• Tuberculosis and Respiratory Diseases
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• 제55권1호
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• pp.21-30
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• 2003

서 론 : MUC genes의 증가와 배상세포의 증식 기전에 성장인자(growth factor)인 상피세포 성장인자 및 수용체(epidermal growth factor receptor; EGFR)가 배상세포의 증식이나 이형성에 관여한다. EGFR의 ligands 중의 한 종류인 heparin binding EGF(HB-EGF)는 세포막에 존재하는 pro-heparin binding EGF(pro-HB-EGF)로부터 유리된다. HB-EGF의 유리는 G-protein과 연관이 있다. 따라서, 본 연구는 그람 음성세균의 lipopolysaccande(LPS)에 의한 기도 점액 과생성의 기전을 밝히고, 기도점액 과분비에서 EGFR과 G-protein의 연관성을 밝혀 기도 점액 과분비 기전을 밝히고자 한다. 연구방법 : NCI-H292 세포배양에서 LPS단독 투여 또는TGF-${\alpha}$와 병합 투여한 후 MUC5AC의 당단백질을 ELISA법으로 측정하였다. LPS에 의한 MUC5AC 당단백질의 생성 기전을 밝히기 위해서 heterotrimeric G-protein 억제제인 mastoparan을 투여하고 TNF-${\alpha}$와 MUC5AC를 ELISA법으로 각각 측정하였다. MUC5AC의 생성에서 G-protein과 EGFR의 연관성을 확인하기 위하여 EGFR이 항상 발현되어 있고 MUC5AC를 분비할 수 있는 NCI-H292 세포에 G-protein 자극제인 mastoparan-7로 자극한 후 MUC5AC의 생성을 측정하였다. G-protein이 활성화하여 metalloproteinase가 세포막에 있는 HB-EGF를 유리하여 EGFR이 활성화하여 MUC5AC가 생성여부를 확인하기 위하여 ADAM10으로 NCI-H292세포에 자극하여 MUC5AC의 생성을 측정하였다. MUC5AC 생성이 EGFR과 연관성을 확인하기 위하여 특이 EGFR tyrosine kinase 억제제인 AG1478과 중화 polyclonal EGF 항체를 전처치 후 MUC5AC를 측정하였다. 결 과 : LPS의 자극에 의한 MUC5AC의 생성은 LPS 농도에 유의하게 증가 되지 않았으나, EGFR의 ligand인 TGF-${\alpha}$를 동시 투여한 경우는 LPS의 농도에 비례하여 유의하게 증가하였다. LPS의 자극은 TNF-${\alpha}$의 생성을 유의하게 증가시켰으며, G-protein 억제제인 mastoparan을 전처치한 경우는 TNF-${\alpha}$가 유의하게 감소 되었다. LPS 자극 전에 TNF-${\alpha}$ antibody, AG1478 또는 mastoparan을 전처치한 경우는 MUC5AC의 생성이 유의하게 억제되었다. MUC5AC의 생생에서 G-protein과 EGFR의 연관성에 대한 실험에서 MUC5AC의 생성이 mastoparan-7의 농도에 따라 유의하게 증가되었으며, EGF의 중화항체를 사용한 경우는 MUC5AC의 생성이 감소되었다. 또한 Matrix metalloproteinase인 ADAM10의 농도에 비례하여 MUC5AC의 생성을 증가시켰다. 결 론 : LPS에 의한 MUC5AC의 분비는 LPS가 TNF-${\alpha}$를 생성시키고, TNF-${\alpha}$가 EGFR의 발현을 유도하여 MUC5AC가 분비되었다. 또한 MUC5AC의 생성에 있어서 G-protein의 활성은 matrix metalloproteinase에 의하여 EGFR의 ligand 인 HB-EGF가 유리되어 EGFR의 transacti vation으로 MUC5AC가 생성되는 것으로 사료된다.

• Natural Product Sciences
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• 제23권3호
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• pp.201-207
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• 2017

Angelica decursiva has been utilised as remedy for controlling the airway inflammatory diseases in folk medicine. We investigated whether nodakenin, columbianadin, and umbelliferone isolated from the roots of Angelica decursiva inhibit the gene expression and production of MUC5AC mucin from human airway epithelial cells. Confluent NCI-H292 cells were pretreated with nodakenin, columbianadin or umbelliferone for 30 min and then stimulated with epidermal growth factor (EGF), phorbol 12-myristate 13-acetate (PMA) or tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$) for 24 h. The MUC5AC mucin gene expression was measured by reverse transcription - polymerase chain reaction (RT-PCR). Production of MUC5AC mucin protein was measured by enzyme-linked immunosorbent assay (ELISA). The results were as follows: (1) Nodakenin did not affect the expression of MUC5AC mucin gene induced by EGF, PMA or $TNF-{\alpha}$. Columbianadin inhibited the expression of MUC5AC mucin gene induced by EGF or PMA. However, umbelliferone inhibited the expression of MUC5AC mucin gene induced by EGF, PMA or $TNF-{\alpha}$; (2) Nodakenin also did not affect the production of MUC5AC mucin protein induced by EGF, PMA or $TNF-{\alpha}$. Columbianadin inhibited the production of MUC5AC mucin protein induced by PMA. However, umbelliferone inhibited the production of MUC5AC mucin protein induced by EGF, PMA or $TNF-{\alpha}$. These results suggest that, among the three compounds investigated, umbelliferone only inhibits the gene expression and production of MUC5AC mucin stimulated by various inducers, by directly acting on airway epithelial cells, and the results might explain the traditional use of Angelica decursiva as remedy for diverse inflammatory pulmonary diseases.

• Tuberculosis and Respiratory Diseases
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• 제77권5호
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• pp.203-208
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• 2014

Background: In this study, we investigated whether lobetyolin, lobetyol, and methyl linoleate derived from Codonopsis pilosula affect MUC5AC mucin secretion, production, and gene expression from airway epithelial cells. Methods: Confluent NCI-H292 cells were pretreated with lobetyolin, lobetyol, or methyl linoleate for 30 minutes and then stimulated with phorbol 12-myristate 13-acetate (PMA) for 24 hours. The MUC5AC mucin gene expression, and mucin protein production and secretion were measured by reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Results: Lobetyolin, lobetyol, and methyl linoleate inhibited the gene expression of MUC5AC mucin induced by PMA; lobetyolin did not affect PMA-induced MUC5AC mucin production. However, lobetyol and methyl linoleate inhibited the production of MUC5AC mucin; lobetyolin and lobetyol did not significantly affect PMA-induced MUC5AC mucin secretion from NCI-H292 cells. However, methyl linoleate decreased the MUC5AC mucin secretion. Conclusion: These results suggest that among the three compounds, methyl linoleate can regulate gene expression, production, and secretion of MUC5AC mucin by directly acting on the airway epithelial cells.

• Biomolecules & Therapeutics
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• 제18권3호
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• pp.294-299
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• 2010

In this study, we tried to investigate whether platycodin D significantly affects MUC5AC mucin production and gene expression induced by epidermal growth factor (EGF), phorbol ester (PMA) and tumor necrosis factor-$\alpha$ (TNF-$\alpha$) from human airway epithelial cells. Confluent NCI-H292 cells were pretreated with varying concentrations of platycodin D for 30 min and then stimulated with EGF, PMA and TNF-$\alpha$ for 24h, respectively. MUC5AC mucin gene expression and mucin protein production were measured by RT-PCR and ELISA. The results were as follows: (1) Platycodin D was found to inhibit the production of MUC5AC mucin protein induced by EGF, PMA, and TNF-$\alpha$, respectively. (2) It also inhibited the expression of MUC5AC mucin gene induced by the same inducers. These results suggest that platycodin D can regulate mucin gene expression and production of mucin protein, by directly acting on human airway epithelial cells.

• Natural Product Sciences
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• 제21권1호
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• pp.59-65
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• 2015

In this study, we investigated whether cynaroside, cynarin and linarin derived from Chrysanthemum indicum L. affect the secretion, production and gene expression of MUC5AC mucin in airway epithelial cells. Confluent NCI-H292 cells were pretreated with cynaroside, cynarin or linarin for 30 min and then stimulated with PMA (phorbol 12-myristate 13-acetate) for 24 h. The MUC5AC mucin gene expression, mucin protein production and secretion were measured by RT-PCR and ELISA, respectively. Effect of linarin on EGF (epidermal growth factor) - or TNF-${\alpha}$ (tumor necrosis factor-${\alpha}$)-induced MUC5AC mucin gene expression and mucin protein production was also examined. The results were as follows: (1) Cynaroside and cynarin did not significantly affect PMA-induced MUC5AC mucin secretion from NCI-H292 cells. However, linarin decreased MUC5AC mucin secretion; (2) Cynaroside did not affect PMA-induced MUC5AC mucin production and gene expresion from NCI-H292 cells. However, cynarin and linarin inhibited the production and gene expression of MUC5AC mucin; (3) Linarin also inhibited the production and gene expression of MUC5AC mucin induced by EGF- or TNF-${\alpha}$ from NCI-H292 cells. These results suggest that linarin can regulate the gene expression, production and secretion of mucin, by directly acting on airway epithelial cells.

• Tuberculosis and Respiratory Diseases
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• 제69권5호
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• pp.348-353
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• 2010

Background: In this study, we tried to investigate whether carbenoxolone, prunetin, and silibinin affect tumor necrosis factor (TNF)-${\alpha}$-induced MUC5AC mucin production and gene expression from human airway epithelial cells. Methods: Confluent NCI-H292 cells were pretreated with each agent (carbenoxolone, prunetin, and silibinin) for 30 min and then stimulated with TNF-${\alpha}$ for 24 hours. The MUC5AC mucin gene expression and mucin protein production were measured by reverse transcription-polymerase chain reaction and enzyme linked immunosorbent assay, respectively. Results: Carbenoxolone, prunetin and silibinin inhibited the production of MUC5AC mucin protein induced by TNF-${\alpha}$; the 3 compounds also inhibited the expression of MUC5AC mucin gene induced by TNF-${\alpha}$. Conclusion: This result suggests that carbenoxolone, prunetin and silibinin can inhibit mucin gene expression and production of mucin protein induced by TNF-${\alpha}$, by directly acting on airway epithelial cells.

• Journal of Microbiology and Biotechnology
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• 제25권5호
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• pp.589-597
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• 2015

It has been reported that overexpression of MUC5AC induced by excessive inflammation leads to airway obstruction in respiratory diseases such as chronic obstructive pulmonary disease and asthma. 15-Hydroxyeicosatetraenoic acid (15-HETE) has been reported to have anti-inflammatory effects, but the role of 15-HETE in respiratory inflammation has not been determined. Therefore, the aim of this study was to investigate the effects of 15-HETE on MUC5AC expression and related pathways. In this study, phorbol-12-myristate-13-acetate (PMA) was used to stimulate NCI-H292 bronchial epithelial cells in order to examine the effects of 15-HETE. 15-HETE inhibited PMA-induced expression of MUC5AC mRNA and secretion of MUC5AC protein. Moreover, 15-HETE regulated matrix metallopeptidase 9 (MMP-9), mitogen-activated protein kinase kinase (MEK), and extracellular signal-regulated kinase (ERK). In addition, 15-HETE decreased the nuclear translocation of specificity protein-1 (Sp-1) transcription factor and nuclear factor κB (NF-κB). Furthermore, 15-HETE enhanced the transcriptional activity of peroxisome proliferator-activated receptor gamma (PPARγ) as a PPARγ agonist. This activity reduced the phosphorylation of protein kinase B (PΚB/Akt) by increasing the expression of phosphatase and tensin homolog (PTEN). In conclusion, 15-HETE regulated MUC5AC expression via modulating MMP-9, MEK/ERK/Sp-1, and PPARγ/PTEN/Akt signaling pathways in PMA-treated respiratory epithelial cells.

• Tuberculosis and Respiratory Diseases
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• 제80권1호
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• pp.60-68
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• 2017

Background: Mucus hypersecretion from airway epithelium is a characteristic feature of airway inflammatory diseases. Tumor necrosis factor ${\alpha}$ (TNF-${\alpha}$) regulates mucin synthesis. Glucocorticoids including mometasone fuorate (MF) have been used to attenuate airway inflammation. However, effects of MF on mucin production have not been reported. Methods: Effects of MF and budesonide (BUD) on the phorbol-12-myristate-13-acetate (PMA)-induction of mucin and TNF-${\alpha}$ in human airway epithelial cells (NCI-H292) were investigated in the present study. Confluent NCI-H292 cells were pretreated with PMA (200 nM) for 2 hours. Subsequently, the cells were stimulated with MF (1-500 ng/mL) or BUD (21.5 ng/mL) for 8 hours. Dexamethasone ($1{\mu}g/mL$) was used as the positive control. Real-time polymerase chain reaction was used to determine MUC2 and MUC5AC mRNA levels. The level of total mucin, MUC2, MUC5AC, and TNF-${\alpha}$ in culture supernatants were measured using enzyme-linked immunosorbent assay. Results: MF and BUD significantly suppressed MUC2 and MUC5AC gene expression in PMA-stimulated NCI-H292 cells. The inhibitory effects of the two steroid drugs were also observed in the production of total mucin, MUC2 and MUC5AC proteins, and TNF-${\alpha}$. Conclusion: Our findings demonstrated that MF and BUD attenuated mucin and TNF-${\alpha}$ production in PMA-induced human airway epithelial cells.

• The Korean Journal of Physiology and Pharmacology
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• 제22권6호
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• pp.671-677
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• 2018

In the present study, we investigated whether tussilagone, a natural product derived from Tussilago farfara, significantly affects the production and gene expression of airway MUC5AC mucin. Confluent NCI-H292 cells were pretreated with tussilagone for 30 min and then stimulated with EGF (epidermal growth factor) or PMA (phorbol 12-myristate 13-acetate) for 24 h or the indicated periods. The MUC5AC mucin gene expression was measured by RT-PCR. Production of MUC5AC mucin protein was measured by ELISA. To elucidate the action mechanism of tussilagone, effect of tussilagone on PMA-induced $NF-{\kappa}B$ signaling pathway was investigated by western blot analysis. Tussilagone significantly inhibited the production of MUC5AC mucin protein and down-regulated the expression of MUC5AC mucin gene, induced by EGF or PMA. Tussilagone inhibited PMA-induced activation (phosphorylation) of inhibitory kappa B kinase (IKK), and thus phosphorylation and degradation of inhibitory kappa Ba ($I{\kappa}B{\alpha}$). Tussilagone inhibited PMA-induced phosphorylation and nuclear translocation of nuclear factor kappa B ($NF-{\kappa}B$) p65. This, in turn, led to the down-regulation of MUC5AC protein production in NCI-H292 cells. These results suggest that tussilagone can regulate the production and gene expression of mucin by acting on airway epithelial cells through regulation of $NF-{\kappa}B$ signaling pathway.

• Biomolecules & Therapeutics
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• 제18권4호
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• pp.396-401
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• 2010

We conducted this study to investigate whether baicalin, baicalein or schizandrin significantly affect MUC5AC mucin production induced by epidermal growth factor (EGF) or phorbol ester (PMA) in human airway epithelial cells. Confluent NCI-H292 cells were pretreated with varying concentrations of baicalin, baicalein or schizandrin for 30 min and then stimulated with EGF or PMA for 24 h, respectively. MUC5AC mucin protein production was measured by ELISA. The results were as follows: (1) Baicalin was found to inhibit the production of MUC5AC mucin protein induced by both EGF and PMA. (2) Baicalein, the aglycone of baicalin, also inhibited MUC5AC mucin production. (3) Schizandrin, derived from Schizandrae Fructus, inhibited MUC5AC mucin production by the same inducers. These results suggest that baicalin, baicalein and schizandrin can regulate the production of mucin protein by directly acting on human airway epithelial cells.