Biomedical subjects

Ja Seok Koo

Publications and source records attributed to Ja Seok Koo.

13 recordsLinked to original sources

PubMed2006

Up-regulation of S100A8 and S100A9 protein in bronchial epithelial cells by lipopolysaccharide.

Increased serum levels of the S100A8 (MRP-8) protein have been reported in inflammatory conditions including bacterial infection, arthritis, and cystic fibrosis (CF). This protein is expressed constitutively with S100A9 (MRP-14) in neutrophils and is regulated by inflammatory stimulants. It has been hypothesized that increased inflammatory response to persistent bacterial infection is a major feature of CF lung disease. Therefore, the authors wished to determine the involvement of these two proteins in the innate defense response of the bronchial epithelium to lipopolysaccharide (LPS). Human bronchial epithelial cells (16HBE14o-) and primary bronchial epithelial cells (NHBE) were grown at air-liquid interface (ALI) and stimulated for up to 96 hours with LPS from Pseudomonas aeruginosa. The 16HBE14o- cells responded to LPS with a 2.9-fold increase in S100A8 mRNA production after 12 hours. S100A9 mRNA production was increased by 1.8-fold after 12 hours and 2.9-fold after 24 hours. It was also found that the S100A8 and S100A9 proteins were increased in the secretions of the 16HBE14o- and NHBE cells after LPS stimulation. This finding suggests that S100A8 and S100A9 are involved in the innate defense of the bronchial epithelium.

Bronchi↗

PubMed2005

Nonclassical action of retinoic acid on the activation of the cAMP response element-binding protein in normal human bronchial epithelial cells.

Vitamin A (retinol) is essential for normal regulation of cell growth and differentiation. We have shown that the retinol metabolite retinoic acid (RA) induces mucous cell differentiation of normal human tracheobronchial epithelial (NHTBE) cells. However, early biological effects of RA in the differentiation of bronchial epithelia are largely unknown. Here, we showed that RA rapidly activated cAMP response element-binding protein (CREB). However, RA did not use the conventional retinoic acid receptor (RAR)/retinoid X receptor (RXR) to activate CREB. RA activated CREB in NHTBE and H1734 cells in which RARs/RXR were silenced with small interfering RNA (siRNA) targeting RAR/RXR expression or deactivated by antagonist. Inhibition of protein kinase C (PKC) or extracellular regulated kinase (ERK1/2) blocked the RA-mediated activation of CREB. In addition, depletion of p90 ribosomal S6 kinase (RSK) via siRSK1/2 completely abolished the activation, suggesting that PKC, ERK, and RSK are required for the activation. Altogether, this study provides the first evidence that RA rapidly activates CREB transcription factor via PKC, ERK, and RSK in a retinoid receptor-independent manner in normal bronchial epithelial cells. This noncanonical RA signaling pathway may play an important role in mediating early biological effects in the mucociliary differentiation of bronchial epithelia.

Bronchi↗

PubMed2005

Bile acids induce MUC2 overexpression in human colon carcinoma cells.

BACKGROUND: Mucin alterations are a common feature of colonic neoplasia, and alterations in MUC2 mucin have been associated with tumor progression in the colon. Bile acids have been linked to colorectal carcinogenesis and mucin secretion, but their effects on mucin gene expression in human colon carcinoma cells is unknown METHODS: Human colon carcinoma cells were treated </= 6 hours with 10-200 microM deoxycholate, chenodeoxycholate, or ursodeoxycholate. MUC2 protein was assayed by Western blot analysis and MUC2 transcription was assayed using a MUC2 promoter reporter luciferase construct. Transcription activator protein 1 (AP-1) activity was measured using an AP-1 reporter construct and confirmed by Western blot analysis for c-Jun/AP-1. RESULTS: MUC2 transcription and MUC2 protein expression were increased three to fourfold by bile acids in a time and dose-dependent manner with no effect on cell viability. AP-1 activity was also increased (deoxycholate > chenodeoxycholate > ursodeoxycholate). Treatment with the putative chemopreventive agent curcumin, which decreased AP-1 activity, also decreased MUC2 transcription. Cotransfection with a dominant negative AP-1 vector decreased MUC2 transcription, confirming the significance of AP-1 in MUC2 induction by deoxycholate. Calphostin C, a specific inhibitor of protein kinase C (PKC), greatly decreased bile acid-induced MUC2 transcription and AP-1 activity, whereas inhibitors of MAP kinase had no effect. CONCLUSIONS: Bile acids induced mucin expression in human colon carcinoma cells by increasing MUC2 transcription through a process involving MAP kinase-independent, PKC-dependent activation of AP-1.

Antineoplastic Agents↗

PubMed2005

Galectin-3 modulates MUC2 mucin expression in human colon cancer cells at the level of transcription via AP-1 activation.

BACKGROUND & AIMS: Galectin-3 and MUC2 intestinal mucin each have been correlated with the malignant behavior of colon cancer cells. Galectin-3 modulates expression of MUC2 protein, but the specific regulatory mechanisms are unknown. This study sought to determine how galectin-3 increases MUC2 expression. METHODS: Galectin-3 levels in human colon cancer cells of high and low metastatic ability were manipulated via expression of galectin-3 complementary DNA in sense or antisense orientation. Galectin-3 and MUC2 protein expression were determined by Western analysis and immunocytochemistry. Transient transfections of promoter reporter constructs were used to monitor MUC2 transcription and AP-1 activity. Electrophoretic mobility shift assays, site-directed mutagenesis, and chromatin immunoprecipitation were used to monitor the participation of AP-1 in MUC2 transcription. RESULTS: Alterations in galectin-3 levels correlated with both MUC2 protein expression and transcription. By using MUC2 promoter constructs of different lengths, galectin-3 responsiveness was found between 1500 and 2186 bp upstream of the translation start site, a region that contains 1 consensus AP-1 binding site. AP-1 activity paralleled MUC2 transcription in the different cell lines. Mutation in the AP-1 site markedly decreased MUC2 promoter activity, and MUC2 transcription was inhibited by cotransfection with a dominant-negative AP-1 vector. Electrophoretic mobility shift assays, co-immunoprecipitation, and chromatin immunoprecipitation analyses suggested an association between galectin-3, c-Jun, and Fra-1 in forming a complex at the AP-1 site on the MUC2 promoter. CONCLUSIONS: Galectin-3 up-regulation of MUC2 transcription occurs at the level of transcription through AP-1 activation. This may have important implications for understanding the role of galectin-3 and MUC2 in colon cancer metastasis.

Cell Line, Tumor↗

PubMed2004

Prostaglandin E2 induces MUC8 gene expression via a mechanism involving ERK MAPK/RSK1/cAMP response element binding protein activation in human airway epithelial cells.

MUC8 gene expression is overexpressed in nasal polyp epithelium and is also increased by treatment with inflammatory mediators in nasal epithelial cells. These data suggest that MUC8 may be one of important mucin genes expressed in human airway. However, the mechanisms of various inflammatory mediator-induced MUC8 gene expression in normal nasal epithelial cells remain unclear. We examined the mechanism by which prostaglandin E(2) (PGE2), an arachidonic acid metabolite, increases MUC8 gene expression levels. Here, we show that ERK mitogen-activated protein kinase is essential for PGE2-induced MUC8 gene expression in normal human nasal epithelial cells and that p90 ribosomal S 6 protein kinase 1 (RSK1) mediates the PGE2-induced phosphorylation of cAMP-response element binding protein. Our results also indicate that cAMP-response element at the -803 region of the MUC8 promoter is an important site of PGE2-induced MUC8 gene expression. In conclusion, this study gives insights into the molecular mechanism of PGE2-induced MUC8 gene expression in human airway epithelial cells.

Cell Line, Tumor↗

PubMed2004

A comparative analysis of data generated using two different target preparation methods for hybridization to high-density oligonucleotide microarrays.

BACKGROUND: To generate specific transcript profiles, one must isolate homogenous cell populations using techniques that often yield small amounts of RNA, requiring researchers to employ RNA amplification methods. The data generated by using these methods must be extensively evaluated to determine any technique dependent distortion of the expression profiles. RESULTS: High-density oligonucleotide microarrays were used to perform experiments for comparing data generated by using two protocols, an in vitro transcription (IVT) protocol that requires 5 microg of total RNA and a double in vitro transcription (dIVT) protocol that requires 200 ng of total RNA for target preparation from RNA samples extracted from a normal and a cancer cell line. In both cell lines, about 10% more genes were detected with IVT than with dIVT. Genes were filtered to exclude those that were undetected on all arrays. Hierarchical clustering using the 9,482 genes that passed the filter showed that the variation attributable to biological differences between samples was greater than that introduced by differences in the protocols. We analyzed the behavior of these genes separately for each protocol by using a statistical model to estimate the posterior probability of various levels of fold change. At each level, more differentially expressed genes were detected with IVT than with dIVT. When we checked for genes that had a posterior probability greater than 99% of fold change greater than 2, in data generated by IVT but not dIVT, more than 60% of these genes had posterior probabilities greater than 90% in data generated by dIVT. Both protocols identified the same functional gene categories to be differentially expressed. Differential expression of selected genes was confirmed using quantitative real-time PCR. CONCLUSION: Using nanogram quantities on total RNA, the usage of dIVT protocol identified differentially expressed genes and functional categories consistent with those detected by the IVT protocol. There was a loss in sensitivity of about 10% when detecting differentially expressed genes using the dIVT protocol. However, the lower amount of RNA required for this protocol, as compared to the IVT protocol, renders this methodology a highly desirable one for biological systems where sample amounts are limiting.

Algorithms↗

PubMed2003

Induction of MUC8 gene expression by interleukin-1 beta is mediated by a sequential ERK MAPK/RSK1/CREB cascade pathway in human airway epithelial cells.

Mucins are the major components of the mucus layer that covers and protects the respiratory, digestive, and reproductive tracts. Our previous studies showed that MUC8 gene expression was overexpressed in in vivo polyp epithelium in chronic sinusitis and was also increased by treatment with inflammatory mediators in an in vitro culture condition. However, the mechanisms by which the inflammatory mediators-induced MUC8 gene expression in normal nasal epithelial cells evolved remain unclear. We examined the mechanism by which the important proinflammatory mediator, interleukin (IL)-1 beta, increases MUC8 gene expression levels. We found that pharmacologic and genetic inhibition of ERK MAPK pathway abolished IL-1 beta-induced MUC8 gene expression in normal human nasal epithelial cells. Moreover, the overexpression of wide-type or of the dominant-negative mutant of p90 ribosomal S6 protein kinase 1 (RSK1) enhanced or suppressed, respectively, IL-1 beta-induced MUC8 gene expression. RSK1 was found to directly phosphorylate cAMP-response element-binding protein (CREB), and this event led to the stimulation of subsequent CRE-mediated gene transcription. In conclusion, IL-1 beta was found to induce MUC8 gene expression via a sequential ERK/RSK1/CREB pathway in human airway epithelial cells.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2003

Interleukin-1 beta and tumor necrosis factor-alpha induce MUC5AC overexpression through a mechanism involving ERK/p38 mitogen-activated protein kinases-MSK1-CREB activation in human airway epithelial cells.

Mucin hypersecretion is commonly observed in many inflammatory diseases of the respiratory tract. MUC5AC is generally recognized to be a major airway mucin because MUC5AC is highly expressed in the goblet cells of human airway epithelium. Moreover, it is regulated by various inflammatory cytokines. However, the mechanisms by which the interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha induce MUC5AC gene expression in normal nasal epithelial cells, and the signal molecules involved, especially in the downstream signaling of mitogen-activated protein (MAP) kinases, remain unclear. Here we show that pharmacologic or genetic inhibition of either ERK or p38 MAP kinase pathway abolished IL-1beta- and TNF-alpha-induced MUC5AC gene expression in normal human nasal epithelial cells. Our results also indicate that the activation of mitogen- and stress-activated protein kinase 1 (MSK1) and cAMP-response element-binding protein and cAMP-response element signaling cascades via ERK and p38 MAP kinases are crucial aspects of the intracellular mechanisms that mediate MUC5AC gene expression. Taken together, these studies give additional insights into the molecular mechanism of IL-1beta- and TNF-alpha-induced MUC5AC gene expression and enhance our understanding on mucin hypersecretion during inflammation.

Cell Line↗

PubMed2003

Proteomic and transcriptomic analysis of interleukin-1beta treated lung carcinoma cell line.

Mucin hypersecretion is one of the main symptoms of inflammatory disease in the respiratory tract. We previously reported that the pleiotypic pro-inflammatory cytokine, interleukin (IL)-1beta, plays a significant role in respiratory tract inflammation by inducing mucins. However, the molecular mechanism for mucin hypersecretion in the respiratory tract remains still unclear. In order to understand the mechanisms of mucin hypersecretion in the airway epithelium, the differentially expressed proteins and genes in the lung mucoepidermoid carcinoma cell line (NCI-H292 cells), which were treated for 6 and 24 hours with IL-1beta (10 ng/mL) were identified using two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) proteomics and cDNA microarray analysis (8.6K). In the 2-D PAGE, eight differentially expressed proteins and 14 post-translational modification proteins were identified at 6 and 24 hours after the IL-1beta-treatment. Four hundred and thirteen genes (6.6%) and 115 genes (2.0%) were differentially expressed, respectively, at 6 and 24 hours after the IL-1beta-treatment by microarray analysis. The differentially expressed genes and proteins that were regulated by the IL-1beta-treatment were mostly in the metabolic pathway rather than in the regulatory pathway. These results clearly show that the transcript levels have little value in predicting the extent of protein expression.

Carcinoma↗

PubMed2003

Differential regulation of nonsteroidal anti-inflammatory drug-activated gene in normal human tracheobronchial epithelial and lung carcinoma cells by retinoids.

In this study, we analyze the effect of several retinoids on the expression of nonsteroidal anti-inflammatory drug-activated gene (NAG-1) in normal human tracheobronchial epithelial (HTBE) cells and several lung carcinoma cell lines. The retinoid 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (AHPN) greatly enhances the expression of NAG-1 mRNA and protein in a time- and dose-dependent manner in human lung adenocarcinoma H460 cells and several other carcinoma cell lines. This induction was specific for AHPN because retinoic acid, a retinoic acid receptor-, and a retinoid X receptor pan-agonist were unable to induce NAG-1, suggesting that this induction is not mediated through activation of retinoid receptors. Although NAG-1 is a p53-responsive gene, AHPN-induced NAG-1 expression does not require p53. The induction of NAG-1 expression by AHPN is caused at least in part by an 8-fold increase in the stability of NAG-1 mRNA. In contrast to carcinoma cells, NAG-1 expression is effectively induced by retinoic acid and the RAR-selective pan-agonist in normal HTBE cells and accompanies the inhibition of squamous differentiation and the initiation of normal differentiation. In vivo, NAG-1 expression was observed in the normal tracheobronchial epithelium, whereas no expression was found in either squamous metaplastic tracheal epithelium or in sections of human lung tumors. Our results suggest that the induction of NAG-1 expression by retinoids in normal HTBE and lung carcinoma cells is regulated by distinct mechanisms and is associated with different biological processes. The linkage between AHPN treatment and NAG-1 expression revealed in this study provides a new mechanism for the antitumorigenic activity of AHPN.

Adenocarcinoma↗

PubMed2002

IL-13 suppresses MUC5AC gene expression and mucin secretion in nasal epithelial cells.

Recently, it was found that IL-13 is involved in allergic inflammation and mucus hypersecretion in in vivo animal models. However, the role of IL-13 in in vitro cellular models has yet to be determined. This study aimed to investigate the direct effect of IL-13 on mucin gene expression and mucin secretion in cultured normal human nasal epithelial cells. After treatment with IL-13, total mucin and MUC5AC mucin levels were measured using an immunoblotting assay. MUC2, MUC5AC and MUC8 mRNA expression were determined using reverse transcription-polymerase chain reaction. Also, immunostaining was performed using a MUC5AC antibody on histologic and cytospin slides. After treatment with > or = 5 ng/ml IL-13, the level of total mucin and MUC5AC mucin secretion decreased substantially. The expression of MUC2 and MUC8 mRNA increased with higher concentrations of IL-13, but the expression of MUC5AC mRNA decreased. On the 7th day after IL-13 treatment, a significant decrease in the number of MUC5AC-positive cells was confirmed by immunostaining. These findings indicate that IL-13 suppresses MUC5AC mucin gene expression and mucin secretion in cultured normal human nasal epithelial cells in vitro.

Cell Count↗

PubMed2002

Upregulation of MUC8 and downregulation of MUC5AC by inflammatory mediators in human nasal polyps and cultured nasal epithelium.

Polyps are believed to be the source of mucus hypersecretion in chronic inflammation of the sinus. However, it is not clear which mucins are responsible for the hypersecretion of mucus by nasal polyps. We describe the over-expression of MUC8 mRNA in nasal polyps and the upregulation of MUC8 mRNA expression and downregulation of MUC5AC mRNA expression by inflammatory mediators. We found that the level of MUC8 mRNA, but not the level of MUC5AC mRNA, increased in nasal polyps. We also found that there was an increase in intracellular mucin in nasal polyps, compared to the normal nasal inferior turbinate. A mixture of inflammatory mediators increased MUC8 mRNA expression and decreased MUC5AC mRNA expression in cultured normal human nasal epithelial cells. Among inflammatory mediators, IL-4 is responsible for the decrease in MUC5AC mRNA and MUC5AC mucin secretion. These results indicate that MUC8 may be one of the major mucins secreted from the polyp epithelium and that it may play an important role in the pathogenesis of mucus hypersecretion in chronic sinusitis with polyps.

Cells, Cultured↗

PubMed2002

Overexpression of mucin genes induced by interleukin-1 beta, tumor necrosis factor-alpha, lipopolysaccharide, and neutrophil elastase is inhibited by a retinoic acid receptor alpha antagonist.

Proinflammatory cytokines, lipopolysaccharide (LPS), and neutrophil elastase (NE) have been implicated in the induction of hypersecretion of respiratory mucus. In this study, we demonstrated that interleukin-1beta (IL-1beta) increased MUC2 and MUC5AC mRNA levels 2- to 3-fold in a time- and dose-dependent manner in NCI-H292 cells. In contrast, MUC5B mRNA was not significantly changed. A transcription inhibitor blocked the stimulation of MUC2 and MUC5AC gene expression by IL-1beta. A translation inhibitor did not interfere with the induction of MUC2 mRNA expression, whereas stimulation of MUC5AC mRNA was blocked, suggesting de novo protein synthesis is required for the stimulation of MUC5AC mRNA. We previously reported that induction of MUC2, MUC5AC, and MUC5B gene expressions by retinoic acid is mediated by the retinoic acid receptor (RARalpha), and inhibited by the specific RARalpha antagonist Ro 41-5253. Here, we demonstrate that the RARalpha antagonist can effectively inhibit IL-1beta-induced MUC2 and MUC5AC gene expression and reduce intracellular MUC5AC protein. Further investigation showed that the RARalpha antagonist also inhibited the stimulation of MUC2 and MUC5AC mRNA expression by tumor necrosis factor-alpha, LPS, and NE.

Benzoates↗