Mucin through Regulating Nuclear Factor-Kappa B Signaling Pathway in Airway Epithelial Cells.

Seo HS, Sikder MA, Lee HJ, Ryu J, Lee CJ - Biomol Ther (Seoul) (2014)

Bottom Line: Apigenin significantly inhibited MUC5AC mucin production and down-regulated MUC5AC gene expression induced by TNF-α in NCI-H292 cells.To elucidate the action mechanism of apigenin, effect of apigenin on TNF-α-induced nuclear factor kappa B (NF-κB) signaling pathway was also investigated by western blot analysis.Apigenin also has an influence on upstream signaling of IKK because it inhibited the expression of adaptor protein, receptor interacting protein 1 (RIP1).

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Medicine, Chungnam National University, Daejeon 301-131, Republic of Korea.

ABSTRACT
In the present study, we investigated whether apigenin significantly affects tumor necrosis factor-α (TNF-α)-induced production and gene expression of MUC5AC mucin in airway epithelial cells. Confluent NCI-H292 cells were pretreated with apigenin for 30 min and then stimulated with TNF-α for 24 h or the indicated periods. The MUC5AC mucin gene expression and mucin protein production were measured by reverse transcription - polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Apigenin significantly inhibited MUC5AC mucin production and down-regulated MUC5AC gene expression induced by TNF-α in NCI-H292 cells. To elucidate the action mechanism of apigenin, effect of apigenin on TNF-α-induced nuclear factor kappa B (NF-κB) signaling pathway was also investigated by western blot analysis. Apigenin inhibited NF-κB activation induced by TNF-α. Inhibition of inhibitory kappa B kinase (IKK) by apigenin led to the suppression of inhibitory kappa B alpha (IκBα) phosphorylation and degradation, p65 nuclear translocation. This, in turn, led to the down-regulation of MUC5AC protein production in NCI-H292 cells. Apigenin also has an influence on upstream signaling of IKK because it inhibited the expression of adaptor protein, receptor interacting protein 1 (RIP1). These results suggest that apigenin can regulate the production and gene expression of mucin through regulating NF-κB signaling pathway in airway epithelial cells.

No MeSH data available.


Related in: MedlinePlus

Effect of apigenin on TNF-α-induced nuclear translocation of NF-κB p65 and phosphorylation of p65. NCI-H292 cells were either untreated or pretreated with 20 μM apigenin for 24 h at 37°C and then stimulated with TNF-α (50 ng/mL) for the indicated periods. Nuclear protein extracts were prepared and resolved on 10% SDS-PAGE, transferred onto a PVDF membrane, probed with antibody against p65 (A) or phospho-specific p65 (Ser 536) antibody (B). The results shown are the representative of three independent experiments. To ensure equal protein loading, the membrane was reprobed with anti-p84 antibody. *significantly different from TNF-α alone at each time point (p<0.05, Student’s t-test) (cont: control, Api: apigenin, TNF: TNF-α).
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f2-bt-22-525: Effect of apigenin on TNF-α-induced nuclear translocation of NF-κB p65 and phosphorylation of p65. NCI-H292 cells were either untreated or pretreated with 20 μM apigenin for 24 h at 37°C and then stimulated with TNF-α (50 ng/mL) for the indicated periods. Nuclear protein extracts were prepared and resolved on 10% SDS-PAGE, transferred onto a PVDF membrane, probed with antibody against p65 (A) or phospho-specific p65 (Ser 536) antibody (B). The results shown are the representative of three independent experiments. To ensure equal protein loading, the membrane was reprobed with anti-p84 antibody. *significantly different from TNF-α alone at each time point (p<0.05, Student’s t-test) (cont: control, Api: apigenin, TNF: TNF-α).

Mentions: To determine the effect of apigenin on NF-κB activation followed by TNF-α treatment, we examined whether apigenin inhibits TNF-α-induced nuclear translocation of NF-κB p65. As shown in Fig. 2A, nuclear translocation of NF-κB p65 by TNF-α was inhibited by pretreatment with 20 μM of apigenin. In the nuclear fraction of the cells treated with TNF-α only, there was an increase in nuclear translocation of p65 gradually and reached optimal level at 30 min. However, in the cells treated with apigenin plus TNF-α, the level of p65 was gradually decreased as compared to the cells treated with TNF-α only. Next, transcriptional activity of p65 largely depends on its phosphorylation. For this reason, we investigated the effect of apigenin on TNF-α-induced phosphorylation of p65. As shown in Fig. 2B, TNF-α-induced phosphorylation of p65 was gradually increased and reached optimal level at 30 min. However, apigenin completely blocked the phosphorylation of p65.


Apigenin Inhibits Tumor Necrosis Factor-α-Induced Production and Gene Expression of Mucin through Regulating Nuclear Factor-Kappa B Signaling Pathway in Airway Epithelial Cells.

Seo HS, Sikder MA, Lee HJ, Ryu J, Lee CJ - Biomol Ther (Seoul) (2014)

Effect of apigenin on TNF-α-induced nuclear translocation of NF-κB p65 and phosphorylation of p65. NCI-H292 cells were either untreated or pretreated with 20 μM apigenin for 24 h at 37°C and then stimulated with TNF-α (50 ng/mL) for the indicated periods. Nuclear protein extracts were prepared and resolved on 10% SDS-PAGE, transferred onto a PVDF membrane, probed with antibody against p65 (A) or phospho-specific p65 (Ser 536) antibody (B). The results shown are the representative of three independent experiments. To ensure equal protein loading, the membrane was reprobed with anti-p84 antibody. *significantly different from TNF-α alone at each time point (p<0.05, Student’s t-test) (cont: control, Api: apigenin, TNF: TNF-α).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4256032&req=5

f2-bt-22-525: Effect of apigenin on TNF-α-induced nuclear translocation of NF-κB p65 and phosphorylation of p65. NCI-H292 cells were either untreated or pretreated with 20 μM apigenin for 24 h at 37°C and then stimulated with TNF-α (50 ng/mL) for the indicated periods. Nuclear protein extracts were prepared and resolved on 10% SDS-PAGE, transferred onto a PVDF membrane, probed with antibody against p65 (A) or phospho-specific p65 (Ser 536) antibody (B). The results shown are the representative of three independent experiments. To ensure equal protein loading, the membrane was reprobed with anti-p84 antibody. *significantly different from TNF-α alone at each time point (p<0.05, Student’s t-test) (cont: control, Api: apigenin, TNF: TNF-α).
Mentions: To determine the effect of apigenin on NF-κB activation followed by TNF-α treatment, we examined whether apigenin inhibits TNF-α-induced nuclear translocation of NF-κB p65. As shown in Fig. 2A, nuclear translocation of NF-κB p65 by TNF-α was inhibited by pretreatment with 20 μM of apigenin. In the nuclear fraction of the cells treated with TNF-α only, there was an increase in nuclear translocation of p65 gradually and reached optimal level at 30 min. However, in the cells treated with apigenin plus TNF-α, the level of p65 was gradually decreased as compared to the cells treated with TNF-α only. Next, transcriptional activity of p65 largely depends on its phosphorylation. For this reason, we investigated the effect of apigenin on TNF-α-induced phosphorylation of p65. As shown in Fig. 2B, TNF-α-induced phosphorylation of p65 was gradually increased and reached optimal level at 30 min. However, apigenin completely blocked the phosphorylation of p65.

Bottom Line: Apigenin significantly inhibited MUC5AC mucin production and down-regulated MUC5AC gene expression induced by TNF-α in NCI-H292 cells.To elucidate the action mechanism of apigenin, effect of apigenin on TNF-α-induced nuclear factor kappa B (NF-κB) signaling pathway was also investigated by western blot analysis.Apigenin also has an influence on upstream signaling of IKK because it inhibited the expression of adaptor protein, receptor interacting protein 1 (RIP1).

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Medicine, Chungnam National University, Daejeon 301-131, Republic of Korea.

ABSTRACT
In the present study, we investigated whether apigenin significantly affects tumor necrosis factor-α (TNF-α)-induced production and gene expression of MUC5AC mucin in airway epithelial cells. Confluent NCI-H292 cells were pretreated with apigenin for 30 min and then stimulated with TNF-α for 24 h or the indicated periods. The MUC5AC mucin gene expression and mucin protein production were measured by reverse transcription - polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Apigenin significantly inhibited MUC5AC mucin production and down-regulated MUC5AC gene expression induced by TNF-α in NCI-H292 cells. To elucidate the action mechanism of apigenin, effect of apigenin on TNF-α-induced nuclear factor kappa B (NF-κB) signaling pathway was also investigated by western blot analysis. Apigenin inhibited NF-κB activation induced by TNF-α. Inhibition of inhibitory kappa B kinase (IKK) by apigenin led to the suppression of inhibitory kappa B alpha (IκBα) phosphorylation and degradation, p65 nuclear translocation. This, in turn, led to the down-regulation of MUC5AC protein production in NCI-H292 cells. Apigenin also has an influence on upstream signaling of IKK because it inhibited the expression of adaptor protein, receptor interacting protein 1 (RIP1). These results suggest that apigenin can regulate the production and gene expression of mucin through regulating NF-κB signaling pathway in airway epithelial cells.

No MeSH data available.


Related in: MedlinePlus