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Background and Aim: TFF1 (pS2) is expressed at a high level in gastric epithelial cells and plays an important role in protecting the gastric mucosa. However, the regulatory mechanisms of TFF1 expression are not fully understood. The aim of this study was to investigate the effect of TNF-alpha, a representative proinflammatory cytokine, on TFF1 expression., Methods: MKN45 and AGS cells, derived from human gastric carcinoma, were used. Endogenous TFF1 mRNA expression was analyzed by real-time quantitative RT-PCR. The sequences of the human TFF1 promoter were cloned into the pGL3-basic vector and reporter gene assays were performed. Nuclear factor (NF)-kappaB activity was monitored using a reporter vector that contained multiple copies of NF-kappaB responsive element upstream of the luciferase gene. Interaction between NF-kappaB and TFF1 cis-element was examined by electophoretic mobility shift assay (EMSA)., Results: TNF-alpha activated NF-kappaB and up-regulated endogenous TFF1 mRNA expression as well as the transcription of the TFF1 reporter genes in a dose-dependent manner. IL-1beta, another proinflammatory cytokine, also up-regulated TFF1 expression. TNF-alpha responsive element was mapped between -342 and -147 of the human TFF1 promoter and a putative NF-kappaB binding site was identified at -231. When this element was deleted, the reporter genes became almost insensitive to TNF-alpha treatment. EMSA showed binding of NF-kappaB to this element., Conclusions: Inflammatory stimuli that activate NF-kappaB appear to up-regulate TFF1 expression in gastric epithelial cells. This mechanism may aid in the protection of the gastric mucosa under inflammatory conditions.

Trefoil factor family (TFF) is a group of peptides that play critical roles in maintaining gastric mucosal integrity. In real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) and reporter gene assays, we show that indomethacin and aspirin upregulate TFF2 expression in MKN45 gastric cells. These drugs also activated peroxisome proliferator-activated receptor gamma (PPARgamma) at concentration ranges that increase TFF2 expression, and upregulated TFF2 expression was suppressed by GW9662, a specific inhibitor of PPARgamma. These results suggest that indomethacin and aspirin upregulate gastric expression of TFF2 through activation of PPARgamma. This mechanism may be important in reducing the extent of gastric mucosal injury caused by the administration of non-steroidal anti-inflammatory drugs (NSAIDs).

Reactive oxygen species appears to be involved in the pathogenesis of ethanol-induced gastric mucosal injury in vivo. Because ingested ethanol diffuses into the gastric mucosa, targeting both epithelium and endothelium, in the present study we examined the possible protective effect of antioxidants on ethanol damage in gastric epithelial cells and endothelial cells in vitro. Cytotoxicity by ethanol was quantified by measuring 51Cr release. The effects of impairment of the glutathione redox cycle and of inhibition of cellular catalase were examined. The generation of superoxide was assessed by the reduction in cytochrome c. Ethanol caused a time- and dose-dependent increase in 51Cr release from epithelial cells. Incubation of cells with DL-buthionine-(S,R)-sulfoximine, while reducing glutathione production, dose dependently enhanced ethanol-induced injury. 1,3-Bis(chloroethyl)-nitrosourea, while inhibiting glutathione reductase activity, also sensitized cells to ethanol. In contrast, the inhibition of catalase with 3-amino-1,2, 4-triazole did not alter the susceptibility of epithelial cells to ethanol. Ethanol induced damage to endothelial cells in a similar fashion. In endothelial cells, however, neither impairment of the glutathione cycle nor inhibition of catalase influenced ethanol-induced damage. Epithelial cells, when exposed to ethanol, increased superoxide production as a function of ethanol concentration, whereas endothelial cells did not. The glutathione redox cycle, but not cellular catalase, plays a critical role in protecting epithelial cells against ethanol damage, whereas neither antioxidant seems to play a role in protection of endothelial cells. The distinct difference in antioxidant protection against ethanol appears to depend on the capability of each cell to produce cytotoxic oxygen species in response to ethanol exposure.

Trefoil factor family (TFF) is a group of small peptides secreted by gastrointestinal epithelial cells. Among three known TFF peptides, TFF1 (formerly pS2) is expressed at a high level in gastric epithelial cells and plays an essential and critical role in maintaining the integrity of the gastric mucosa. Recent evidence also suggests that TFF1 acts as a tumour suppressor gene in the stomach. TFF1 was originally discovered as an oestrogen-inducible gene in MCF-7 breast cancer cells, and its expression is dependent on oestrogen in MCF-7 and other hormone-dependent breast cancer cells. Although gastric epithelial cells express oestrogen receptors (ERs), gastric TFF1 expression appears to be independent of oestrogen signalling. Instead, several cis-regulatory elements are involved in the regulation of gastric TFF1 expression and balanced signalling from gp130, a common IL-6 family coreceptor, has been shown to be necessary for the proper expression of TFF1 in the stomach. Epigenetic regulation, such as DNA methylation in the promoter region of the TFF1 gene, may be also important for tissue-specific expression of TFF1 in the stomach. However, further studies are still needed to fully understand the detailed regulatory mechanisms of TFF1 expression in gastric epithelial cells. [ABSTRACT FROM AUTHOR]

: Trefoil factor family peptides are expressed in gastrointestinal epithelial cells and play a critical role in maintaining mucosal integrity. Although non-steroidal anti-inflammatory drugs (NSAIDs) are important causative agents of gastric mucosal lesions, few data are available about the effect of NSAIDs on trefoil family peptides in gastric mucosa. : To examine whether indometacin, a widely used NSAID, affects trefoil factor family expression in gastric epithelial cells. : MKN45, a cell line derived from human gastric cancer, was used. TFF1, TFF2, and TFF3 mRNA expression was assessed by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR). TFF2 gene transcription was also examined by luciferase reporter gene assay. : Relative expression level of TFF1, TFF2, TFF3 mRNA was 616: 12: 1 in unstimulated MKN45 cells. Although indometacin (1–250 µmol / L) had no significant effect on the expression of TFF1 and TFF3 mRNA, it up-regulated TFF2 mRNA expression in a dose- and time-dependent manner. Luciferase reporter gene assay confirmed the up-regulation of TFF2 gene transcription by indometacin. Indometacin-induced up-regulation of TFF2 expression was not antagonized by externally applied prostaglandin E2. : These results suggest that indometacin up-regulates gastric epithelial cell TFF2 expression through a COX-independent mechanism. Since TFF peptides play an important role in gastric mucosal protection, indometacin-induced TFF2 may reduce the degree of gastric mucosal damage induced by indometacin. [ABSTRACT FROM AUTHOR]

Summary Background : Although trefoil factor family peptides (TFF peptides) are assumed to play important roles in gastric mucosal protection, the regulatory mechanism of gastric TFF expression has not been fully understood yet. Recent reports showed gastric expression of peroxisome proliferator-activated receptor γ (PPARγ), a nuclear receptor known to be involved in the regulation of cell growth and differentiation in other cell types, such as adipocytes. Aim : To determine whether PPARγ affects the expression of TFF in gastric epithelial cells. Methods : MKN45 gastric cells were used as a model of gastric epithelial cells. DNA synthesis of the cells was determined by the measurement of BrdU incorporation. The effects of PPARγ ligands, 15-deoxy-Δ[sup 12,14] -prostaglandin J[sub 2] (15d-PGJ[sub 2] ) and troglitazone (TGZ) on TFF expression were assessed by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR). Results : MKN45 cells expressed a significant amount of PPARγ. Both 15d-PGJ[sub 2] and TGZ suppressed DNA synthesis of the cells in a dose-dependent manner. In the control condition, MKN45 cells most abundantly expressed TFF1 and the relative expression level of TFF1, TFF2, and TFF3 mRNA was 1700:32:1. TFF1 and TFF2 mRNA levels were significantly up-regulated by the incubation of the cells with 15d-PGJ[sub 2] (10 µm) or TGZ (30 µm), whereas TFF3 mRNA level was not affected. Conclusion : The results of the present study suggest a possible role of PPARγ in the regulation of TFF expression in gastric epithelial cells. [ABSTRACT FROM AUTHOR]

Backgournd: Peroxisome proliferator-activated receptors (PPAR) are a family of three nuclear receptors (PPARα, PPARδ, and PPARγ). Although recent evidence suggests a role for PPARγ in the regulation of colonic epithelial cell growth, the role for PPARγ in the stomach has not been established. Aim: To examine the expression of PPARγ and the effects of PPARγ ligands on the viability of gastric epithelial cells. Methods: MKN45 cells and primary cultured rat gastric epithelial cells were used. Troglitazone (TGZ) and 15-deoxy-Δ[sup 12, 14 ] -prostaglandin J[sub 2] (15d-PGJ[sub 2] ) were used as PPARγ ligands. Expression of PPARγ was examined by RT–PCR and Western blot analysis. Cell viability was measured by WST-1 assay and TUNEL assay was performed to detect apoptosis. Results: MKN45 cells expressed all subtypes of PPAR. PPARγ ligands decreased cell viability and induced cell death in a dose-dependent manner, whereas ligands for PPARα and PPARδ had no significant effect. TUNEL assay showed that this cell death is apoptosis. Primary cultured rat gastric epithelial cells also expressed PPARγ and activation of PPARγ decreased cell viability. Conclusion: These results suggest that PPARγ plays an important role in the regulation of cell growth and cell death in gastric epithelial cells. [ABSTRACT FROM AUTHOR]

Many cytokines are involved in the repair of damaged tissue, and one of these, hepatocyte growth factor (HGF), is involved not only with liver regeneration but also in the repair of other tissues. To investigate the importance of HGF in the repair of the small intestine, we evaluated its effect and that of other growth factors in IEC-6 cells, an intestinal epithelial cell line derived from normal rat small intestine. Round "wounds" were made in confluent monolayers of IEC-6 by silicon rubber-tipped steel rods and various cytokines; transforming growth factor alpha (TGF-alpha), transforming growth factor beta1 (TGF-beta1), keratinocyte growth factor (KGF), and HGF, were added. We photographed the repaired monolayers every 24 h and calculated the ratios of areas not covered by cells to initial areas. Cell proliferation with TGF-alpha, TGF-beta, KGF, or HGF was examined in terms of [3H]-thymidine uptake. Finally, we determined c-met (the HGF receptor) mRNA in the IEC-6 cells by Northern blot hybridization. HGF was the most potent of the cytokines in accelerating repair of the damaged monolayer of IEC-6. HGF was also 1.34 times more effective than control the medium for inducing cell proliferation of IEC-6. By Northern blot hybridization, three bands of mRNA bound to c-met cDNA. These results suggest that HGF is important in the repair of the small intestine. [ABSTRACT FROM AUTHOR]